BIOL200 RNA Processing II and Transport

Podcast

Play an AI-generated podcast conversation about this lesson

Questions and Answers

What is the role of Ran-GTP in nuclear export?

It is hydrolyzed in the nucleus to release cargo proteins.

It facilitates the transport of mRNA through the cytoplasm.

It directly transports proteins out of the cell.

It binds to exportin1 to increase affinity for cargo protein. (correct)

What must proteins possess to be exported from the nucleus?

A nuclear localization signal (NLS).

A promoter binding region.

A ribosomal binding site (RBS).

A nuclear export signal (NES). (correct)

Which component stimulates the hydrolysis of GTP in the cytoplasm?

Ran-GEF.

Ran-GAP. (correct)

Free exportin1.

Export transporter.

What is the function of mRNP in nuclear export?

It binds to FG repeats for transport across the nuclear pore complex. Signup and view all the answers

How does exportin1 recognize the cargo protein for export?

By forming a complex with Ran-GTP in the nucleus. Signup and view all the answers

What is the role of the 5' cap in mRNA processing?

Protects mRNA from degradation Signup and view all the answers

Which sequence is recognized by cleavage and polyadenylation factors during mRNA processing?

Poly(A) signal Signup and view all the answers

During mRNA splicing, which components interact first with the mRNA?

U1 and U2 snRNPs Signup and view all the answers

What occurs during the slow phase of polyadenylation?

Addition of 12 A residues Signup and view all the answers

Which step occurs right after the cleavage of the 3' end during polyadenylation?

3' end is recognized by Poly A Polymerase Signup and view all the answers

What is the primary function of the proteins CPSF and CStF in mRNA processing?

Recognition of the poly(A) signal Signup and view all the answers

What is the result of the transesterification reactions during splicing?

Formation of the lariat structure and joined exons Signup and view all the answers

Which component exits the spliceosome during the splicing process?

U1 and U4 snRNPs Signup and view all the answers

What is the function of the U2AF protein in mRNA splicing?

It helps with splicing efficiency. Signup and view all the answers

Which of the following is true about self-splicing introns?

They can splice themselves independently. Signup and view all the answers

How does the presence of multiple introns impact gene expression?

It allows for the creation of multiple protein isoforms. Signup and view all the answers

In Drosophila, the presence of the Sxl protein in females leads to which splicing outcome?

Exon 3 of the sex-lethal mRNA is skipped. Signup and view all the answers

What role do exonic splicing enhancers (ESEs) play in mRNA splicing?

They are sequences within the exons that promote splicing Signup and view all the answers

What is a consequence of the absence of early Sxl protein in male Drosophila?

The mRNA contains a stop codon. Signup and view all the answers

How does the splicing mechanism differ between female and male Drosophila regarding the Dsx protein?

Females produce a shorter, female-specific Dsx protein. Signup and view all the answers

Which statement is true regarding RNA editing as mentioned in the content?

It alters nucleotide sequences post-transcriptionally. Signup and view all the answers

What is the primary role of the nuclear pore complex (NPC)?

Facilitating the transport of molecules between the nucleus and cytoplasm Signup and view all the answers

Which modification involves the conversion of Cytosine to Uracil?

Editing of pre-mRNA Signup and view all the answers

What is required for proteins to enter the nucleus?

A nuclear localization signal (NLS) Signup and view all the answers

What happens to Ran-GTP after it interacts with importin in the nucleus?

It hydrolyzes GTP to GDP, promoting the release of importin Signup and view all the answers

At what molecular weight do molecules start requiring transport through the nuclear pore complex?

60 kDA Signup and view all the answers

What type of molecules freely diffuse through the nuclear pore complex?

Molecules up to 60 kDA Signup and view all the answers

What is the role of FG nucleoporins in the nuclear pore complex?

To form a permeability barrier Signup and view all the answers

Which of the following is a characteristic of the amino acid sequence of importin?

Interacts with FG repeats for transportation Signup and view all the answers

Study Notes

RNA Processing II & Nucleocytoplasmic Transport

RNA Processing II and Nucleocytoplasmic Transport lecture
Lecture is PL2/3
Lecturer: Holly J
Course: BIOL200
Date: November 20th, 2024

Housekeeping

No lecture on December 4th
End-of-term instructor review
Submit questions by December 3rd at noon
Sessions will be recorded
Tutorials are one week behind the lecture schedule

mRNA Processing

Modifications at the 5' and 3' ends of pre-mRNA are crucial for stability and protection
Three major co-transcriptional steps:
- 5' capping
- 3' cleavage and polyadenylation
- RNA splicing

5' Capping

Nascent mRNA emerges from the RNA exit channel (25bps)
A methylguanalate cap is added by a capping enzyme that interacts with RNA Pol II CTD
Protects the mRNA, facilitates nuclear export, allows recognition by translation initiation factors

3' Cleavage & Polyadenylation

Poly(A) signal = AAUAAA, + G/U
Polyadenylation factors (CPSF, CStF, CFI, CFII) recognize the signal
3' end is cleaved
3' end is recognized by Poly A Polymerase (PAP)
Slow Phase: 12 A residues added to 3' end
Rapid Phase: PolyA Binding Protein (PABP) adds around 200 A residues

mRNA Splicing

U1 (intron splice site) and U2 (branch point) interact with mRNA
U4, U5, U6 are recruited
U1 & U4 exit (active spliceosome)
Transesterification Reactions
- OH at branch point attacks 5' phosphate at the 1st intron residue, forming a lariat
- 3' end of exon attacks 5' end of following exon
Release of mature mRNA, snRNAs, and intron
Self-splicing introns: in some cases, introns have the capacity to self-splice without external snRNAs

mRNA Splicing Specificity

Exon cross recognition complex
U2AF: Important for splicing efficiency
SR Proteins: Contain RRM domains and arginine/serine rich domains
Cooperative binding of U1 to the 5' splice site and U2 to the branch point
Binding to exonic splicing enhancers (ESEs)
ESES: sequences within exons that promote splicing

Alternative Splicing

The presence of multiple introns allows for multiple variations of a protein from a single gene (protein isoforms)
Example: Sexual differentiation in Drosophila
- Sxl protein is only present in female embryos (early development)
- Binding site (intronic splicing silencer) blocks use of splice site
- Later in development, the gene is active in males and females
- Males:
  - Absence of early Sxl protein => sex-lethal mRNA is spliced to make mRNA with a stop codon => no functional Sxl protein
  - No functional transformer protein
- Females:
  - Presence of early Sxl protein => exon 3 is skipped => functional Sxl protein => binding of Sxl to ISS skips exon 2 => functional transformer protein
Dsx protein (double-sex gene)
- Females: complex of tra, Rbp1 and tra2 direct splicing, cleavage, and alt polyadenylation => short, female-specific Dsx
- Binding site = exotic splicing enhancer
- Males: no tra => exon 4 is skipped => longer, male-specific Dsx

RNA Editing

Sequence of mature mRNA sometimes differs from the DNA's coding regions sequence
Adenosine to Inosine (A -> I) OR Cytosine to Uracil (C -> U)
Ex: Editing of apoB pre-mRNA changes C->U at position 666 in intestinal cells
Shorter apoB-48 has N-terminal domains associated with lipids

Nucleo-Cytoplasmic Transport

Nuclear Pore Complex

Proteins synthesized in the cytoplasm need to exit the nucleus (and potentially re-enter)
Nuclear Pore Complex (NPC): made up of nucleoporins
- Nuclear basket
- Aqueous pore
- FG nucleoporins: extended regions (Phe-Gly repeats + hydrophilic regions) form permeability barrier
- Molecules up to 60kDA freely diffuse; larger molecules need to be transported

Nuclear Import

Proteins wanting to enter the nucleus require a nuclear localization signal (NLS)
Free importin binds to NLS on cargo protein, forming a complex
Complex interacts with FG repeats in NPC channel
Importin interacts with Ran-GTP causes cargo protein release (decreased affinity)
Importin-Ran-GTP complex diffuses back to cytoplasm
Ran interacts with Ran-GAP, hydrolyzes GTP to GDP, importin is released
Ran-GDP returns to nucleus, interacts with Ran-GEF, causing GDP to be exchanged for GTP

Nuclear Export

Proteins wanting to exit the nucleus require a nuclear export signal (NES)
Free exportin1 forms complex with Ran-GTP and cargo protein (increased affinity)
Complex interacts with FG repeats in NPC channel
In cytoplasm, complex encounters Ran-GAP, stimulating GTP hydrolysis, cargo is released
Exportin1 and Ran-GDP transported back to nucleus
Ran-GEF stimulates conversion of Ran-GDP to GTP

mRNP Export

Once mRNA processing is complete, it remains associated with proteins (mRNP complex)
Exporter binds RNA with mRNP proteins during elongation (prior to processing)
Forms a domain that interacts with FG repeats, allowing diffusion across NPC

Questions?

Studying That Suits You

Use AI to generate personalized quizzes and flashcards to suit your learning preferences.

Description

Explore the essential processes of RNA processing and nucleocytoplasmic transport in this BIOL200 lecture by Holly J. Understand mRNA modifications, including 5' capping and 3' polyadenylation, that are vital for mRNA stability and functionality. This quiz will test your knowledge of the intricacies involved in these cellular mechanisms.