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)
How does rapid polyadenylation occur in the cleavage/polyadenylation complex?
- PABII binds to short A tail.
- Accelerates A addition by PAP (rapid polyadenylation).
- PABII signals PAP to stop polymerization after 200-250 residues
What is the basis of transcription initiation and polyadenylation?
- transcription initiation = involves DNA sequences binding to General Transcription Factors (GTFs).
- Polyadenylation = involves conserved sequences in pre-mRNA binding to proteins
How do interactions differ in RNA processing and transcription initiation?
- RNA processing = involves interactions between RNA sequences & proteins.
- Transcription initiation = involves interactions between DNA sequences & proteins
Why is gene expression carefully regulated in cells?
- Gene expression controlled to ensure only necessary genes expressed
- Proteins produced in varying amounts based on cell’s needs
- Correct gene expression allows cells to perform specialized functions
How does gene expression regulation occur?
- Gene expression regulated in response to cell’s function & needs.
- Key regulatory stages include differentiation, development, and environmental responses
What are key stages of gene regulation and their examples?
- Occurs during differentiation, development, and environmental responses.
- Example: Cyclins needed for cell division expressed at specific cell cycle stages
How can problems in gene expression lead to diseases?
- Cancer: Absence of transcription off switch leads to improper gene expression and cell division.
- Developmental diseases: Mistakes in protein synthesis can result in organ developmental problems.
- Chronic diseases like obesity: Abnormal gene expression due to incorrect signals
How is transcription initiation regulated in eukaryotic gene expression?
- Transcription initiation controlled by interactions between DNA sequences & regulatory proteins
- Regulatory proteins (trans) have DNA binding & transcription activation domains.
- Regulatory sequences (cis) are short DNA sequences influencing transcription
What are the types of gene transcription and their characteristics?
- Constitutive genes: Constantly expressed (house-keeping genes).
- Regulated genes: Selectively expressed based on conditions or cell types
How do different types of transcription factors function?
- Two groups of TFs: general and sequence-specific.
- Proximal promoters and TATA box play roles in transcription initiation
How are regulated genes controlled?
- 5’ end of regulated genes contains regulatory cis sequences and promoters.
- Enhancers, promoters, & specific TFs control gene expression.
- Combination of protein factors in TIC regulates gene expression
How does hormone regulation affect gene expression?
- Hormones from endocrine system switch genes on/off.
- Hormones bind to receptors, initiate signaling cascades.
- Hormones stimulate synthesis of proteins in target tissues
How can hormones affect gene expression?
- Hormones can bind to cell membrane receptors, starting signaling cascade.
- Some hormones enter cells, interact with DNA, stimulate transcription.
- Hormones regulate and coordinate changes in gene expression.
How can mRNA processing be regulated?
- U1A protein’s regulation involves inhibition of its pre-mRNA polyadenylation.
- U1A protein binds to its pre-mRNA, inhibits polyadenylation.
- Binding prevents mRNA processing, and mRNA doesn’t mature
