virology exam 3

retroviruses (family Retroviridae) such as HIV-1 are spherical w/ distinctive nucleocapsid and matrix layers inside an envelope with prominent spikes

(+) RNA has all the typical features of eukaryotic mRNA: 5' methylated cap and 3' poly-A tail; genomes have long 5' and 3' UTR

still working on universal IAV vaccine, problems isolating one part

(ex. HA spike stalk promising, but have problems w/ stalk's head)

eukaryotic cell cycle

**4 stages: M, G1, S, and G2; progression thru the cell cycle is controlled by 3 checkpoints where cells can exit the cell cycle and undergo apoptosis; differentiated cells are in G0, which branches off from G1; some cells in G0 can be stimulated by growth factors to re-enter G1 and begin cycling

overlapping coding sequences commonly used by viruses to encode less abundant proteins w/ regulatory or enzymatic functions

**HIV accessory proteins have low abundance compared w/ polyproteins

expression of immediate-early, early, and late proteins = linked to layer of epithelium occupied by infected host cell (as cell moves thru layers)

**HPV gene expression and genome replication are tied closely to the differentiation status of the host cells in a stratified epithelium

typical sites of assembly in eukaryotic viruses = cytoplasm, plasma membrane, nucleus

**different viruses assemble at various different subcellular locations

antigenic proteins of flu A virus

**hemagglutinin (HA) & neuraminidase (NA)

if DNA damage = normal, uninfected cells exit cell cycle and undergo apoptosis

**key protein regulator of apoptosis in response to DNA damage = p53; stress caused when a virus forces a cell into S phase resembles DNA damage stress-- can trigger the p53 system (both DNA damage and viral infections activate p53)

most recent innovation w/ phages treating bacterial infections = use of enzymes produced by phages as drugs (not entire phage), e.g.

**lysins = enzymes that degrade peptidoglycan; useful in killing gram-positive bacteria

early transcription proceeds unidirectionally, terminating approx. halfway around the circular genome

**major mechanism by which SV40 encodes so many different proteins w/ small amount of DNA = alternative splicing (3 different mature early mRNAs that encode either small t antigen, large T antigen, or the 17-kt protein)

large T antigen interacts with:

**pRB, preveting pRB from interacting with E2F transcription factors and thus derepressing S phase genes; **interacts with p53, which prevents p53 from stimulating apoptosis

virus budding = capsid (in the cytoplasm) remains interior to the membranous structure that buds away

**when a virus buds, the exterior surface of plasma membrane remains exterior, the interior contents of the budding virus, which are contiguous w/ the cytoplasm, remain on the interior of the cell

whole genome replication involves host Pol II transcribing entire viral genome into single (not spliced) mRNA in nucleus-

-cytoplasm--encapsulated for new virions

misincorporation is responsible for much more genetic change than physical & chemical assaults, but UV light still important source of mutation

1. UV light causes formation of covalent bonds between atoms of adjacent pyrimidines (bases t, c, & u); 2. disrupts normal hydrogen bonding between strands of DNA & replication of DNA or RNA; 3. same issues w/ human DNA & sunlight

retroviral integrase inserts the viral cDNA into a chromosome, forming proviral DNA that can be transcribed by host Pol II

1. after the reverse transcriptase reactions, HIV cDNA is in complex w/ virus & host proteins called preintegration complex

virology unites the biosphere

1. all cells evolved awash in viruses and subviral entities; 2. viruses are actually central component of biosphere; 3. discipline of virology reveals the fundamental connectedness of all nucleic acid-base life

antiviral drugs target proteins unique to viruses & essential for their replication cycle

1. antiviral drugs bind to and inhibit viral proteins in infected cells; 2. ideal situation = antiviral drug does not bind to any normal human proteins- in practice this goal is rarely achieved; 3. good antiviral drug binds to target virus protein more tightly than it binds to human proteins-- at levels of drug in body, most of drug binds to viral proteins; 4. viral proteins that play essential role at any point during replication are particularly good possible targets b/c inhibiting them slows viral replication

viral evolution occurs in response to selective pressure from antiviral drugs

1. antiviral drugs exert strong selective pressure on viruses-- result in emergence of viruses resistant to drug used to treat them; 2. resistant viruses usually have genetic variations--alter the amino acids that would otherwise be involved w/ target protein & drug

attenuated vaccines = highly immunogenic b/c they can still replicate

1. are 3 different types of vaccines: attenuated, inactivated, and subunit (most to least immunogenic)

inactivated vaccines = composed of nonreplicating virions

1. are safer than attenuated b/c virus in vaccine treated so unable to replicate; 2. inactivate virus by physical assaults (heat, radiation) or chemical treatment; 3. e.g. poliovirus treated w/ formaldehyde-- forges covalent bonds between capsomeres-- prevents virus from uncoating; 4. inactivated vaccine provides immune system w/ foreign antigens that can provoke specific response against the virus

medically dangerous emerging viruses are zoonotic

1. as human population increases--spread into new landscapes--puts humans in contact w/ many wild (undomesticated) animal viruses, some of which can jump species barrier; 2. in addition, human expansion also = contact between pets and domesticated farm animals w/ many wild animal viruses; 3. zoonotic diseases = typically reproduce in nonhuman animal (usually vertebrate) but can be transmitted to humans

eukaryotic virus assembly must take cellular protein localization into account

1. b/c eukaryotic viruses assemble at different subcellular locations, protein localization is very important aspect of viral biology; 2. synthesis and trafficking of cytoplasmic proteins, nuclear proteins, & transmembrane (across membranes) proteins are very important; 3. cytoplasm of a eukaryotic cell is very crowded and diffusion is much slower than in water; 4. diffusion is too slow for period of time needed for the completion of replication cycle; 5. therefore, viruses rely on the cytoskeleton (including microtubules) and motor proteins; 6. pharmaceuticals or other treatments that disrupt microtubules or microtubule-dependent motors interfere w/ viral assembly

dna viruses can cause productive lytic infections, cellular transformation, or latent infections

1. baltimore class I viruses 2. infection by most groups of dsDNA viruses can result in 3 distinct outcomes: a. lytic infection (productive infection) b. cellular transformation (host cell begins to be transformed into a cancer cell; cell becomes less responsive to physiological cues meant to regulate cell growth, differentiation, and death; can be dead end for virus, if no progeny virions produced) c. latent infection

repeated elements in any promoter region usually sites for protein binding

1. bind to ubiquitous host transcription factors such as SP1-- binds to the GC-rich sequences in 21-bp repeat; 2. 72-bp repeat functions as binding site for host proteins that enhance transcription

eukaryotic capping, splicing, and polyadenylation occur co-transcriptionally

1. during transcription elongation, Pol II is in a complex w/ capping, splicing, & polyadenylation enzymes 2. eukaryotic primary transcripts contain introns that must be spliced out for mRNA to encode protein-- splicing occurs at same time as transcription

**only enveloped viruses use budding

1. enveloped viruses exit host cell by pinching off; 2. host cell may remain alive for time until energetic costs of viral replication take their toll & cell dies despite not being lysed

viruses acquire envelopes thru one of two pathways:

1. enveloped viruses have more complex assembly than naked viruses for 2 reasons: a. they express both soluble cytoplasmic proteins and transmembrane proteins, which are translated at different sites, but must come together; b. they acquire their membranous envelope at a site distinct from either the nucleus or the cytoplasm, where new genomes accumulate

eukaryotic transcription is affected by the state of the chromatin

1. eukaryotic DNA is packed around nucleosomes forming chromatin-- when condensed, can interfere w/ transcription

unlike cells, viruses assemble from their constituent parts

1. every living cell comes from a preexisting parental cell (from division of a parental cell); 2. viruses arise by assembly, not by division of parental viruses; 3. x-ray crystallographer, Rosalind Franklin, was instrumental in determining that tobacco mosaic virus virions are composed of 2,310 identical capsomers that are arranged in a helical pattern surrounding a single genomic RNA molecule

new pandemic flu A strains may be able to arise thru mutation

1. gain of function studies have shown mutation can alter H5N1 HA protein so virus can attach to human cells more efficiently; a. could allow transmission directly from birds to humans; b. could make human-to-human transmission easier; 2. 2013 outbreak of avian H5N1 flu among Cambodians, researchers found potentially dangerous sequence variations that arose thru mutations of the viruses while they replicated in infected people

most baltimore class I viruses including polyomaviruses manipulate the eukaryotic cell cycle

1. generally, genome replication among class I dsDNA viruses, relies heavily on eukaryotic cell cycle; 2. host DNA replication proteins present exclusively during S phase; lasts about 1/3 of lifetime of actively cycling cell

subunit vaccines are composed of selected analgesic proteins

1. have only some of the component parts of the infectious microbe; 2. current inactivated vaccines in use for hep. B and seasonal flu

compact retroviral genome is used economically to encode many proteins through the use of polyproteins, alternative splicing, and translation of polycistronic mRNA

1. have small genomes--need strategies to encode more than one protein using same sequence

hepadnaviruses are enveloped and have genomes containing both DNA and RNA in an unusual arrangement

1. includes hepatitis B (HBV); 2. virions = spherical w/ icosahedral capsids, enveloped; envelope has 3 different types of spike proteins

lytic herpesvirus replication involves a cascade with several waves of gene expression

1. includes herpes simplex virus 1, varicella-zoster virus (human herpesvirus 3), epstein-barr virus (human herpesvirus 4), and kaposi's sarcoma virus (human herpesvirus 8); **can cause latent infections

adenoviruses are large dsDNA viruses w/ 3 waves of gene expression

1. infect respiratory organs, eyes, kidneys, gastrointestinal tract, and blood cells; 2. can cause acute bronchitis, pneumonia, pink eye, common cold (mostly rhinoviruses), and others; 3. linear dsDNA genomes, 4-8 times longer than small polyomaviruses and papillomaviruses dsDNA viruses; 4. after entry into its host cell, virus is progressively disassembled as it travels to nucleus--uncoating introduces viral genomic DNA into nucleus

lysis

1. is an option regardless of virus structure & enveloped or non-enveloped; 2. all assembled virions released in one large burst; 3. host cell is killed; ultimately explodes b/c of osmotic pressure

some viruses require maturation reactions during release in order to form infectious virions

1. maturation = any irreversible process that occurs during or after viral release that takes assembled virus from noninfectious state to infectious one

release from bacterial cells usually occurs by lysis

1. most living things, including bacteria, plants, & some protists, have rigid cell walls covering their plasma membranes (PM); a. cell walls squeeze back against osmotic pressure-- protects PM from osmotic lysis; 1. cell in hypotonic environment (solutes outside < solutes inside), water enters cell-- PM can lyse w/o cell wall or other mechanism; b. walls also provide protection against viruses except where viruses have evolved mechanisms to bypass cell wall during penetration & release of offspring virions

release from animal cells can occur by lysis

1. no cell wall to obstruct virus egress, instead, cytoskeleton is key for maintaining cell integrity; 2. viruses that exit thru lysis must disrupt cytoskeleton; 3. late protein (a protease) cleaves the intermediate filament cytokeratin-- intermediate filament no longer able to assemble properly; a. effect = deprives cell of mechanical integrity and makes it much more susceptible to lysis

capsids & nucleocapsids associate with genomes using one of two general strategies

1. once capsomers and genomes arrive at same location within cell, still must assemble

pandemic flu A strains have arisen thru recombination

1. origins of flu A strains that caused pandemics since 1918 have been studied; use approaches that rely on genome sequencing and bioinformatics; 2. viral genomes recovered from tissues of flu victims from 1918-1920 & samples preserved from all subsequent pandemics; 3. growing libraries of swine, domestic bird, and migratory bird samples for comparison; 4. where evolutionary history could be reconstructed, all pandemics caused by antigenic shift when avian influenza recombined w/ other strains of flu

release from plant cells often occurs thru biting arthropods

1. plant viruses must surmount thick cell wall when they exit cell

e. coli cell envelope

1. pm, then periplasm filled w/ loose network of peptidoglycan, & then 2nd lipid bilayer; 2. peptidoglycan = gel-like, v. large molecule, surrounds cell in 3 dimensions like a net-- protects the cell from osmotic lysis

polyomaviruses are small DNA viruses with early and late gene expression

1. polyomaviruses are models for viruses w/ some of shortest known genomes to replicate in animal cells; 2. associated with post-organ kidney transplant malfunction and w/ Merkel cell carcinoma; 3. naked spherical virions-- only 50 nm in diameter; 4. small genome (4-5 kbp) composed of circular dsDNA packaged into nucleosomes; 5. after attachment & penetration, uncoating results in introduction of viral genome w/ its 24 incomplete nucleosomes into nucleus; 6. after nuclear histone H1 joins the viral dsDNA genome-- nucleosomes are same host's chromatin-- viral genome is ready for transcription

poxviruses are extremely large dsDNA viruses that replicate in host cytoplasm

1. poxviruses are largest dsDNA viruses that infect humans (both physical size of virions & length of genomes)

hepadnaviruses use reverse transcription to amplify their genomes

1. replication cycle has many steps associated with gene expression & genome replication-- occur in nucleus

SV40 genome replication requires viral and host proteins to form active DNA replication forks

1. replication of SV40 DNA is arguably the best understood eukaryotic replicon to date; 2. many aspects of SV40 genome replication are similar to those of host DNA replication; unique aspects of viral replication involve large T antigen: a. regulatory roles b. direct enzymatic role during DNA replication

virions more structurally complex than TMV also reproduce by assembly, not by division

1. replication thru self-assembly of component parts makes viruses radically different from host cells; 2. other research provides evidence that more complex viruses also reproduce by assembly

metagenomics will revolutionize evolutionary understanding of viruses

1. results in discovery of 1000s of new viral genomes every year--pace of discovery is accelerating

retroviruses-- viral reverse transcriptases have polymerase and RNase H activity

1. retroviruses have genome of (+) ssRNA; 2. cause HIV among other diseases; 3. viral reverse transcriptase enzymes use an RNA template to synthesize DNA (reverse transcription)-- use DNA template to synthesize DNA (DNA polymerase enzyme)

existing drugs not very effective

1. seasonal flu A viruses already resistant to M2 protein inhibitors (no longer bind to M2 proteins); 2. NA inhibitors may shorten symptoms by one day if taken early during infection, but may not reduce hospitalizations or deaths; **new anti-flu drugs targeting viral enzymes such as RdRp are in development

papillomavirus replication cycle is tied closely to the differentiation status of its host cell

1. small circular DNA viruses; 2. important causes of human disease: a. lytic replication cycle causes warts, including genital warts, b. human papillomaviruses are leading cause of cervical cancer; 3. vaccines now prevent most common forms of cervical cancer by immunizing the population, including males, against HPV

gene therapy = engineered viruses could be used (in principle) to treat cancer and other conditions

1. some diseases caused by inheritance of faulty alleles that encode nonfunctional or dysfunctional proteins/mRNAs; 2. in principle, should be possible to treat genetic diseases thru gene therapy, by providing functional wild-type alleles to cells that need it; 3. b/c viruses are experts at binding to and introducing genes into host cells, are ideal vectors (carriers) for selective delivery of therapeutic transgenes; 4. if therapeutic transgenes could alter the genetic code of stem cells, then single treatment could in principle cure inherited disorder by providing functional gene expressed by correct cells

SV40 forces the host cell to express S phase & uses large T antigen and host proteins for genome replication

1. sv40 in particular: the large T and small t antigens have multiple functions, including manipulation of cell cycle and apoptotic pathways; 2. host cells susceptible to infection by SV40 typically in G0 prior to infection (so do not contain nuclear DNA replication proteins needed by virus); 3. virus must force cell to express S phase genes to replicate its genome; 4. changes in gene expression are accomplished by early nonstructural proteins-- large T antigen and small t antigen; 5. large T antigen interacts with pRB-- prevents pRB from interacting with EF2 transcription factors-- the S phase genes are no longer repressed; 6. large T antigen also interacts with p53-- prevents p53 from promoting apoptosis; 7. small t antigen inhibits a phosphatase complex (PP2A) which would otherwise dephosphorylate pRB; small t antigen therefore further biases the system toward expression of S phase genes

most class I animal viruses rely on host transcription machinery for gene expression

1. transcription by class I animal viruses catalyzed by host enzymes 2. transcription occurs in the nucleus; is catalyzed by RNA polymerase

late SV40 transcription is regulated by both host and viral proteins

1. transcripts initiate from a second promoter in NCCR-- transcription proceeds in opposite direction around genome from early transcriptions

viral genetic diversity arises through mutation and recombination

1. two major sources of change in the nucleotide sequences over time: mutation and recombination

drugs to treat flu target uncoating and release stages of viral replication

1. uncoating = existing drugs that treat influenza infections target M2 & NA; are envelope-associated proteins unique to virus; a. after flu virus enters cell thru endocytosis, cell acidifies the endosome; b. M2 matrix protein allows protons to cross viral envelope-- triggers rearrangements of HA viral spike; c. rearranged spike activates process of fusion of viral envelope w/ host endosome membrane-- genome is released into cytoplasm

polyomavirus replication cycle

1. uncoating releases the genome with nucleosomes into the nucleus; 2. early mRNAs are synthesized by host Pol II; 3. they exit the nucleus; 4. and are translated; 5. making early proteins that manipulate the host cell; 6. early proteins enter the nucleus; 7. and enable genome replication; 8. late mRNAs are produced by host Pol II; 9. they exit the nucleus; 10. and are translated; 11. late proteins enter the nucleus; 12. where they assemble with new genomes; 13. and are ultimately released thru host cell lysis

last stages of virus replication cycle are assembly, release, and maturation

1. viral assembly: a. refers specifically to assembly of capsids w/ genomes; b. is universal to all viruses (although varies w/ viral group); c. enveloped viruses must also acquire a membrane during assembly; 2. release: a. similar in all viruses- 1. result is extracellular infectious particles; 2. different methods of release = lysis and budding

synthesis of mRNA in SV40 is controlled by the noncoding control region

1. virus DNA encodes seven proteins; 2. genomes expressed in nucleus, where host factors needed for mRNA production are abundant 3. genome has 2 promoters facing in opposite directions; are [art of noncoding control region (NCCR)

might be possible to develop bacteriophage therapy to treat people w/ antibiotic-resistant bacterial infections

1. viruses not only microbes that evolve resistance to drugs used to treat them; antibiotic resistance among pathogenic bacteria is an increasing problem; 2. microbiologists interested in developing bacteriophages as pharmaceuticals to treat infection for almost 100 years; 3. USSR was innovative in developing phages as treatment for bacterial infections; 4. phages commonly used in food industry to reduce bacterial loads during food processing

6 large T antigen proteins form hexamers--

2 hexamers bind to ori-- facilitates melting DNA at the origin (separate the strands); **DNA replication then proceeds bidirectionally (aka theta replication)

in principle- rational vaccine design includes antigens that should provoke immune response to protect against all IAV

= development of universal IAV vaccine

concatamer

DNA molecules w/ multiple genomes covalently attached to each other in tandem

checkpoint during G1 is particularly important for DNA virus replication

G1 checkpoint is controlled by growth factors that can stimulate the cell to enter S phase; many host cells susceptible to viral infection are differentiated, thus exist in G0 and therefore have not passed the G1 checkpoint; DNA viruses that replicate in the nucleus typically affect the cell cycle by causing a differentiated host cell to pass the G1 checkpoint in the absence of external growth factor stimulation; dsDNA viruses target cell cycle proteins (growth factors, etc.) to manipulate cell cycle & control DNA replication

all retroviruses encode 3 polyproteins

Gag, Gag-Pol (processed by viral protease enzyme), and ENV (processed by host protease enzyme); HIV-1 also encodes 6 accessory proteins

HSV-1 does the opposite: actively prevents host cell from expressing S phase genes

HSV-1 must encode all proteins needed for DNA replication b/c no analogous host proteins are available; **proteins include those that participate directly in replication as well as enzymes for deoxribonucleotide triphosphate (dNTP) synthesis

what genome type is IAV & what type of polymerase does it need initially

IAV genome = (-) ssRNA and uses RdRp (rna dependent rna polymerase)

small t antigen interacts with:

PP2A (phosphatase) complex and inhibits its activity, thereby preventing repression of S phase genes

what host enzyme is used for transcription of DNA to mRNA

Pol II

why would fewer people infected reduce the chances of avian flu developing the ability to be transmitted from person to person

RNA-based viruses have higher mutation rate b/c RdRp cannot proofread, the more people infected, the more viral replication occurs, and the more opportunities exist for a mutation that will allow it to be contagious from one person to another-- note an increase in probability, does not mean that it can't happen in small population of infected individuals

process of synthesizing the first cDNA strand is discontinuous; the reverse transcriptase begins copying one part of the (+) RNA template before jumping to distal part of template and continuing DNA synthesis, skipping over intervening (+) RNA

RNase H domain essential for jumping reactions

why isn't viral transcription enzyme RdRp, as with many rna-based viruses?

RdRp is an rna-dependent enzyme (template to be transcribed is RNA). Class I viruses are dsDNA, so the host enzyme can transcribe it

vaccines = critical components of effective public health

WHO estimates vaccines prevent 2.5 million deaths each year; vaccine is medical treatment that provokes an immune response

macrophage

a large phagocytic cell found in stationary form in the tissues or as a mobile white blood cell

origin of 1977 influenza pandemic strain is possible exception to antigenic shift as cause

a. 1977 strain is very similar to 1918 H1N1 strain archived in 1950s; b. 1977 epidemic probably caused by accidental release of a frozen lab stock of old H1N1 viruses

several mechanisms trigger switch to late transcription

a. 1st: late transcription typically repressed by several copies of different host proteins; collectively called initiator-binding proteins (lbps) that bind to NCCR; b. initiator-binding proteins remain at constant levels in the nucleus; c. after viral genome has replicated many times, lbps distribute themselves among the many copies of NCCR-- lbp-mediated repression of late transcription alleviated when not enough copies of initiator-binding proteins to bind to all NCCRs; d. 2nd, accumulation of large T antigen in nucleus stimulates late transcription and represses early transcription by binding to parts of NCCR

5 anti-flu drugs on the market

a. 2 (amantadine and rimantidine) inhibit the M2 protein; b. other 3 (oseltamivir, zanamivir, peramivir) inhibit NA enzyme & block budding

vaccinia virus structure is complex

a. 2 different infectious vaccinia virions exist (intracellular mature virus, IMV, and extracellular enveloped virus, EEV); b. both forms contain a core, core wall, and lateral bodies surrounded by an inner membrane; c. larger EEV form has another membranous external envelope w/ viral spikes; d. large linear genome inside composed of 194 kbp dsDNA w/ inverted terminal repeats-- form covalently closed hairpin termini

H1N1 strain of 2009 pandemic is multiple recombinant

a. 3 gene segments including HA originate from swine-H1N1 (contracted from humans in 1918 pandemic); b. 2 genome segments including one encoding NA originate from different swine virus- has recent ancestors in birds; d. last genome segment descended from H3N2 human viruses; originated w/ 1968 pandemic

1957 H2N2 virus is recombinant

a. 5 of its genome segments descend from 1918 H1N1 virus; b. 3 segments, including the 2 encoding HA and NA, originated from a different avian influenza strain

viruses use ESCRT machinery to bud from and exit cells

a. ESCRT machinery is recruited to form an ever-tightening spiral underneath budding virion; b. finally, a cellular ATPase disassembles the ESCRT machinery; c. result = release of enveloped virus

vaccine that protects against HBV:

a. HBV S gene cloned in yeast, where large S spike protein is overexpressed; b. S protein then purified and used in combination with an adjuvant in vaccine

also have an RNase H domain that removes RNA from RNA-DNA hybrids

a. RNase H (or RNH) = non-sequence-specific enzymes that catalyze the cleavage of RNA in an RNA/DNA substrate; b. uses a hydrolytic mechanism (hydrolysis- molecule of water breaks down one or more chemical bonds)

spillover = when disease jumps species barrier, zoonotic spillover includes:

a. SARS-CoV; b. MERS-CoV; c. HIV (chimps to humans); d. pandemic flu A; e. 2013-2016 ebola in Africa started from transmission from bat to toddler; f. SARS-CoV-2, origin unknown, bat viruses have similarities

mRNAs leave the nucleus as normal mRNP particles-- translated in the usual way-- produce the regulatory T antigens

a. T antigens have nuclear localization signals-- cause them to be imported into the nucleus; b. in nucleus, affect gene expression and genome replication; e.g., large T antigen binds to sequences in NCCR-- represses early mRNA production

2nd lambda lysis protein (endolysin) degrades peptidoglycan (in the periplasm)

a. access to periplasm thru pores made by holing; b. result: mesh of the peptidoglycan full of holes large enough to allow virions to escape

recombinant vaccines = attenuated vaccines where attenuated virus engineered to express proteins from another species, ex. adenoviruses

a. adenoviruses (dsDNA) have capacity to incorporate large amounts of DNA from other sources; b. have long been favored as recombinant vaccine vectors and gene therapy agents; possible candidate for universal IAV vaccine

maturation stage (only in some specific viruses)

a. assembled proteins undergo proteolysis (breakdown of proteins or peptides into amino acids by the action of enzymes-- obligatory for virion to be infectious- if type to use proteolysis); b. other cases: proteins must be rearranged in some way for the virions to be infectious; c. maturation events are irreversible; d. maturation occurs during or after release in eukaryotic viruses

once virions are leaving or leave cell surface and successfully bud away, need to avoid:

a. binding to outside cell surface (w/ spike proteins) so escape from cell is impossible; b. re-infecting same host cell

viruses that infect animals (bats, camels, chimps, swine) already adapted to many conditions that make it possible to replicate in humans

a. can replicate at human-like body temps.; b. have also evolved to respond to (defend against) mammalian defenses

at each checkpoint, cell assesses whether completion of cell cycle w/o any mutations is likely, then:

a. continues thru the cell cycle; b. if there is something irreparably wrong, exits the cell cycle-- typically undergoes apoptosis

release: NA is 1 of 2 viral spikes; is enzyme needed during release from host cells

a. during budding, HA spikes on newly formed virions can get stuck to neuraminic acid on surface of former host cell; b. HA enzyme (neuraminidase) cleaves the surrounding sialic acid sugars-- releases virus into extracellular space

following assembly:

a. enveloped virus exits host cell by budding; b. last steps of maturation actually occur at the same time or slightly after budding

two different strategies by which some viruses acquire envelopes

a. formation of internal structures of virion that occur in coordination w/ envelope acquisition process, e.g. HIV; -or- b. acquisition of envelope occurs sequentially after assembly of capsid and genome finished; assembled capsid interacts w/ a cellular membrane that also contains virus spike proteins & pinches off, e.g. flu virus

2 cases where a single HIV-1 mRNA encodes > one protein by manipulating translation

a. genome-length mRNA is used to translate either Gag polyprotein or Gag-Pol polyprotein; b. accessory vpu gene overlaps from env gene, Vpu protein is translated from the vpu-env mRNA thru use of an alternative start codon; c. production of a less abundant Gag-Pol polyprotein requires a -1 ribosomal frameshift

zoonotic flu strain at first not adapted to humans, some viruses unable to replicate in humans or only poorly, but sometimes virus replicates so ferociously that death rate is much higher than for human-adapted flu strains, like H5N1 avian flu virus

a. has 60% mortality rate in people (typical seasonal flu rate is < 0.1%); b. can only be contracted directly from birds; is not transmitted from human to human

NCCR

a. has no nucleosomes; b. plays a crucial role in gene expression & regulation; c. specific sequences & certain host proteins required for early viral gene expression; d. regulatory DNA includes AT-rich sequences, a 21-bp repeat region containing 2 copies of GC-rich sequence, & single 72-bp repeat; e. AT-rich region serves to initiate assembly of the Pol II transcription complex (TATA box)

3rd protein (spanin) needed to breach outer membrane

a. has transmembrane segments embedded in plasma membrane and in outer membrane (spanin spans the periplasm); b. fuses the inner membrane w/ outer membrane

covalent modification of histones alters chromatin

a. histone acetyl transferase enzymes add negatively charged acetyl groups to histones-- move apart due to repelling like charges b. histone deacetylation enzymes remove acetyl groups-- enable close nucleosome packing

sampling diverse environments leads to discovery of many new virus genomes, e.g.

a. human gut is rich source of plant virus genomes b/c of omnivorous diets; b. deep sea hydrothermal vents have diverse virus communities w/ esp. large populations of dsDNA bacteriophages & archaeal phages

bacteriophage lambda (release from host w/ cell wall)

a. infects bacteria that stain negative in the gram reaction-- means they have particular type of cell wall; a. E. coli is gram negative and the host for phage lambda

integrated proviral DNA from a retrovirus is maintained for lifetime of host cell; provides longest lasting expression of therapeutic gene

a. insertion of proviral DNA can also cause cancer, so carries risks

has repeated sequences such as inverted repeats and shorter tandem repeats, found at species-specific locations

a. inverted repeat = reverse order complement; b. tandem repeats = series of DNA bp repeated ≥ 2 times in a row

1st protein (holing) attacks PM

a. is a transmembrane protein; b. clumps together w/ other holing molecules; c. many holing proteins rearrange themselves-- form huge holes in PM

effective antiviral drugs also have good pharmacological properties

a. lack of/minimal side effects (often caused when drug binds to and interferes w/ normal human protein function); b. whether can be taken orally; c. length of time it persists in human body; d. how human body breaks down or otherwise modifies the drug

early transcripts in alternative splicing

a. large T antigen plays many regulatory roles; **b. small t antigen and large T antigen both required to drive cell out of G0 and pass the G1/S cell cycle checkpoint

circular genomes w/ dsDNA, but with gap where DNA is single-stranded

a. longer strand of DNA has terminal protein covalently attached to 5' end; b. shorter DNA is actually a DNA-RNA chimera w/ smaller number of RNA nucleotides at 5' end

what are functions of viral regulatory proteins

a. manipulate the host cell cycle; b. regulate viral gene expression; c. participate in viral DNA replication

HPV

a. naked spherical virions w/ single type of capsomer; b. encodes ≥ 12 proteins, **replication cycle is very similar to that of polyomavirus: after uncoating (including introduction of viral genome into nucleus), host Pol II transcribes viral early mRNAs

1918 H1N1 virus origin in birds

a. not known if it was an antigenic shift; b. after the human pandemic began, humans transmitted it to swine; is important for evolution of later pandemic strains

mutation

a. occurs when mistake is made during replication or a physical or chemical assault changes a nucleotide; b. cellular DNA polymerases have editing functions that keep rate of nucleotide substitutions very low, but many viral polymerases have no editing functions- viral RdRps are particularly mutagenic

viral genomes have size limit based on capsid size as well as mechanism of packaging the genome during assembly and maturation

a. packaging capacity of gene therapy vector = length of nonviral genes that can be inserted into vector

viruses have evolved mechanisms to prevent sticking to or re-infecting host, ex.

a. particular proteins collaborate to remove receptors from cell surface or prevent them from being displayed there initially (so nothing to remove); b. cells infected w/ enveloped viruses: budding in combination w/ metabolic strain on host causes its death

vaccinated people become sick b/c:

a. person has underlying immune deficiency; b. had another infection that altered response to the vaccine; c. attenuated vaccine acquired a new mutation that somewhat restored ability to replicate in humans; d. vaccine strain recombined w/ natural virus and co-infected person; **thus, inactivated vaccines are sometimes preferred over attenuated ones

not all maturation processes are well-understood; ex. studies of viral maturation

a. picornaviruses & HIV- maturation is caused by a viral protease found inside the maturing procapsid; b. procapsid = immature viral capsid that does not yet contain a genome

recombination between different influenza A viruses occurs often in swine

a. pigs more susceptible to avian influenza than humans; b. also susceptible to human influenza; c. when avian influenza replicates in pigs, is selection for offspring viruses that better infect pigs (alpha 2,3 linkage vs. alpha 2,6 linkage in sialic acids)-- also increases susceptibility to humans (due to similarities between pigs & humans, both mammals)

HIV uses 3 main strategies to encode proteins:

a. polyproteins- reduced amount of mRNA b/c mRNA encoding polyprotein has only one 5' UTR, one start codon, one stop codon, and one 3' UTR for entire polyprotein; b. alternative splicing to express proteins w/ overlapping coding sequences; c. translation of polycistronic mRNA

DNA viruses usually block p53

a. prevents apoptosis; b. enables phosphorylation of pRB-- forces the host cell past the G1 checkpoint

viral gene expression cannot occur until viral DNA becomes inserted into a host chromosome

a. process is catalyzed by a viral enzyme = integrase; b. integrase was packaged into virion and traveled w/ viral nucleic acids from cell periphery into nucleus; c. early steps use viral enzymes; some later steps catalyzed by cellular enzymes (vs. viral integrase)

gain of function experiments-- start w/ avian viruses and select those w/ improved transmission among lab mammals

a. purpose of experiments = determine genetic changes that might be necessary for an avian strain to become pandemic--predict likelihood, i.e. number of mutations needed, that pandemic will occur & consider location & human demographics **also to understand molecular bio of transmission

promising strategies in molecular bio to try to reduce chance that resistance can evolve

a. sequence many strains of virus, compare, & look for conserved regions among them; b. if conserved sequences (less than or = 1 amino acid) occur in active site of enzyme or in other surface-exposed regions of protein, can design something to bind/block function of protein

two general methods by which capsids and genomes associate w/ one another during assembly

a. sequential assembly = capsid forms separately from the genome (hollow shell) and is subsequently filled in another step; B. concerted assembly = capsomers & genome come together during genome synthesis assembly-- no separate empty capsid to fill

virions have only a few basic arrangements of capsomers and nucleic acids--determines the mechanism of assembly

a. shell-like capsid w/ nucleic acid inside, ex. icosahedral capsids; b. nucleocapsid in which capsomers coat the genome (tight physical association between the capsomers and genome); c. capsid or nucleocapsid to be surrounded by an envelope

classic experiments of Hershey and Chase

a. sought to determine whether DNA or proteins entered host to direct synthesis of new phages; b. used radioactive building blocks of dNTPs and amino acids to track chemical processes; c. results from measuring new infections: DNA entered host cells and directed synthesis of new bacteriophages, whereas protein remained outside of the host cell, far from the cytoplasmic site of bacteriophage production; d. if some form of division were responsible for virus replication, entire parental virus should have entered host cell (not the case)- also, the radioactive material would both be distributed to offspring phages in about equal amounts (also not true); **later, other research showed that even the complex bacteriophage T4 s/ icosahedral head, helical tail, and tail fibers could assemble spontaneously

new capsids either:

a. trafficked back to nucleus-- release new relaxed circular DNA into nucleus-- more genome copies; -OR- b. targeted for assembly of new enveloped virions

2 well-known poxviruses that infect humans

a. variola virus (smallpox, 1979 = eradicated from natural circulation, samples still exist in US and Russia); b. vaccinia virus (cow pox, used to immunize people to protect against smallpox, very closely related to variola-- induces immunity against both variola & vaccinia viruses, but does not cause disease in healthy people)

cells in G0 contain a complex of pRB protein

a. when bound to other factors-- represses expression of proteins needed for DNA replication; b. when phosphorylated (add phosphate group), pRB cannot bind to other factors that prevent DNA replication-- DNA replication proceeds

membrane invaginating (turning inside) to form vesicle inside multivesicular body is pinched off from outer membrane thru process known as: abscission

abscission catalyzed by protein complexes = ESCRT machinery (endosomal sorting complexes)

preintegration complex also includes at least some capsid proteins-- enable the DNA and its associated proteins to dock w/ a nuclear pore complex

after docking, preinitiation complex is released into the nucleoplasm (inside nucleus)

after entering host cells, capsids w/ genome inside transported to nucleus

after relaxed circular DNA is released into nucleus thru nuclear pore, host enzymes convert it into covalently closed circular DNA (cccDNA)

to lyse host cell, bacteriophage lambda encodes different proteins that each attack one of the layers

all 3 proteins expressed late in infection-- is time for assembly of offspring phages before lysis occurs

genomes can be transcribed and translated by software & the proteins they encode can be subjected to bioinformatics analysis

all w/o ever culturing a single virus

what is a protease

an enzyme that breaks down proteins & peptides

antigenic definition

antibody or immune response triggered by antigens on a particular virus-- antigenic shift is a change in antigens that trigger an immune response- often replacement of the HA or NA w/ novel HA or NA

antigenic =

antibody or immune response triggered by antigens on a particular virus; antigenic variation = virus presents many different triggers for the immune system, including many that the immune system is not able to recognize

definition of mutation

any change to DNA or RNA sequence of nucleotide bases

bacteriophages, picornaviruses, and reoviruses

assemble, express/replicate genomes in the cytoplasm

vaccines provoke long-lasting immunity and require few booster shots to repopulate pool of memory lymphocytes

b/c attenuated viruses can replicate in the body & administered to millions of people, attenuated viruses cause small number of people to get sick

gain of function experiments = controversial

b/c of danger that lab-selected strain could escape lab; disagreement regarding whether risks of gain-of-function transmission experiments outweigh public health benefits of knowledge gained

viral world is very diverse

by any measure, e.g. virion size, composition, shape, and enveloped or non-enveloped (naked)

human papmillomaviruses encode about 13 proteins that are translated from polycistronic mRNA

can also encode variants of some proteins

enveloped HSV-1 enters host cell by fusion w/ plasma membrane

capsid is transported to nuclear pore-- viral DNA is released into nucleus (capsid remains in cytoplasm); cellular enzymes in nucleus ligate ends of viral genome together to make circular DNA

many viruses infect terminally differentiated cells such as macrophages or neurons

cell nuclei in G0 phase lack most DNA replication proteins; **progression thru cell cycle not automatic-- is controlled by 3 checkpoints

most class I viruses prevent or delay cellular apoptosis

cells have a mechanism to detect and respond to DNA damage; e.g. body's natural defense against cell transformation that can lead to cancer

inactivated vaccines contain adjuvants

chemicals that enhance immune response by simulating danger (i.e cell death without killing cells)

approx. 147 bp (depending on species) of eukaryotic DNA are wrapped around each histone

chromatin resembles beads (of nucleosomes) on a string in electron micrographs; **histones packed together by a 5th histone protein that binds to the linker DNA between nucleosomes

**unlike polyomavirus: important aspect of papillomavirus replication cycle--

closely tied to the differentiation status of host cells; **papillomaviruses can only infect stratified squamous epithelia, such as that found in human skin; cells in each layer are uniquely differentiated to serve unique functions of each layer

nucleosome

consists of a segment of DNA wound around 8 histone proteins (2 each of 4 different proteins), resembles thread wrapped around a spool

both enzymes also able to copy the (-) cDNA to synthesize (+) DNA--

creates dsDNA copy of a (+) RNA template

although adjuvant improves immune response, inactivated vaccines typically require more boosters over person's lifetime to repopulate pool of memory lymphocytes

current inactivated vaccines in use for one of two types of polio vaccine

cellular machinery that moves viral components from cell periphery to nucleus

cytoskeleton, with motor proteins

have been instances where something went wrong during inactivation process & people injected w/ infectious virions...

danger makes subunit vaccines more attractive b/c safer, although less immunogenic

can also attenuate viruses thru genetic engineering

delete genes needed to cause infection in humans but are not required for growth in culture (then force mutation, rather than waiting for one to occur)

method to stop evolution of more dangerous descendants of H5N1 avian flu =

destroy infected poultry when there is an epizootic (pandemic in nonhuman animals)

disadvantage w/ phages:

difficult to create collection of phages that could reasonably be expected to kill a particular pathogen during a natural infection

subunit vaccines = very safe, b/c do not originate as infectious microbes- however-

do not provoke a very strong response on their own, so boosters and strong adjuvants are necessary

NCCR of SV40 includes many regulatory sites

early promoter drives transcription to the left, and the late promoter drives transcription to the right; **NCCR has an AT-rich region, 3 21-bp sequences, and 2 72-bp sequences, all of which are multifunctional in that they interact with a variety of different proteins depending on the stage of the virus replication cycle

how do most plant viruses infect plants (mode of entry)

entry thru some type of cell damage, usually thru chewing animals, esp. arthropods

retroviruses and rhabdoviruses

enveloped, assemble at plasma membrane

coronaviruses and poxviruses

enveloped, assemble on internal membrane

herpesviruses and influenza virus

enveloped, assemble some components in nucleus & other components at the plasma membrane or at an internal membrane site

what is an endosome/its function

eukaryotic intracellular membrane-bound vesicle, used for transporting molecules & substances around cell

gene expression and genome replication among hepadnaviruses also involves reverse transcription

even though genomes are predominantly DNA

urgent need for new/better vaccines to prevent viral infection

ex. no HIV, MERS, SARS, cancer-causing viruses, or cold vaccines

as w/ RNA viruses w/ RdRp, influenza A has high misincorporation rate (mistakes in copying/mutations)

exhibits high rates of antigenic variation

after entering cell, core of vaccinia virion is released into cytoplasm--

forms a virus factory very similar to VRCs

cccDNA is maintained as an extrachromosomal (not in a chromosome) genetic element throughout lytic infection

gene expression begins w/ host's Pol II synthesizing viral mRNAs and viral pregenomic RNA

w/ many viruses:

genes encoding most abundant structural proteins do not overlap w/ other coding sequences

some viral mRNAs are spliced, whereas the full-length genomic RNA is not spliced

genomic DNA exported to cytoplasm = translation

attenuated vaccines replicate slowly in human body

gives adaptive immune response time to develop

1st gene therapy trials happened in last 25% of 20th century

have been ~2000 trials

virus also expresses a regulatory RNA during late gene expression

helps shut down production of early proteins

groups of viruses that have prodigy virions that escapae by budding

herpesviruses, (-) ssRNA including influenza

terminally differentiated cell

highly specialized cell that does not typically undergo mitosis; have exited the cycle, passing from G1 into resting phase (called G0), vs. continuing to S phase; some differentiated cells remain in G0 for the rest of their life; others can respond to signals (e.g. growth factors)-- induce cell to exit G0 and re-enter the cycle at G1 stage-- then progress thru G1, S, G2, and M

viral replication complex

host proteins & viral nucleic acids assembled on a specific membrane

release from animal cells can occur by budding

in secretory vesicle (material to be secreted from cell), fusion event results in the interior contents of the vesicle from being released; viral budding from PM is opposite of this process

universal flu vaccine desperately needed- to protect people and agricultural animals against most forms of IAV

including seasonal & zoonotic strains

viral genome replication likely proceeds first thru theta replication and secondary thru rolling circle replication

infected cell nuclei ultimately fill w/ concatamers of viral dna

what is latent infection?

infections where host cell contains a viral genome & a few other viral macromolecules, but it is mostly inactive for a long time (i.e., years) and no virions are produced

although seasonal flu vaccines are useful, universal flu vaccine = highly sought after

influenza A (IAV) is major pathogen; causes millions of serious infections and 250-500k deaths every year worldwide

outcome of dsDNA virus infection determined by:

interaction of particular virus and host cell; same virus can have different outcomes in different host cells

first wave of gene expression = production of proteins to counteract host immunity in various ways

intermediate gene expression = induces replication of viral genome; late gene expression = focused on production of structural proteins-- assemble into progeny virions in nucleus-- leave cell thru lysis

rational vaccine design

involves study of immune response to virus to determine which viral antigens are capable of inducing a strong sterilizing adaptive immune response

single infected cell typically produces 100,000 progeny virions

is 10-100 times more than burst size of most viruses that infect animal cells

when cell is co-infected w/ 2 different viruses, can create recombinant (hybrid) offspring w/ new combo of genome segments

is origin of most pandemic influenza strains

polyprotein

large proteins that are digested into individual protein components, each w/ unique activity

DNA replication coincides w/ late gene expression

late genes encode structural and assembly proteins that allow new virions to assemble and ultimately escape from the host cell by budding

late transcription does not occur until genome replication has begun

late genes encode structural proteins needed to complete replication cycle

attenuated vaccines, cont.

made from microbes that can still replicate, to some extent, in human body but cannot cause symptomatic infection in healthy people; attenuated by propagating virus in nonhuman host or host cell under different environmental conditions than those in human body; in situation other than humans, virus must acquire mutations that enable replication

multivesicular endosomes (MVEs) are complex intracellular organelles that function in endocytosis

major function of cell's endocytic pathway: sort internalized macromolecules and membrane proteins; interior contents of vesicular body remains interior to vesicle after pinching off

IAV has segmented genome where major antigenic proteins are encoded by different segments

makes possible for 2 different flu viruses to enter cell at same time

both mutation & recombination (forms of genetic variation) = problematic for vaccine development

manufacturers produce ~500 million doses of vaccine yearly; volume needed means vaccine manufacturers begin many months before flu season; a. manufacturers make educated guess about which forms of flu will be circulating during following flu season-- part of data collection = US manufacturers monitor flu strains in Australia during summer before fall flu season in US (their winter flu)

herpesviruses have very large enveloped virions and large linear dsDNA genomes

many w/ large spherical enveloped virions w/ several distinctive layers, including envelope spikes, outer & inner tegument, and inner icosahedral capsid

ultimately, new virion assembly occurs in nuclear viral factories

mature virions exit host cell thru lysis

RNA vaccines only have to cross plasma membrane (not nuclear envelope as w/ DNA)

may improve possibility of successful in vivo (inside organism) transfection- under certain conditions, mRNA is taken by cells

current attenuated vaccines

measles, mumps, rubella, seasonal flu, chicken pox, polio, rotavirus, yellow fever, and rabies

adenoviruses produce 2nd protein (death protein) that kills cells when it accumulates during late stages of infection & causes lysis

mechanism not understood; other animal viruses that exit by lysis encode proteins that enhance or cause lysis of plasma membrane

most common gene therapy vectors in development today =

modified adenoviruses (dsDNA), parvoviruses (ssDNA), or retroviruses

polycistronic =

molecule of mRNA that can encode for ≥ 2 proteins

natural selection & pesticide resistance-- same principle applies to viruses & anti-viral drugs

more drastic the difference between normal & mutated individuals, the stronger the selection; by contrast, if the pesticide only reduced reproduction somewhat, the selective pressure would not be as strong & it would take longer for the mutant strain to increase significantly in the population

plant cell walls are metabolically inert layers; provide protection from osmotic shock, mechanical strength, and rigid support for whole plant

most plant viruses also require some type of cell damage to exit cells; asexual reproduction in clonal plants can spread virus to different clones arising from same rootstock (virus does not need to cross thru cell wall)

b/c of the differences, viruses cannot use secretory machinery for budding

must use proteins designed for pinching off so that the contents inside of vesicle remain on the interior; such a system exists among host's endosomes b/c contents of endosomes becomes pinched off during normal endosome maturation to form multivesicular bodies (multiple vesicles)

polyomaviruses, papillomaviruses, & adenoviruses

naked, assemble in nucleus

first wave of HSV-1 transcription (immediate early) promotes transcription of next wave of gene expression, also interferes w/ host's immune response

next wave (early gene expression) encodes proteins needed for replication of viral genomes

before Pol II can initiate transcription, chromatin has to be remodeled (spread apart) so the promoter DNA is available

next, transcription initiation requires assembly of a complex array of basal transcription factors

when prodigy virions escape by budding, does the cell usually recover and continue living?

no, cell usually dies- its amino acids, nucleotides, & ATP are depleted, so it cannot maintain homeostasis

do walls protect cells in a hypertonic environment from drying out?

no, cell walls do not prevent water from leaving the cell

checkpoint during G1

normal cell = s phase promoters repressed--no DNA replication (cell stays in G0 phase w/o passing checkpoint), vs. w/o repression of S phase promoters-- DNA replication (cell passes checkpoint into S phase); **the tp53 gene (human version of p53) is the most frequently mutated gene (>50%) in human cancer

late gene expression

occurs after genome replication has begun; late genes encode structural proteins needed to complete virus reproductive cycle

offspring viruses collected and infection under non-human conditions repeated many times =

passaging; virus becomes attenuated b/c now adapted to favor infection of nonhuman cells-- viruses collected from patient and then propagated in avian tissue culture cells at a temp. cooler than the human body, shed viruses are collected and procedure repeated many times-- as virus adapts to in vitro passaging, it becomes attenuated for replication in the human body)

how is peptidoglycan critical in the bacteriophage replication cycle

peptidoglycan degraded as part of new virion release

advantage to using phages to treat bacterial infections =

phages extremely specific- kill pathogen w/o affecting commensal and beneficial microbiota (unlike antibiotics)

DNA replication always begins at the same seq. of DNA, the origin of DNA replication (ori)

present in non-coding control region

prophylactic/preventative vaccines

prevent disease

late transcription terminates approx. halfway around genome--

produces 2 alternatively spliced mRNAs that together encode 4 proteins

viral genes are transcribed by host enzyme (Pol II)--

produces typical eukaryotic capped and polyadenylated mRNA

different forms of the polymerase are specialized for

producing mRNA, rRNA, and tRNA; **RNA polymerase II (abbreviated as Pol II) catalyzes mRNA synthesis

early gene expression

production of nonstructural proteins-- manipulate cell cycle, regulate viral gene expression, & ultimately initiate viral genome replication

gene expression occurs in somewhat overlapping waves

progeny virions assemble in dedicated cytoplasmic viral factories-- released by both lysis and budding

what is apoptosis

programmed cell death; natural pathway used during development and during immune responses; also a defense to protect whole animal from uncontrolled cell proliferation (aka cancer); protect whole animal from a virus attempting to complete its replication cycle

stratified epithelium

proxmial/first layer of cells = basal stem cells, actively dividing to give rise to the rest of the epithelium; midzone cells are nondividing; upper level formed by dead keratinized cells = stratum corneum; **virus can only infect the living basal cell layer; injury is necessary to allow tiny virion to bypass upper layers of epithelium

what is a frameshift

reading frames (3-base codon) are shifted by one or more individual bases, either upstream or downstream, in this case, -1, the ribosome slips back one nucleotide base and continues translation

early viral transcripts

regulatory (non-structural) vs. viral proteins (structural)

herpesvirus genome results in concatamers

replication of HSV-1 DNA takes place in different biochemical context than polyomavirus, papillomavirus, and adenovirus, which force cell to express S phase genes

diseases eradicated thru vaccination

rinderpest (cattle) & smallpox (humans)

what type of assembly would include a procapsid

sequential assembly process (vs. concerted)

sometimes manufacturers guess on target, other times is a mismatch between vaccine and most prevalent form of flu

situation can discourage public from getting vaccinated at all

b/c supply of basal transcription factors is limited, they cannot assemble on all promoters simultaneously

situation provides opportunity for regulating gene expression

alternative splicing used to produce multiple HIV-1 pre-mRNA molecules w/ same promoter

some of exons for one protein overlap w/ exons or introns from other genes

retroviruses are enveloped and have RNA genomes yet express their proteins from dsDNA

some steps of retrovirus reproductive cycle occur in cytoplasm; others occur in nucleus

metagenomics =

study of genetic material recovered directly from environmental samples (collective genome of microorganisms from an environmental sample)

reverse transcriptases do not have 3' to 5' exonuclease editing activity--

their mutation rate is about 10,000 times higher than the DNA polymerase used during host DNA replication: a. reverse-transcribing viruses give rise to offspring populations w/ tremendous genetic diversity

sterilizing vaccinated person completely protected from acquiring viral disease

then viral antigens classified as exhibiting high or low levels of antigenic variation, **best candidates for effective universal vaccine = proteins capable of evoking a strong adaptive immune response and also exhibit low antigenic variation

b/c inactivated virus cannot replicate--does not induce cell death--may be recognized as foreign, but is not recognized as dangerous...

therefore does not result in strong innate immune response (required for long-lasting immunity)

unlike all other groups of dsDNA viruses, poxvirus expresses its genome and replicates its DNA entirely in cytoplasm

therefore, nuclear host proteins not available for gene expression and genome replication, e.g., a. host splicing machinery confined to nucleus; b. b/c poxviruses do not encode splicing machinery, vaccinia mRNAs do not have introns

how do plant viruses infect other cells in the same plant?

thru plasmodesmata

gene therapy can also be used to treat cancer, e.g.

transform cancer cells w/ gene to make them more sensitive to apoptosis or sensitive to pharmaceutical

therapeutic vaccines

treat existing infection

viral replication also causes enough destruction that damage-associated molecular patterns are produced

triggers additional immune response

pregenomic RNA is encapsilated, then reverse transcribed inside new capsid-

ultimately forming new relaxed circular DNA

most diversity at level of molecular genetics

use every possible strategy for encoding protein & many employ unique gene expression & genome replication strategies not otherwise found within molecular hosts

COVID-19 vaccines are only authorized/approved mRNA vaccine

use mRNA to direct cells to produce copies of a spike protein; **researchers studying how mRNA might be used to develop vaccines for additional infectious diseases (research has been ongoing for decades)

after attachment, is partial uncoating of the virus genome; next is long process of reverse transcription where infecting (+) mRNA genome is converted into dsDNA

viral dsDNA enters nucleus-- viral integrase enzyme inserts viral dsDNA genome almost randomly into host genome (unique)

besides high mortality rate, zoonotic flu has possibility that w/ only few mutations in right places--

virus might gain ability to be passed from human to human and still maintain high mortality rate

as part of host antiviral response, host packages viral DNA into heterochromatin (condensed form of chromatin)

viruses have mechanisms to counteract problem-- changes its genome into euchromatin (loose form of chromatin) during lytic infection

recombination (larger scale genetic change than typically classified as mutation)

when 2 viruses coinfect same cell and offspring virus contains combo of genes that originate with both parents (vs. one parent); aka reassortment (sometimes used only for segmented RNA viruses; recombination applicable to all viruses); **form of horizontal transfer

zoonotic disease

when flu or other illness jumps from animals to humans/crosses species barrier

when histones are tightly packed together-- transcription is blocked, vs.

when histones are associated more loosely w/ DNA-- transcription machinery (i.e. polymerase) can access DNA

dna of viral origin, once inserted into host chromosome = provirus

when provirus has formed, viral gene expression can occur

mRNA vaccines

work by introducing piece of mRNA that corresponds to a viral protein, usually for small piece of protein found on virus' outer membrane

reducing # of people infected by avian flu important for reducing possibility of virus developing ability to be transmitted from person to person

zoonotic IAVs that have gained ability to be transmitted from human to human are very dangerous b/c they can cause pandemics w/ high fatality rates