Viro Study guide
Poxviruses - cytoplasmic, pathogenesis, biological weapon, etc. (not the details of replication cycle)
Cytoplasmic: o Cytoplasmic replication o No access to host transcription or replication machinery (has its own) - therefore, big!!! o Virion has all enzymes for transcription, polyadenylation and capping (RNA pol, Transcription factors , capping enzyme, ATPases, DNA helicases, ligases, topoisomerases, protein kinases) · Pathogenesis o High mortality rate o Variola major (20%+), variola minor (2-5%) o Spread by inhalation (respiratory), gets to skin o Forms open sores o Resistant to desiccation (contamination lingers in clothing, bedding, etc.) o Does not induce chronic infection - gets cleared or death of the host. Survivors are immune. · Biological weapon • Eradicated in 1970's. Only stocks in Russia and U.S.???? • Susceptibility is worldwide, outbreaks are possible • Availability of vaccines - $$$$ + resources • Mistakes/problems in dissemination are expected • Experts are alive who can reconstruct it • Political solution preferred • Need to inform & empower the population
EBV and immortalization of B cells & pathology
EBV is Epstein-Barr Virus, which is a lymphotropic herpesvirus that infects B lymphocytes primarily. It is also known to infect epithelial cells in the nasopharynx. EBV infection in adolescents and adults can lead to sever mononucleosis (Mono). Both primary and latent infectons with EBV are characterized by the expression of a viral gene product that interferes with the induction of apoptosis in B lymphocytes.
Temporal regulation of HSV-1 gene expression (order of gene expression).
Earliest genes expressed are important in "priming" the cell for further viral gene expression, in mobilizing cellular transcriptional machinery, and in blocking immune defenses at the cellular level This phase is then followed by the expression of a number of genes that are either directly or indirectly involved in viral genome replication & Finally upon genome replication, viral structural proteins are expressed high abundance
Temporal gene regulation by T4 phage (sequential use of the same polymerase)
Four stages: immediate-early, delayed-early, quasi-late and late virus modifies the specificity of host RNA polymerase for temporal regulation of viral gene regulation by the expression of phage-specific σ factorand by modification of the core enzyme by phage-encoded enzymes.\
What is HHV-8?
HHV-8 is a type of human herpesvirus that is associated with the human cancer called Kaposi's sarcoma (KS). Was known as a rare disease of very old men in the late 19th century. Marked by slow formation of sarcomas made up of highly pigmented epithelial cells. Is not invasive but eventually leads to death. Occurrence is associated with loss of immune capacity and specific geographical and genetic factors. HHV-8 is known to encode for bcl-2 gene, which inhibits apoptosis, (Oncogenic transformation) a G protein-coupled receptor, which is active and can cause transformation of cultured cells.
1. How DNA viruses avoid end problems
Herpesviruses and many bacterial DNA viruses have genomes with repeated sequences at their terminals so that the viral genome can become circular via a recombination event following infection. Thus, even though the virion DNA is linear, replicating viral DNA in the cell is either circular or joined end to end in long concatemers. These structures are then resolved to linear ones when viral DNA is encapsidated. -Adenovirus, on the other hand, has solved the problem by using a primer that is covalently bound to a vial protein that binds to the viral DNA's end. Further, adenovirus DNA proceeds only continuously; there is no discontinuous strand synthesis. -Small single-stranded (ss) DNA viruses, like parvoviruses, have solved the problem by encoding a complementary repeat sequence at the end that allows the genome to form a "hairpin loop" at the end; thus, the end of the molecule is not free. A similar solution is seen in the genome structure of poxvirus. Like chromosomal DNA, this linear DNA genome is covalently closed at its ends. Thus, in effect, replication just proceeds "around the corner" onto the complementary strand.
Temporal gene regulation in T7 phage (sequential use of different polymerases)
Insert genomes into cell starts transcription of early genes Host RNA polymerase makes 5 mRNAs producing T7 RNA polymerase for delayed early and late genes Immediate early gene phosphorylates host RNA polymerase to shut down host transcription delayed early genes makes T7 DNA polymerase for replication Late genes makes structural, lysis genes for assembly/exit
How HSV-1 evades immune response during reactivation
a. the inhibition of MHC class-I mediated antigen presentation at the surface of the infected cell by the α47 protein slows the host's ability to detect the earliest stages of productive infection. b. HSV encodes a protein, ICP34.5 that blocks interferon's inhibition of translation in the infected cell by inhibiting phosphorylation of the translational initiation factor eIF-2. This result in ensuring efficient translation of the small amounts of viral transcripts expressed in this limited infection. c. Virus encodes a protein that inhibits the infected cell's tendency to undergo apoptosis
Herpesvirus - neurotropic and lymphotropic, examples
Many herpesviruses are neurotropic, meaning they actively infect nervous tissue. All such viruses are collectively termed alpha-herpesviruses. i. Three types of human herpesviruses belong to this group, they are herpes simplex 1 and 2 and varicella-zoster virus (VZV) Five human herpesviruses are lymphotropic, meaning they replicate and establish latency in tissues associated with the lymphatic system. These herpes are divided into beta and gamma herpesvirus groups based on the specifics of their genome structure and replication. a. an example of beta-herpesvirus is human cytomegalovirus, which is linked both to a form of infectious mononucleosis and to congenital infections of the nervous system. b. An example of gamma-herpesviruses is EBV Epstein-Barr Virus and Kaposi's Sarcoma herpesvirus or human herpesvirus-8, all of which are linked to human cancer. c. Herpesvirus genome contains about 60-80 genes (80-240 kbp genome). There is a large variation of genomic structure. Despite this, gene order is maintained within large blocks of the genome "and varying degrees of genetic homology. A general feature that herpesviruses share is their regions of inverted sequences. The size of herpesvirus genomes varies from 80kbps to 240kbps.
HSV-1 budding cascade
Mature capsids bud through the inner nuclear membrane that contains the viral glycoproteins. In early maturation process in the nucleus, capsids appear to be surrounded by the primary tegument protein, UL31, and this directs the budding through the inner nuclear membrane into which the UL31 phosphorylated membrane protein has been inserted. The primary envelope capsids then bud through the outer nuclear membrane where the primary envelope is lost. The cytoplasmic capsids then associate with tegument proteins of the mature virion. Final envelopment takes place as mature capsids associated with the tegument proteins bud into exocytotic vesicles.
Papillomavirus and wart
Papillomavirus forms benign tumors. Papillomavirus replication involves both productive and abortive schemes like in polyomavirus. These viruses cause warts or papillomas that tend to be benign and can spread from host to host through sexual intercourse. There are two types of human papillomaviruses, they are HPV16 and HPV18. a. HPV-16 is about 7900 base pairs long and is similar to that of SV40 but contains more early ORFs. The early and late promoters are found in several regions within the genome. i. How are warts form in HPV-16? 1. The virus enters the basal cells of the epithelium. The virus then expresses genes that induce cells to replicate their DNA rather more frequently than would an uninfected epithelial cell. Papillomavirus DNA remains in the infected cell nucleus as an episome or "mini" chromosome where it can replicate when cell DNA replicates, but not in high numbers, until the SV40 virus which replicates and dies off. (Plasmid like replication). The virus change the functions of cells. For example, K5 and K14 keratins are terminated while K1 and K10 of the suprabasal skin cells are expressed. Since replication is highly localized and differentiation is sped up, a wart is formed. ii. For HPV-16 and HPV-18, the growth enhancement is a function of proteins encoded by E5, E6 and E7 gene products that inactivate normal functions of p53 and Rb proteins. iii. HPV-6 and HPV-11 are also other strains that can infect humans
Papovavirus = ""Papilloma, polyoma, vacuolating"
Papovavirus stands for : Papilloma, polyoma, Vacuolating * Two distinct families: Papilloma virus & polyoma virus families * Icosahedral capsid * Circular genome * Chronic infection * Ability to remain with the host for long period. * Need to alter cell growth in host cell response to neighboring cells for viral replication
SV40 & how SV40 synthesizes large and small T antigens & functions of large T antigen
Polyoma virus (simian Vacuolating Agent 40). * Forms tumors in animals, transforms primary cells * Persistent infection with little evidence of disease. * Kills cells very slowly. * No global virus induced shutdown of host function. * SV40 virus originally was found as a contaminant of African Green monkey kidney cells (AGMK) in which poliovirus was being grown for vaccine purposes. sup bbe (hey higlight what you think will be in free response!!!! SV40's early region transcription of early mRNA encodes small t and large T antigens. Large T antigen migrates to nucleus and carries out numerous functions: Activation of cellular DNA and RNA synthesis by binding to the cellular growth control gene products named Rb and p53. This binding stops these control proteins from keeping the cell contact inhibited. This function causes the infected cell to begin a round of DNA replication. Blockage of apoptosis that is normally induced in cells where p53 is inactivated at inappropriate times in the cell cycle. Binding to the SV40 ori to initiate viral DNA replication. Shutting off early viral transcription by binding to regions in and near the early promoter. Activating late transcription. Playing a role in virion assembly.
T4 phage morphogenesis (how the components of virus come together to make progeny)
Process of head assembly and DNA encapsidation similar to that of the other large DNA viruses described. After filling of the head, the other components of the complex virion, which have preassembled to form subassemblies, come together to form the complete particle. all steps are simple biochemical reactions driven by mass action mature phage is released from the infected cell by expression of a late lysozyme that disrupts the bacterial cell wall, releasing virus.
Role of large T antigen in virus-induced carcinogenesis - integration and stable transformation
Roles of the large T antigen include: The large T antigen alters the host cell to allow it to replicate viral DNA. The large T antigen also binds to the SV40 ori to allow DNA replication to begin and to shut off synthesis of early mRNA. Every round of DNA replication requires T antigen to bind to the origin of DNA replication and initiate a round of DNA synthesis. a. activation of cellular DNA and RNA synthesis by binding to the cellular growth control gene products named Rb and p53. Keeps cell contact inhibited. b. Blockage of apoptosis that is normally induced in cells where p53 is inactivated at inappropriate times in the cell cycle. c. Binding to the SV40 ori to initiate viral DNA replication d. Shutting off early viral transcription by binding to regions in and near the early promoter e. Activating late transcription f. Playing a role in virion assembly.
SV40 genome & how different proteins are produced
SV40 Genome: contains 5243 base pairs dsDNA genome 4 genomes Early region 1 mRNA precursor with 2 ORFs Alternate splicing to express the ORFs when the larger region is spliced out, you get a larger protein due to the removal of the stop codon alternate splicing produces different proteins
How T even phages avoid bacterial restriction defenses:
T-even phages use unusual base: 5-hydroxymethylcytosine (5-OH MeC) instead of C 5-hydroxyMeCTP precursor is made by viral enzyme (T4 hydroxymethylase) - first viral enzyme discovered in 1957 DNA is then modified by glycosylation (1 or 2 glucose residues added to hydroxymethyl residues) Modified DNA evades bacterial restriction defenses.
HSV-1- type of genome, replication cycle - productive infection (acute phase) vs. latency and reactivation. HSV-1 replication: role of vhs,TIF, rolling circle mechanism
TIF: alpha-trans-inducing factor protein; also called VP16 or UL48 Functions in enhancing immediate-early viral transcription It does this by interacting cellular proteins such as Oct1 (octamer-binding proteins) It is an extremely powerful transcriptional activator with very broad specificity Its C-terminal region contains a very large number of acidic amino acids that activate transcription by mobilizing RNA polymerase bound to the preinitiation complex at the promoter in the vicinity of the enhancer region Vhs: virion-associated host shut-off protein; also called UL 41 remains in cytoplasm where it causes disaggregation of polyribosomes and degradation of cellular and viral RNA. Rolling Circle: (in theory) generates a continuous concatemeric strand of newly synthesized viral DNA that is available for encapsidation In actuality, as DNA is being replicated, new synthesis begins at any one of a number of ori's and highly concatenated, linked networks of DNA are formed in the infected cell. Linear genome, 152 kb long (152,000 base pairs) Uses circular intermediate 100 transcripts, mostly unspliced. 70 + ORFs Each gene has a promoter and polyA signal "dispensable genes" for pathogenesis 6 gene regions in genome 1. Ends: repetitive DNA sequences for packaging 2. RL regions: long repeat (9 kb) for immediate early regulatory protein and promoter for latency associated transcript (LAT) for reactivation. 3. Long unique region (UL): 56 + proteins (replication, capsid and others) 4. Rs regions: immediate early protein a4 that is a transcription activator (for viral transcription) 5. Ori's: 3 short regions. 6. Us: unique short region. 12 ORFs for glycoprotein and response to host defense. Productive Infection (Vegetative Cycle) · o 1.Entry: Sequential interactions to attach & enter HVEM (herpes virus entry mediators) -Unknown function in normal cells Membrane fusion Tegument protein transports nucleocapsid to nucleus Vhs 2. Immediate early gene expression Alpha-TIF and OctI activate gene expression. 5 protein products for transcriptional regulation, post-transcriptional processing, and inhibition of antigen presentation by MHCI o 3. Early gene expression Early beta-genes, activated by alpha genes, for viral DNA replication and other "non-essential" functions o 4. Genome replication and late gene expression High level of replication at the replication compartments in the nucleus...cell destruction Early genes turn off, late genes are turned on. o 5. Virus assembly and release: 30+ structural genes assemble around viral scaffolding proteins and interact with viral DNA in nucleus. Latent Infection The viral genome of HSV and many (but not all) other alpha-herpesviruses is not fully shut off during latent infection The HSV LAT is a large transcript that is weakly expressed during productive infection, but unlike all other known productive cycle transcripts, it does not "shut down" in latently infected cells. HSV-Latent Infection Neurotropic Enters sensory nerve axon near site of infectiion Migrates to nucleus of neuron HSV-1 favors lip/facial (trigeminal ganglion) HSV-2 favors sciatic nerve ganglia Productive infection leads to cell death During latency, HSV DNA is wrapped with histone as mini-chromosome or episome Neurons don't replicate so HSV is dormant Stress activates host transcription & virus activation ... some viral replication Travels down axon and reinfects the area it first infected Reactivation doesn't mean damage to trigeminal nerve ganglion Reactivation: Replication Cycle attachment enters through breakage in the skin attachment and entry Endocytosis into an endosome acidification allows virion release virion migrates to a nuclear pore complex early genes regulate virus genome replication by shutting off host translation late genes make the capsid capsid is released via exocytosis.
