Ch.7, Part 1 - DNA to RNA Flashcards
What are the chem and struc diffs b/w DNA and RNA?
- RNA differs fr DNA chemically: (1) ribonucleotides; (2) uracil (U) instead of thymine (T), wh can still base-pair by H-bonding w A.
- RNA also differs fr DNA structurally: RNA is single-stranded → can fold into diff shapes (unlike DNA) → diversity of funcs, incl structural, regulatory, and catalytic roles.
T/F: All RNA in cell is made by transcription.
True
All RNA in cell is made by transcription.
Briefly summarize the process of transcription.
Transcription:
- Open/unwind small portion of DNA double helix to expose bases on ea DNA strand.
- One of two strands of DNA double helix acts as template for RNA synth.
- Ribonucleotides are added, one by one, to growing RNA chain
- Nucleotide seq is det by complem bp (H-bonding) w DNA template → incoming ribonucleotide covalently linked to growing RNA chain via RNA pol.
- RNA transcript has ntide seq exactly complem to template DNA.
Summarize how transcription differs fr DNA replication.
Xcr differs fr DNA repl:
- RNA transcript does not remain H-bonded to template
- RNA is single-stranded.
- RNAs are copied fr limited region of DNA → much shorter than DNA.
- RNA pol
- Like DNA pol, RNA pols catalyze formation of phosphodiester bonds that link ntides t/g and form sugar–P backbone of RNA chain.
- RNA pol moves stepwise along DNA → unwinds DNA helix just ahead to expose new region of template strand for base-pairing → growing RNA extended one ntide at a time in 5′-to-3′ direction.
- Incoming ribonucleoside triphosphates (ATP, CTP, UTP, and GTP) provide energy to drive rxn fwd.
Summarize the diffs b/w RNA and DNA polymerases.
Diff b/w RNA/DNA pol:
- Both catalyze formation of phosphodiester bonds that link ntides t/g and form sugar–P backbone.
- Both grow one ntide at a time in 5’-to-3’ direction.
- RNA pol uses ribonucleoside for phosphates as substrates → catalyzes linkage of ribonucleotides, not deoxyribonucleotides.
-
RNA pol can initiate synth w/o primer.
- Likely evolved bc xcr need not be as accurate as DNA repl bc RNA not used as permanent storage form of GI → mistakes in RNA xcrs have relatively minor consequences for cell.
- Rate of mutation: RNA pol = one mistake per 104 ntides; DNA pol = one mistake per 107 ntides.
The RNA transcript is almost immediately released from the DNA template as it is synthesized. How does this mechanism influence the rate of transcription?
The immediate release of the RNA transcript after synthesis results in many RNA copies can be made fr same gene in relatively short time.
- I.e. synth of next RNA typ begins before synth of prev RNA completed.
- E.g. medium-sized gene (~1500 ntide pairs) xcr’s in ~50 seconds.
- Many RNA pols work simult along single stretch of DNA → 1000+ transcripts synth’d w/I 1 hour.
The vast majority of genes in a cell’s DNA specifies AA seqs of proteins → encoded by RNA → directs protein synth (via messenger RNAs; mRNAs). How do mRNAs differ in euks and bacteria?
Vast majority of genes in cell’s DNA specify AA seqs of proteins → encoded by RNA → directs protein synth (via messenger RNAs; mRNAs).
- Euks: ea mRNA typ carries info xcr’d fr just one gene → single protein
-
Bac: set of adj genes often xcr’d as single mRNA → carries info for several diff genes/proteins.
- Recall: operons.
Non-protein coding genes result in RNAs wh serve many roles in cell, incl regulatory, structural, and catalytic. Describe of few types of these ncRNAs.
Non-protein-coding RNAs:
- Ribosomal RNAs (rRNAs) - struc/catalytic core of ribosomes → translate mRNAs into protein.
- Transfer RNAs (tRNAs) - adaptors; select specific AAs and hold in place on ribosome for incorporation into protein.
- MicroRNAs (miRNAs) - key regulators of euk gene expression.
Transcription initiation is critical as it is the main point at wh cell selects wh proteins/RNAs to prod. Describe the recognition process (xcr initiation) in bacteria.
Xcr initiation in bacteria:
- RNA pol randomly collides w DNA → sticks weakly to double helix → slides rapidly along its length.
- RNA pol links tightly only after encountering promoter: gene region; contains specific ntide seq that lies immediately upstream of starting point for RNA synth.
- RNA pol bound tightly to promoter → RNA pol opens double helix immediately in front of promoter to expose ntides on ea strand of a short stretch of DNA.
Consider a bacterial cell in wh RNA pol has tightly bound the promoter and opens the double-helix immediately in front of promoter to expose ntides on ea strand of a short stretch of DNA. What occurs next in bacterial transcription?
Xcr elongation/termination in bacteria:
- One exposed DNA strand acts as template → two incoming ribonucleoside triphosphates joined t/g by RNA pol to begin synth of RNA transcript.
- Elongation continues until RNA pol encounters terminator (stop site): gene region; RNA pol halts and releases both DNA template and RNA transcript.
- Terminator seq is contained w/I gene → xcr’d into 3ʹ end of new RNA transcript.
What are bacterial promoters and terminators?
Bacterial promoters and terminators and gene regions w specific ntide seqs that are recognized by RNA polymerase.
-
Promoters:
- Polarity of promoter orients the RNA pol and dets wh DNA strand is xcr’d.
- All bacterial promoters contain DNA seqs at -10 and -35 ntide positions (see Fig)
- Not xcr’d into RNA transcript.
- Note the typ TATA box.
-
Terminators:
- Xcr’d into RNA transcript.
T/F: transcription of a bacterial gene typ starts at the promoter sequence.
False
Promoter seqs allow RNA pol to recognize DNA strand to be xcr’d, but promoter isn’t xcr’d into an RNA transcript. I.e. xcr initiates further downstream of promoter seqs.
What are sigma factors wrt bacterial RNA pols?
Sigma factors are a subunit of bacterial RNA pols wh are primarily responsible for recognizing promoter seqs.
- Recog promoters w/o unwinding DNA double-helix by scanning unique, exterior features of bases themselves.
How does the “polarity” of promoters affect transcription?
Promoters have certain polarity: two diff ntide seqs upstream of xcr start site → position RNA pol and ensure binding in only one orientation.
- RNA pol can only synth RNA in 5′-to-3′ direction → must use DNA strand oriented in 3′-to-5′ direction as template.
T/F: On a single chromosome, transcription always proceeds in the same direction.
False
While xcr always occurs in 5’-to-3’ direction, genes can be transcribed fr either strand. Thus, xcr proceeds in either direction on a single chromosome.
Describe two ways in wh RNA polymerases are diff b/w bacteria/euks.
Diffs in bac/eukRNA pol:
- Bac contain single type of RNA pol; euks have three types RNA pol (I, II, III) → ea resp for xcr diff types of genes.
- RNA pol II xcr’s vast majority of euk genes, incl all that encode proteins and miRNAs.
- Bac RNA pol (along w sigma subunit) is able to initiate xcr on its own; euk RNA pols req assistance of many accessory proteins.
- E.g. general transcription factors (GTFs): must assemble at ea euk promoter (along w RNA pol) before xcr initiation.
How are general transcription factors involved in euk transcription?
GTFs are accessory proteins that assemble on promoter (gene region) whr they help position RNA pol and pull apart DNA double helix to expose template strand → allows RNA pol to initiate xcr.
- Incl TFIIB, TFIID, etc.
Euk transcription initiation assembly process typ begins w binding of TFIID (GTF) to a short segment of DNA double helix composed primarily of _______ ntides, also called a _______.
Euk transcription initiation assembly process typ begins w binding of TFIID (GTF) to a short segment of DNA double helix composed primarily of T/A ntides, also called TATA box.
- TATA box is typ located ~25 ntides upstream fr promoter.
- Subunit of TFIID—TATA-binding protein (TBP)—recognizes and binds TATA seq.
Euk xcr initiation begins w TFIID (GTF) binding a TATA box upstream of the promoter seq. What happens as a result of this binding?
TFIID causes dramatic local distortion in DNA, wh helps direct other proteins to promoter → other factors (TFIIB) and RNA pol II assemble to form complete xcr initiation complex.
Before euk xcr can begin, RNA pol II must dissoc fr the complex of GTFs at the promoter. How does this occur?
Liberation of RNA pol II is initiated by GTF TFIIH, wh contains a protein kinase subunit → adds P groups to RNA pol II “tail”
Euk mRNA transcripts (pre-mRNA) must be processed before leaving the nucleus to begin translation. What processing steps must occur, and where are the enzymes that facilitate these steps located?
Post-xcrprocessing—capping,splicing, andpolyadenylation—isconcurrent wxcr.
- Enzymes involved are located on phosphorylated tail of RNA pol II → process transcript as it emerges fr pol II
Describe the post-xcr process of RNA capping.
RNA capping:
- Adds an atypical guanine (7-methyguanosine) to 5’ end of RNA transcript (leading end/synth’d first).
- Capping occurs after RNA pol II has produced ~25 ntides of RNA, i.e. long before xcr finishes.
Describe the post-xcr process of polyadenylation.
Polyadenylation:
- 3′ end of growing mRNA is first trimmed by an enzyme that cuts transcript at a partic seq of ntides → second enzyme adds series of repeated adenines (typ few hundred) to cut end.
T/F: expressed seqs (exons) are typ shorter than introns and represent only small fraction of total length of gene region.
True
expressed seqs (exons) are typ shorter than introns and represent only small fraction of total length of gene region.
- Introns range in length fr 1 to 10,000+ ntides.
- Most euk protein-coding genes have many introns.
- Terms “exon” and “intron” apply to both DNA and corresponding RNA seqs.
Describe the post-xcr splicing mechanism in euks.
Splicing mechanism:
- Snurps recog & bp splice-site signal seqs on ends/tips of introns.
- Carried out mainly by small nuclear RNAs (snRNAs), wh are packaged w proteins to form small nuclear ribonucleoproteins (snRNPs, “snurps”).
- Snurps form core of spliceosome: large assembly of RNA/proteins; carries out RNA splicing.
- 2’-OH of ribose of adenine near 3’ end of intron attacks 5’ splice site of exon 1/intron → cleaves RNA backbone by donating OH gr to 3’ end of exon 1.
- Cut 5’ end of intron covalently links w 2’-C wh donated OH → free 3’-OH end of exon 1 attacks 5’ splice site of exon 2 → cleaves RNA backbone by donating OH gr to 3’ end of intron.
- Intron “lariat” released and exons 1/2 covalently linked → lariat eventually degraded.
Due to intron/exon structure of mRNA transcripts, euks can utilize “alternative splicing”. What does this mean?
Alt splicing:
- Allows many diff proteins to be produced fr same gene.
- Occurs in ~95% of humans genes.
- Thought to have sped up emergence of new/useful proteins, i.e. novel proteins appear to have arisen by mixing/matching of diff exons of pre-existing genes.
- Many proteins in present-day cells resemble patchworks composed fr common set of protein pieces: protein domains.
Poly-A–binding proteins, cap-binding complexes, and exon junction complexes serve what function in post-xcr processing?
Export-ready mRNAs must bind a set of proteins, ea wh targets diff parts of mature mRNA and facilitate export.
- E.g. Poly-A–binding proteins, cap-binding complexes, and exon junction complexes (spliced mRNAs).
- Entire set of bound proteins—rather than any single protein—dets whether mRNA exported.
- “Waste RNAs” are degraded in nucleus → ntides reused for xcr.
- Note: after proteins bind mature mRNA, a nuclear transport receptor (regulator protein) directs transcript toward nuclear pore (ch.15).
