L2: Transcription & processing of eukaryotic genes Flashcards
What are the main components of a transcription initiation complex in eukaryotes?
- RNA polymerase II
- promoters
- general transcription factors (GTFs)
In eukaryotes, what is the order in which transcription is initiated?
- binding of RNA polymerase II to the promoter region of the gene
- followed by recruitment of GTFs
Explain the mechanisms of processing eukaryotic mRNA during transcription initiation
-RNA polymerase II binds to the promoter region
- GTFs facilitate the assembly of the transcription initiation complex, leading to the start of mRNA synthesis
What are the mechanisms involved in termination and polyadenylation during eukaryotic mRNA processing?
- recognition of specific termination sequences
- then the addition of a poly-A tail at the 3’ end of the mRNA
Provide examples of gene expression regulation at different stages of RNA processing
- Gene expression can be regulated during transcription initiation by the binding of TFs to enhancer regions.
- Polyadenylation can be regulated by factors that influence the cleavage and addition of the poly-A tail.
What is the central dogma of molecular biology?
- states the unidirectional flow of genetic information: DNA (genes) undergo transcription to form RNA, which - then translated into proteins.
What is the role of DNA transcription and RNA processing in gene expression?
- DNA transcription = process of synthesizing RNA from a DNA template, and RNA processing = involves modifications to the RNA molecule, such as splicing & polyadenylation.
both play a key role in determining cellular fate and function
How does the gene expression pathway determine cellular fate?
- gene expression pathway regulates which genes are transcribed & processed into RNA
- leads to production of specific proteins that govern cellular functions & determine the fate of the cell
What is the process of eukaryotic transcription?
DNA unwinds to expose bases.
One DNA strand acts as a template.
RNA synthesis directed by the template.
Forms an initiation complex with RNA pol II and promoter.
Transcription factors assist in complex formation.
RNA elongated one nucleotide at a time.
RNA transcribed in the nucleus.
RNA is complementary to template DNA strand.
Uracil (U) replaces thymine (T) in RNA.
RNA helix reforms behind transcription.
RNA released as a single strand.
RNA is a messenger and information carrier.
What are the main types of RNA in eukaryotic cells and their functions?
Messenger RNA (mRNA): Codes for proteins (3% total RNA).
Ribosomal RNA (rRNA): Forms ribosome structure, catalyzes protein synthesis (71% total RNA).
Transfer RNA (tRNA): Acts as adaptors between mRNA and amino acids (15% total RNA).
How can each type of RNA be characterized?
- by size and sedimentation behavior (Svedberg’s coefficient, S).
- Sedimentation behavior influenced by density, mass, and shape
What is the significance of mRNA in a cell?
mRNA is a small percentage of total RNA.
Codes for all functional proteins in the cell.
mRNA is the only coding type of RNA.
DNA → Transcription → mRNA → Translation → Protein
How is DNA transcribed into different types of RNA?
DNA transcribed into mRNA (coding RNA) or non-coding RNA.
rRNA, tRNA, etc. are non-coding types of RNA
What is the transcription initiation complex and how is it formed?
- complex forms upstream of a gene on DNA.
- Interaction between genomic DNA promoters and General Transcription Factors (GTFs) and RNA pol II.
- GTFs needed for transcription initiation
What are promoters and their role in transcription?
- Short sequences upstream of the coding region.
- RNA pol binds to promoters.
Example: TATA box, conserved sequence 25-35bp upstream. - TATA box positions RNA pol II for proper initiation
What is the TATA box and its significance?
- Conserved promoter sequence (TATAAA/TATAAG).
- Positions RNA polymerase II through GTFs.
- Identified by analyzing base frequency in aligned eukaryotic genes
Why can’t RNA pol II directly bind to DNA?
- RNA pol II needs GTFs to bind first.
- GTFs help in positioning RNA pol II at the promoter
What are GTFs?
- Required for RNA pol II transcription initiation.
- Multimeric and conserved proteins.
- Binding follows an ordered manner.
- Position RNA pol II at the promoter.
- Complex process involving hundreds of proteins.
How are DNA-protein interactions involved in transcription initiation?
- DNA-protein interactions form the pol II transcription initiation complex.
- Short DNA sequences (e.g., TATA box) interact with proteins (GTFs).
- GTFs are required for assembly of the transcription initiation complex
What is the step-wise assembly of the transcription initiation complex?
- TATA-binding protein (TBP) binds TATA-box.
- TFIIB binds to the TBP complex.
- TFIIB acts as a bridge for RNA pol II recruitment.
- RNA pol II, TFIIE, and TFIIH associate to form preinitiation complex.
- TFIIH unwinds DNA using ATP hydrolysis and phosphorylation.
- RNA pol II leaves promoter, starts elongation of nascent RNA
How is pre-mRNA processed and what is its significance?
- Entire gene, including introns and exons, is copied into pre-mRNA.
- Pre-mRNA processing involves capping, polyadenylation, and splicing.
- Mature mRNA lacks introns, has a 5’ cap, protein coding region, 3’ UTR, and poly-A tail
What is the process of 3’ end cleavage and polyadenylation of mRNA?
- Cleavage and addition of poly A tail to 3’ end during RNA processing.
- Polyadenylation stabilizes RNA, enhances nuclear export, aids in translation.
- Assembly of complex involves CPSF, CStF, CFI, CFII, and PAP
How does the assembly of the cleavage/polyadenylation complex resemble transcription initiation?
- Both involve multi-protein complexes forming through interactions.
- Assembly cooperatively occurs through protein-nucleic acid and protein-protein interactions
What are the key steps in the cleavage/polyadenylation complex assembly?
- CPSF binds to AU-rich PolyA signal.
- CStF, CFI, CFII proteins bind, causing pre-mRNA bending.
- PAP cleaves 10-35 nucleotides upstream of G/U-rich PolyA signal.
- Free 3’ end is polyadenylated by PAP (slow polyadenylation)
