3600 Quiz 4

how does RNA splicing happen

(A) In the first step, a specific adenine nucleotide in the intron sequence (indicated in red) attacks the 5ʹ splice site and cuts the sugar-phosphate backbone of the RNA at this point. The cut 5ʹ end of the intron becomes covalently linked to the adenine nucleotide, as shown in detail in (B), thereby creating a loop in the RNA molecule. The released free 3ʹ-OH end of the exon sequence then reacts with the start of the next exon sequence, joining the two exons together and releasing the intron sequence in the shape of a lariat. The two exon sequences thereby become joined into a continuous coding sequence. The released intron sequence is eventually broken down into single nucleotides, which are recycled.

RNA capping Is the First Modification of Eukaryotic Pre-mRNAs

1. the 5ʹ end of the new RNA molecule is modified by addition of a cap that consists of a modified guanine nucleotide 2. Three enzymes, acting in suc- cession, perform the capping reaction: one (a phosphatase) removes a phosphate from the 5ʹ end of the nascent RNA, 3. another (a guanyl transferase) adds a GMP in a reverse linkage (5ʹ to 5ʹ instead of 5ʹ to 3ʹ), 4. and a third (a methyl transferase) adds a methyl group to the guanosine Because all three enzymes bind to the RNA polymerase tail phosphorylated at the Ser5 position—the modification added by TFIIH during transcription initiation— they are poised to modify the 5ʹ end of the nascent transcript as soon as it emerges from the polymerase. The 5ʹ-methyl cap signifies the 5ʹ end of eukaryotic mRNAs, and this land- mark helps the cell to distinguish mRNAs from the other types of RNA molecules present in the cell.

how many RNA polymerase do eukaryotes have

3

Polymerase II Also Requires Activator, Mediator, and Chromatin- Modifying Proteins. WHy and what do they do?

4, DNA in eukaryotic cells is packaged into nucleosomes, which are further arranged in higher-order chromatin structures. As a result, transcription initiation in a eukaryotic cell is more complex and requires more proteins than it does on purified DNA. First, gene regulatory proteins known as transcriptional activators must bind to specific sequences in DNA (called enhancers) and help to attract RNA polymerase II to the start point of transcription Second, eukaryotic transcription initiation in vivo requires the presence of a large protein complex known as Mediator, which allows the activator proteins to communicate properly with the polymerase II and with the general transcription factors. Finally, transcription initiation in a eukaryotic cell typically requires the recruitment of chromatin-modifying enzymes, which can increase access to the DNA in chromatin, and by doing so they facilitate the assembly of the transcription initiation machinery onto DNA.

Hill equation

A mathematical expression for the degree of saturation of ligand binding to a molecule with multiple binding sites as a function of the ligand concentration. θ is the fraction of DNA bound, [L] is concentration of the regulatory protein Kd is the apparent dissociation constant, and KA is the ligand concentration producing half occupation Theta on y axis, [L] on x as Ligand concentration increases, degree of saturation increases

Scanning Electron Microscope (SEM)

A microscope that uses an electron beam to scan the surface of a sample, coated with metal atoms, to study details of its topography. - it generally has a greater depth of field. - It detects electrons that are scattered or emitted from the specimen.

RNA

A single-stranded nucleic acid that passes along genetic messages, flexible, odd base pairs, important for ribosomes, protein synthesis

A DNA-binding protein binds cooperatively to DNA by fairly weak homodimerization. The binding of this protein to DNA

A. shows a sigmoidal (S-shaped) binding curve. B. occurs in more of an all or none manner, compared to a protein that is a constitutive dimer (that is in a dimeric state all the time whether DNA bound or not) C. occurs despite the fact that the protein molecules are predominantly monomers in solution.

What are the advantages and disadvantages of developing antibody-based drugs over small molecule based drugs? List at least 2 advantages and 2 disadvantages.

Adv. Known Pharmacokinetics, Know toxicities (less toxicities and less unexpected side effects since a protein naturally in body). Less development cost. Specific binding to target. Disadv. Expensive to make, expensive to store, unstable, expensive to ship, has to be dilute to prevent aggregation,

The following schematic drawing represents the activation of transcription for a eukaryotic gene(gene X). Indicate what sequences B and C might be and the functions of each . Hint: C gets translated by RNA polymerase III.

B. (green section bound to things) cis-regulatory element/sequence. Regulates gene X expression. C. dotted line clear section lncRNA. Lnc RNA can produce functional RNA that suppresses or activated gene expression

Why is it surprising that DNA methylation is NOT different for Igf2 between the maternal and paternal chromosomes?

Because normally one expects the maternal copy that is not expressed to be methylated and suppressed.

The lipid composition of mitochondria in the table below only add up to less than 70%. What is the other major type of lipid in mitochondria (0.25 pts) and why is that component important for mitochondria to have regarding its shape/structure

Cardiolipin. Because of its small head group, it helps membrane maintain a curved structure easier. Important for highly curved mitochondria inner membrane.

what is positive cooperativity

Cooperativity can be positive (if binding of a ligand molecule increases the receptor's apparent affinity, and hence increases the chance of another ligand molecule binding) or negative (if binding of a ligand molecule decreases affinity and hence makes binding of other ligand molecules less likely).

Two haploid budding yeast cells are allowed to mate. One of them carries a mutation in itsmitochondrial DNA that makes the yeast cell resistant to an antifungal drug. If the resulting diploid zygote is allowed to propagate (in the absence of the drug), how do you predict that the fraction ofdrug-resistant cells will change over time in the population?

Curve B (goes from 100 to 50) The zygote (one cells) is resistant to the drug so 100% at beginning. However, mitochondria is mosaic in terms of its genetic make up. Over generations, mitotic segregation of mitochondrial DNA will randomly give rise to cells that only contain the wild-type, non-resistant organelles, and this should occur in about half of the population.

What are enhancers?

DNA sequences that increase the rate of transcription initiation by RNA polymerase II by binding transcription factors DNA sequences that bind other proteins that regulate transcription. They are really far away. do NOT get made into RNA

cis regulatory elements

DNA sequences that modify the expression of other genes that are nearby on the chromosome, often by acting as binding sites for transcription factors

promoter

DNA sequences that proteins bind to /specific region of a gene where RNA polymerase can bind and begin transcription indicates starting point for RNA synthesis

why do introns not show up in mature rna ?

Following transcription, new, immature strands of messenger RNA, called pre-mRNA, may contain both introns and exons. The pre-mRNA molecule thus goes through a modification process in the nucleus called splicing during which the noncoding introns are cut out and only the coding exons remain. Splicing produces a mature messenger RNA molecule that is then translated into a protein

For the α subunit of a trimeric G protein, does a G -protein-coupled receptor (GPCR) acts as a guanine nucleotide exchange factor (GEF) or a GTPase-activating protein (GAP)

GEF

Are G-proteins active when bound to GDP or GTP?

GTP bound state is active Becomes inactive when the G-protein hydrolyzes the GTP to GDP

Explain how PALM/STORM type of super resolution imaging works.

Have to sequentially turn on fluorescence from just a few single molecules at a time. Each time, the molecule can be accurately localized to center of the few spots of fuzzy light. By repeating this localization of a few single molecules many times over, a complete super resolution image is obtained. a super-resolution image of the fluorescent structure is built up as the positions of succesive small groups of molecules are added to the map

The following sequence logo represents the preferred cis-regulatory sequences of an imaginary transcription regulator that functions as a dimer. Would you expect this sequence to be recognized by a homodimer or a heterodimer ? Also explain reason

Heterodimer. Because sequence recognized is different at two positions. Each positions is likely a monomer recognition sequence.

Fibroblasts

In connective tissue, cells that secrete the proteins of the fibers.

lncRNA

Long non coding RNA, can suppress or activate gene expression gene silencing (Xist in x-chromosome inactivation)produce functional RNA that

How does DNA methylation differ between the maternal and paternal chromosome in terms of the H19 locus?

On the maternally inherited chromosome, H19 is transcribed and silences Igf2. This does not occur on the paternal chromosome. As a result, Igf2 is maternally imprinted with no maternal-specific methylation. So H19 is methylated on the paternal locus (not expressed) so does not interfere with Igf2 expression in paternal chromosome.

Replicative cell senescence

Phenomenon in which cell proliferation halts after a finite number of cell divisions, apparently due to loss of telomeres Phenomenon observed in primary cell cultures as they age, in which cell proliferation slows down and finally halts.

what does RNA polymerase II do

RNA polymerase II transcribes most genes, including all those that encode proteins,

what do RNA polymerase I and III do

RNA polymerases I and III transcribe the genes encoding transfer RNA, ribo- somal RNA, and various small RNAs.

Discuss how high positive cooperativity might explain how disease may arise from improper imprinting or X-inactivation.

Relatively small concentration changes (like twice what is needed because of improper imprinting or X-inactivation) might cause quite large effects.

what does RNA splicing do?

Removes Intron (non-coding )Sequences from Newly Transcribed Pre-mRNAs Both intron and exon sequences are transcribed into RNA. The intron sequences are removed from the newly synthesized RNA through the process of RNA splicing. Only after 5ʹ- and 3ʹ-end processing and splicing have taken place is such RNA termed mRNA. RNA SPLICING LEADS TO MRNA, and this mRNA is exported through nuclear pore complex The machinery that catalyzes pre-mRNA splicing is complex, consisting of five additional RNA molecules and several hundred proteins, and it hydrolyzes many ATP molecules per splicing event. This complexity ensures that splicing is accurate,

nucleolus

Ribosome-Producing Factory the site for the processing of rRNAs and their assembly into ribosome subunits. NOT BOUND BY A MEMBRANE it is a large aggregate of differnt RNA's and assembly factors, ribosomal proteins and partly assembled ribosomes rRNA genes have an important role in forming the nucleolus. Size of nucleolus reflects how many ribosomes the cell is producing

nuclear pore complexes (NPCs)

Successfully processed mRNAs are guided through the nuclear pore complexes (NPCs)—aqueous channels in the nuclear membrane that directly connect the nucleoplasm and cytosol. Small molecules can diffuse through but macromolecules require active transport across / through NPC's using energy macromolecules are moved through nuclear pore complexes by nuclear transport receptors, which, depending on the identity of the macromolecule, escort it from the nucleus to the cytoplasm or vice versa.

two regions of unpaired nucleotides situated at either end of the L-shaped molecule are what

T1. cloverleaf tRNA forms L shaped structure two regions of unpaired nucleotides situated at either end of the L-shaped molecule are crucial to the function of tRNA in protein synthesis. One of these regions forms the anticodon, a set of three consecutive nucleotides that pairswith the complementary codon in an mRNA molecule. The other is a short sin-gle-stranded region at the 3ʹ end of the molecule; this is the site where the amino acid that matches the codon is attached to the tRNA.

mRNAS > ______ > proteins

The codons in an mRNA molecule do not directly recognize the amino acids they specify: the group of three nucleotides does not, for example, bind directly to the amino acid. Rather, the translation of mRNA into protein depends on adaptor molecules that can recognize and bind both to the codon and, at another site on their surface, to the amino acid. These adaptors consist of a set of small RNA molecules known as transfer RNAs (tRNAs), each about 80 nucleotides in length. form cloverleaf structure

stop codons

The end of the protein-coding message is signaled by the presence of one of three stop codons (UAA, UAG, or UGA) (see Figure 6-48). These are not recognized by a tRNA and do not specify an amino acid, but instead signal to the ribosome to stop translation.

general transcription factors

The general transcription factors help to position eukaryotic RNA polymerase correctly at the promoter, aid in pulling apart the two strands of DNA to allow transcription to begin, and release RNA polymerase from the promoter to start its elongation mode. referred to as TFIID

How does the packing of cholesterol against other lipids alter the biophysical properties of a membrane?

The rigid core of cholesterol packs against alkyl chains of neighboring lipid molecules, reducing their flexibility.The net effect is to stiffen the membrane bilayer against distortion, while retaining fluidity. A membrane containing cholesterol is less permeable to small molecules, but does not substantially reduce the rate of diffusion of molecules within the membrane. ps cholesterol has hella rings and then a hydrophobic tail , which explains reduced flexibility

Nucleotide Sequences in mRNA Signal Where to Start Protein Synthesis > explain process >> FINISH

The translation of an mRNA begins with the codon AUG, and a special tRNA is required to start translation. This initiator tRNA always carries the amino acid methionine with the result that all newly made proteins have methionine as the first amino acid at their N-terminus, the end of a protein that is synthesized first. (This methionine is usually removed later by a specific protease.) The initiator tRNA is specially recognized by initiation factors because it has a nucleotide sequence distinct from that of the tRNA that normally carries methionine.

initiation of transcription by RNA polymerase II

To begin transcription, RNA polymerase requires several general transcription factors. (A) The promoter contains a DNA sequence called the TATA box, which is located 25 nucleotides away from the site at which transcription is initiated. (B) Through its subunit TBP, TFIID recognizes and binds the TATA box, which then enables the adjacent binding of TFIIB (C). For simplicity the DNA distortion produced by the binding of TFIID is not shown. (D) The rest of the general transcription factors,as well as the RNA polymerase itself, assembl e at the promoter. (E) TFIIH then uses energy from ATP hydrolysis to pry apart the DNA double helix at the transcription start point, locally exposing the template strand. TFIIH also phosphorylates RNA polymerase II, changing its conformation so that the polymerase is released from the general factors and can begin the elongation phase of transcription. As shown, the site of phosphorylation is a long C-terminal polypeptide tail, also called the C-terminal domain

DNA only transposons

Type of transposable element that exists as DNA throughout its life cycle. Many types move by cut-and-paste transposition require transposons short inverted (opposite facing) repeats at each end

differences between bacterial and eukaryotic RNA polymerase

While bacterial RNA polymerase requires only a single transcription- initiation factor (σ) to begin transcription, eukaryotic RNA polymerases require many such factors, collectively called the general transcription factors. 2. Eukaryotic transcription initiation must take place on DNA that is packaged into nucleosomes and higher-order forms of chromatin structure, features that are absent from bacterial chromosomes.

three possible reading frames in protein syntehsis

]In the process of translating a nucleotide sequence (blue) into an amino acid sequence (red), the sequence of nucleotides in an mRNA molecule is read from the 5ʹ end to the 3ʹ end in consecutive sets of three nucleotides. In principle, therefore, the same RNA sequence can specify three completely different amino acid sequences, depending on the reading frame. In reality, however, only one of these reading frames contains the actual message.

Transmission Electron Microscope (TEM)

a microscope that passes an electron beam through very thin sections stained with metal atoms and is primarily used to study the internal ultrastructure of cells

TCA cycle / citric acid

a series of metabolic reactions that break down molecules of acetyl CoA to carbon dioxide and hydrogen atoms

Compare nuclear and mitochondrial DNA in terms of the following. a. presence of non-coding DNA b. Use of RNA to protein codons

a. Mitochondria has hardly any non-coding DNA b. Mitochondria codon usage is loose relative to inside nucleus

What is a housekeeping gene?

constitutive genes that are required for the maintenance of basic cellular function, and are expressed in all cells of an organism under normal and patho-physiological conditions

mRNA polymerase

copies both introns and exons into pre-mRNA, 5' to 3', no replication forks, just brief bubbles that close, no okazaki fragments

You perform flow FISH using these probes on two human cell types, an early primary fibroblast culture and a late secondary culture that is showing the signs of replicative cell senescence.The sorting results are presented in the following histograms. Which histogram (1 or 2) would you expect to correspond to the primary culture.

curve showing highest intensity represents primary culture because During replicative cell senescence, the telomeres become progressively shorter, resulting in weaker fluorescence/intensity in this experiment. once telomeres are too short, cell can no longer proliferate

meosis needs __ for DNA crossover

double strand break

The Spliceosome Uses ___ to Produce a Complex Series of RNA-RNA Rearrangements

extensive ATP hydrolysis is required for the assembly and rearrangements of the spliceosome.

what are reads

https://www.rna-seqblog.com/mapping-rna-seq-reads-visualized/

given a hypothetical set of RNA -seq data, the number of reads is plotted for a region of chromosome containing two genes, from samples obtained from two different tissues. > how can you tell which gene is a houseleepin gene?

if its expressed/ has high reads in both organs

corn has different kernal colors because of

jumping genes

mRNA production

mRNA production is made more efficient in the nucleus by an aggregation of the many components needed for transcription and pre-mRNA processing, thereby producing a specialized biochemical factory. In (A), a postulated scaffold protein holds various components in the proximity of a transcribing RNA polymerase. Other key components are bound directly to the RNA polymerase tail, which likewise serves as a scaffold (see Figure 6-22), but for simplicity these are not shown here. In (B), a large number of such scaffolds have been brought together to form an aggregate that is highly enriched in the many components needed for the synthesis and processing of pre-mRNAs. Such a scaffold model can account for the several thousand sites of active RNA transcription and processing typically observed in the nucleus of a mammalian cell, each of which has a diameter of roughly 100nm and is estimated to contain, on average, about 10 RNA polymerase II molecules in addition to many other proteins. (C) Here, mRNA production factories and DNA replication factories have been visualized in the same mammalian cell by briefly incorporating differently modified nucleotides into each nucleic acid and detecting the RNA and DNA produced using antibodies, one (green) detecting the newly synthesized DNA and the other (red) detecting the newly synthesized RNA.

small nuclear RNAs snRNAs

molecules recognize the nucleotide sequences that specify where splicing is to occur and also catalyze the chemistry of splicing. These snRNPs form the core of the spliceosome, the large assembly of RNA and protein molecules that performs pre-mRNA splicing in the cell. During the splicing reaction, recognition of the 5ʹ splice junction, the branch-point site, and the 3ʹ splice junction is performed largely through base-pairing between the snRNAs and the consensus RNA sequences in the pre-mRNA substrate.

Non-retroviral retrotransposons

moves via RNA intermediate synthesized by nearby promoter require reverse transcriptase and endonuclease polyA at 3' end , 5' end usually truncated

Retroviral-like retrotransposons

moves via an RNA intermediate whose production is driven by a promoter in the LTR require reverse transcriptase and integrase directly repeated long terminal repeats at each end > >

X inactivation *

one of two X chromosomes is randomly inactivated and remains coiled as a Barr body

which RNA is more abundant?

rRNA is 80% of RNA, makes machinery to synthesize proteins mRNA about the other 15%, they regulate gene expression

Discuss why many regulatory proteins might exhibit positive cooperativity and how it might be useful for cells to have things work that way.

regulatory proteins have switch-like behavior, and can switch from off to on with a reasonable concentration change Regulatory proteins bind to specific sequences in the DNA and control which genes to turn on under any particular conditions. Regulatory proteins themselves often receive information by binding small signal molecules, whereupon they change shape which alters their ability to bind DNA.

the ribosome is a ___

ribozyme RNA molecules that possess catalytic activity are known as ribozymes. ribosomal proteins stabilize the RNA core, while permitting the changes in rRNA conformation that are necessary for this RNA to catalyze efficient protein synthesis. The proteins also aid in the initial assembly of the rRNAs that make up the core of the ribosome.

list the RNAS

that small nuclear RNA (snRNA) mole- cules direct the splicing of pre-mRNA to form mRNA, that ribosomal RNA (rRNA) molecules form the core of ribosomes, and that transfer RNA (tRNA) molecules form the adaptors that select amino acids and hold them in place on a ribosome for incorporation into protein. In Chapter 7, we shall see that microRNA (miRNA) molecules and small interfering RNA (siRNA) molecules serve as key regulators of eukaryotic gene expression, and that piwi-interacting RNAs (piRNAs) pro- tect animal germ lines from transposons;

TATA box

the general transcription factors assemble at promoters used by RNA polymerase II. The assembly process begins when TFIID binds to a short double-helical DNA sequence primarily composed of T and A nucleotides. For this reason, this sequence is known as the TATA sequence,

Which region (1 or 2) within gene Y most likely corresponds to an exon and why

the section that is a peak. Introns dont show up in mature rna and therefore will be a trough in the graph

heterodimer

when two polypeptides encoded by different genes bind to each other to form a dimer can recognize a sequence at two different positions

homodimer

when two polypeptides encoded by the same gene bind to each other to form a dimer.