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What is the primary role of Ran-GEF in nuclear transport?

  • To bind to the nuclear export signal (NES)
  • To hydrolyze GTP in the cytoplasm
  • To stimulate the conversion of Ran-GDP to Ran-GTP (correct)
  • To facilitate the binding of exportins to mRNP complexes

    • What occurs after the exportin1 and Ran-GTP complex enters the cytoplasm?

  • Exportin1 dissociates from Ran-GDP
  • The complex binds to FG domains in the nuclear pore complex
  • The cargo protein is transported back to the nucleus
  • Ran-GAP stimulates hydrolysis of GTP, releasing the cargo (correct)

    • What is required for proteins to exit the nucleus?

  • Binding to Ran-GDP
  • Association with mRNP complexes
  • A nuclear export signal (NES) (correct)
  • A nuclear localization signal (NLS)

    • Which component is involved in mRNP export in a Ran-independent mechanism?

    <p>NXFI</p> Signup and view all the answers

    What facilitates diffusion across the nuclear pore complex (NPC) for mRNA?

    <p>Complex formation with FG repeats</p> Signup and view all the answers

    What is the primary function of the 5' cap added to nascent mRNA?

    <p>It protects the mRNA and aids in nuclear export.</p> Signup and view all the answers

    Which factor is directly responsible for recognizing the Poly(A) signal during 3’ cleavage and polyadenylation?

    <p>Cleavage and polyadenylation specificity factor (CPSF)</p> Signup and view all the answers

    What occurs during the first step of mRNA splicing?

    <p>U1 and U2 interact with mRNA.</p> Signup and view all the answers

    What happens during the 'slow phase' of polyadenylation?

    <p>12 A residues are added to the 3' end.</p> Signup and view all the answers

    Which sequence represents the Poly(A) signal recognized during 3’ cleavage and polyadenylation?

    <p>AAUAAA followed by G/U</p> Signup and view all the answers

    What is NOT a role of the 5' cap in mRNA processing?

    <p>Initiating RNA replication.</p> Signup and view all the answers

    During mRNA splicing, what does the OH at the branch point do?

    <p>Attacks the 5' phosphate of the first intron residue.</p> Signup and view all the answers

    Which of the following components is NOT involved in the recruitment process during mRNA splicing?

    <p>Poly A Polymerase</p> Signup and view all the answers

    What role does U2AF play in mRNA splicing?

    <p>It promotes splicing efficiency.</p> Signup and view all the answers

    What is the primary outcome of alternative splicing in relation to a gene?

    <p>It allows for multiple protein isoforms from a single gene.</p> Signup and view all the answers

    Which editing change in the apoB pre-mRNA is specifically noted in intestinal cells?

    <p>Cytosine to Uracil conversion</p> Signup and view all the answers

    How does the presence of Sxl protein influence splicing in female Drosophila embryos?

    <p>It ensures the production of a functional transformer protein.</p> Signup and view all the answers

    What is the role of the Nuclear Transport Receptor in the nuclear import process?

    <p>It binds to the nuclear localization signal (NLS) on cargo proteins.</p> Signup and view all the answers

    What is the effect of early Sxl protein absence in male Drosophila regarding sex-lethal mRNA splicing?

    <p>A stop codon is introduced into the spliced mRNA.</p> Signup and view all the answers

    What structures within the Nuclear Pore Complex (NPC) form the permeability barrier?

    <p>Phe-Gly repeats and hydrophilic regions</p> Signup and view all the answers

    What prevents the usage of splice sites in female Drosophila during early development?

    <p>Exonic splicing silencers attached to Sxl protein.</p> Signup and view all the answers

    Which components are involved in the production of female-specific Dsx protein?

    <p>A complex of tra, Rbp1, and tra2.</p> Signup and view all the answers

    What happens to the Ran-GTP after it aids in the release of the cargo protein in the nucleus?

    <p>It is hydrolyzed to GDP by Ran-GAP.</p> Signup and view all the answers

    Which statement regarding self-splicing introns is true?

    <p>They can splice themselves without external assistance.</p> Signup and view all the answers

    Which of the following statements is true regarding the size of molecules that can freely diffuse through the nuclear pore complex?

    <p>Molecules up to 60kDA can freely diffuse.</p> Signup and view all the answers

    In the nuclear import process, what is the first step taken by free importin when it encounters a cargo protein?

    <p>It binds to the NLS on the cargo protein.</p> Signup and view all the answers

    What is the primary function of exonic splicing enhancers (ESEs)?

    <p>They promote splicing by binding to SR proteins.</p> Signup and view all the answers

    What is formed during the transport of molecules through the nuclear pore complex?

    <p>A protein complex with FG repeats</p> Signup and view all the answers

    What type of domains do proteins need to possess to enter the nucleus?

    <p>Nuclear localization signals (NLS)</p> Signup and view all the answers

    What is the primary sequence recognized as the Poly(A) signal during the cleavage and polyadenylation of pre-mRNA?

    <p>AAUAAA</p> Signup and view all the answers

    Which factor is primarily responsible for adding adenine nucleotides to the cleaved 3' end of pre-mRNA?

    <p>PAP</p> Signup and view all the answers

    What is the immediate consequence of the 3' cleavage of pre-mRNA during polyadenylation?

    <p>Recognition by PAP</p> Signup and view all the answers

    What role does the Poly(A) tail serve in mRNA molecules?

    <p>It stabilizes the mRNA and assists in transport</p> Signup and view all the answers

    What is the role of U1 in the splicing process?

    <p>It aids in the recognition of the 5' splice site.</p> Signup and view all the answers

    Which component within the splicing machinery is responsible for binding to exonic splicing enhancers (ESEs)?

    <p>SR Proteins</p> Signup and view all the answers

    What role does the branch point play in mRNA splicing?

    <p>It is essential for the catalytic activity of the spliceosome.</p> Signup and view all the answers

    Which statement accurately describes the function of U2AF in splicing?

    <p>It increases the efficiency of spliceosome assembly.</p> Signup and view all the answers

    How do SR proteins contribute to the splicing process?

    <p>They mediate cooperative binding of splicing factors.</p> Signup and view all the answers

    What is the main role of the Sxl protein in Drosophila during early development?

    <p>It blocks the use of splice sites in female embryos.</p> Signup and view all the answers

    Which genes are likely expressed differently between male and female Drosophila?

    <p>Sxl and Dsx</p> Signup and view all the answers

    What happens to the sex-lethal gene as Drosophila develop further?

    <p>It is activated in both males and females.</p> Signup and view all the answers

    What is the significance of the intronic splicing silencer in the context of Sxl binding?

    <p>It blocks female-specific splice site usage.</p> Signup and view all the answers

    Which of the following proteins is involved in the somatic sex determination pathway of Drosophila?

    <p>Tra2</p> Signup and view all the answers

    What effect does early Sxl protein have on exon 3 of the sex-lethal mRNA in female Drosophila?

    <p>Exon 3 is skipped during splicing.</p> Signup and view all the answers

    Which protein's production is directly influenced by the binding of Sxl to ISS in female Drosophila?

    <p>Tra</p> Signup and view all the answers

    What is the primary consequence of skipping exon 2 during the splicing of sex-lethal mRNA in female Drosophila?

    <p>Formation of functional transformer protein.</p> Signup and view all the answers

    What is the importance of alternative splicing in the sexual differentiation of Drosophila?

    <p>It generates different protein variants that determine sex.</p> Signup and view all the answers

    What occurs when Sxl protein is absent in male Drosophila during sex-lethal mRNA splicing?

    <p>The mRNA contains a stop codon due to alternative splicing.</p> Signup and view all the answers

    Which protein is directly affected by the splicing of the transformer mRNA in Drosophila males?

    <p>Tra protein</p> Signup and view all the answers

    What is the role of the Sxl protein in female Drosophila?

    <p>To promote the production of functional transformer and doublesex proteins.</p> Signup and view all the answers

    Which gene’s splicing results in determining the developmental pathway of Drosophila based on the Sxl protein presence?

    <p>All of the above</p> Signup and view all the answers

    What is the primary outcome of alternative splicing in the context of Drosophila sex determination?

    <p>The generation of multiple mRNA isoforms from the same gene.</p> Signup and view all the answers

    What characterizes the mRNA differences resulting from RNA editing?

    <p>The mature mRNA sequence can differ from the DNA coding sequence.</p> Signup and view all the answers

    Which of the following edits is specifically noted in the editing of apoB pre-mRNA in intestinal cells?

    <p>C to U conversion</p> Signup and view all the answers

    What distinguishes apoB-48 from apoB-100?

    <p>apoB-48 has a shorter amino acid length compared to apoB-100.</p> Signup and view all the answers

    In which of the following locations is apoB-100 produced?

    <p>Liver</p> Signup and view all the answers

    What role does the N-terminal domain of apoB-48 play?

    <p>It associates with lipids.</p> Signup and view all the answers

    What is the primary function of importin in the nuclear import process?

    <p>To bind cargo proteins containing a nuclear localization signal</p> Signup and view all the answers

    What triggers the release of cargo protein from the importin complex inside the nucleus?

    <p>The interaction of importin with Ran-GTP</p> Signup and view all the answers

    Which step follows the release of cargo protein from the importin complex in the nuclear import pathway?

    <p>Ran-GTP is hydrolized to Ran-GDP</p> Signup and view all the answers

    What is the structure through which the importin-cargo complex passes during nuclear transport?

    <p>Nuclear pore complex (NPC)</p> Signup and view all the answers

    What role does Ran-GDP play after releasing importin from the importin-Ran-GTP complex?

    <p>It returns to the nucleus to exchange GDP for GTP.</p> Signup and view all the answers

    What is the result of the combination of tra, Rbp1, and tra2 proteins in female Drosophila?

    <p>It results in a short, female-specific Dsx protein.</p> Signup and view all the answers

    In male Drosophila, what happens to exon 4 during Dsx protein production?

    <p>Exon 4 is skipped entirely.</p> Signup and view all the answers

    What molecular process occurs at the 3' end of exon 4 during female Dsx protein production?

    <p>Exon 4 is polyadenylated.</p> Signup and view all the answers

    Which of the following accurately describes the outcome of alternative splicing in Drosophila?

    <p>It generates proteins with varying lengths between sexes.</p> Signup and view all the answers

    What is the role of the exotic splicing enhancer in female Drosophila?

    <p>It acts as a binding site for tra proteins.</p> Signup and view all the answers

    What is the main role of the NXF1 protein in mRNP export?

    <p>It serves as the export factor that binds to the mRNP complex.</p> Signup and view all the answers

    Which two components are primarily involved in remolding the mRNP complex for export?

    <p>NXF1 and NXT1</p> Signup and view all the answers

    What is the primary method by which the mRNP complex crosses the nuclear pore complex (NPC)?

    <p>Diffusion facilitated by FG repeats</p> Signup and view all the answers

    What happens to the mRNA once it reaches the cytoplasm during the export process?

    <p>It is released from the mRNP complex.</p> Signup and view all the answers

    In the context of mRNP export, what is the function of the small NXT1 subunit?

    <p>It functions as the export transporter facilitating transport.</p> Signup and view all the answers

    What is the role of the mRNP exporter in the export process of mRNA?

    <p>To bind specific mRNP proteins associated with pre-mRNAs</p> Signup and view all the answers

    Which components make up the mRNP exporter complex?

    <p>NXF1 and NXT1</p> Signup and view all the answers

    What occurs during the remodeling phase of the mRNP complex?

    <p>NXF1 and NXT1 bind to the mRNP complex</p> Signup and view all the answers

    How does the mRNP complex interact with the nuclear pore complex (NPC)?

    <p>It interacts through FG repeats for diffusion</p> Signup and view all the answers

    What happens to the processed mRNA after it passes through the nuclear pore complex?

    <p>It is released from the mRNP complex</p> Signup and view all the answers

    Study Notes

    RNA Processing II & Nucleocytoplasmic Transport

    • The lecture is about RNA processing and nucleocytoplasmic transport.
    • The date of the lecture is November 20th, 2024.
    • The course is BIOL200.
    • The presenter is Holly J.

    Housekeeping

    • There is no lecture on December 4th.
    • The instructor will review the course material at the end of the term.
    • Students must submit questions by December 3rd at noon.
    • All sessions will be recorded.
    • Regular tutorials/lectures will resume, but tutorials will be one week behind schedule.

    mRNA Processing

    • Modifications at the 5' and 3' ends of pre-mRNA are necessary for stability and protection.
    • Three main co-transcriptional steps are capping, cleavage and polyadenylation, and splicing.

    5' Capping

    • Nascent mRNA emerges from the RNA exit channel.
    • A methylguanalate cap is added by a capping enzyme.
    • This enzyme interacts with RNA polymerase II CTD.
    • Capping protects the mRNA and facilitates nuclear export; it leads to recognition by translation initiation factors.
    • Relevant proteins include Serine 2 phosphorylation, splicing factors, polyadenylation factors, export factors, and capping enzymes.
    • Relevant enzymes include capping enzyme, RNA Pol II, and TFIIH.

    3' Cleavage & Polyadenylation

    • A poly(A) signal (AAUAAA + G/U) is recognized by cleavage and polyadenylation factors (CPSF, CStF, CFI, and CFII).
    • The 3' end of the pre-mRNA is cleaved.
    • The 3' end is recognized by poly A polymerase (PAP).
    • A slow phase adds 12 A residues to the 3' end of the pre-mRNA.
    • A rapid phase adds ~200 A residues using PolyA Binding Protein (PABP).
    • Enzymes involved are PAP, CPSF, CStF, CFI, and CFII.

    mRNA Splicing

    • U1 (intron splice site) and U2 (branch point) interact with mRNA.
    • U4, U5, and U6 are recruited.
    • U1 and U4 exit the spliceosome.
    • Transesterification reactions occur.
    • The OH at the branch point attacks the 5' phosphate at the first intron residue to form a lariat.
    • The 3' end of the exon attacks the 5' end of the following exon.
    • Mature mRNA, snRNAs, and intron are released.
    • Relevant enzymes include various PRPF enzymes.

    Self-Splicing Introns

    • In some cases, introns self-splice without external snRNAs.
    • Group II introns are found in mitochondria and chloroplasts and may be the evolutionary predecessors of other introns.

    mRNA Splicing Specificity

    • The exon cross recognition complex helps with splicing efficiency.
    • U2AF helps with splicing efficiency.
    • Spliceosome is involved; U1, U2, U2AF65, Branch point, 3' splice site, 5' splice site, , and cross-exon recognition complex.
    • SR proteins contain RRM domains and arginine/serine-rich domains; they mediate cooperative binding of U1 to the 5' splice site and U2 to the branch point.
    • ESES (exonic splicing enhancers), sequences within the exons, promote splicing.
    • Key enzymes involved are U2AF, and various PRPF enzymes.

    Alternative Splicing

    • The presence of multiple introns allows for multiple variations of a protein from a single gene.
    • Results in protein isoforms.

    Alternative Splicing Ex: Sexual Differentiation in Drosophila

    • Sxl protein is only found in female embryos during early development.
    • In males, splicing results in an mRNA containing a stop codon, leading to no functional Sxl protein.
    • Later in development, the sex-lethal (Sxl) gene is active in both sexes.
    • In males, splicing of tra exon 1 to exon 2 leads to no functional transformer protein.
    • In females, exon 3 of sex-lethal mRNA is skipped, leading to functional Sxl protein, binding of Sxl to a splicing silencer.
    • Diagrams show the location of exons and introns in the mRNA sequences; these diagrams depict splicing in both males and females, including specific components and proteins involved for each sex-determining pathway stages. Diagrams for tra and dsx pre-mRNAs and mRNAs, detailing the difference in splicing patterns based on the presence/absence of Sxl protein.

    RNA Editing

    • The mature mRNA sequence sometimes differs from DNA.
    • Editing mechanisms include Adenosine to Inosine (A -> I) and Cytosine to Uracil (C -> U).
    • Editing of apoB pre-mRNA changes C->U position 6666 in intestinal cells; this leads to a shorter protein isoform, apoB-48, with N-terminal domains that associate with lipids. The diagram shows the location of the changes in the mRNA sequence.

    Nucleo-Cytoplasmic Transport

    • This section covers transport between the nucleus and cytoplasm.

    Nuclear Pore Complex

    • Proteins need to enter and exit the nucleus.
    • The nuclear pore complex (NPC) is made of nucleoporins, creating a basket and aqueous pore.
    • FG nucleoporins have extended disordered regions that form a permeability barrier.
    • Small molecules (<60 kDa) can diffuse freely, while larger molecules need active transport.

    Nuclear Import

    • Proteins with a nuclear localization signal (NLS) require importins to enter the nucleus.
    • Importins bind to the NLS on the cargo protein, forming a complex.
    • The importin-cargo complex interacts with the FG repeats in the NPC.
    • Once in the nucleus, importin interactions with Ran-GTP cause cargo release.
    • Importin and Ran-GTP return to the cytoplasm to repeat the cycle.
    • Ran proteins, GTP/GDP, Ran-GEF, and Ran-GAP facilitate this process. Specific proteins include: Importins, cargo proteins, Ran-GTP, Ran-GDP, Ran-GEF, and Ran-GAP. Diagrams illustrate the importin/Ran cycle. Ran-GTP.

    Nuclear Export

    • Proteins with a nuclear export signal (NES) require exportins to leave the nucleus.
    • Exportins bind to the NES on the cargo protein, forming a complex.
    • The exportin-cargo complex interacts with the FG repeats of the NPC.
    • Ran-GAP stimulates hydrolysis of GTP to GDP.
    • Cargo is released.
    • Exportin and Ran-GDP return to the nucleus. Key components include Ran-GTP/GDP, Ran-GEF and Ran-GAP.

    mRNP Export

    • mRNPs are exported from the nucleus using a Ran-independent mechanism.
    • mRNP exporter proteins (e.g., NXF1 and NXT1) bind to RNA that has associated pre-mRNA proteins; mRNA binding is cooperative.
    • The exporter cooperatively binds to RNA and forms a domain that interacts with FG repeats of the NPC; thus, the proteins diffuse through the NPC.
    • Key components include mRNP exporter complex (NXF1 with NXT1 subunit), ribosome, capping site, and PolyA tail. Diagrams illustrate the components involved in the mRNP export process; arrows show the direction of movement for the proteins involved in the export process. mRNPs associate with proteins and other RNAs in the nucleus. Large proteins (e.g. NXF1) likely play a role in export.

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    RNA Processing II & Nucleocytoplasmic Transport PDF

    Description

    Test your understanding of RNA processing and nucleocytoplasmic transport as discussed in the BIOL200 course. This quiz covers key concepts such as mRNA modifications, capping, and the role of enzymes in RNA stability. Get ready to assess your knowledge on these essential biological processes.

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