L17 Gene Structure and mRNA Processing

Page 1

  • Gene structure: Introns and exons in a gene (example shown). Exons are shorter segments interspersed by introns; overall gene length can be large (e.g., hundreds of thousands of nucleotides in some cases).

  • Exon positions and lengths (illustrative): exons occur at several positions (e.g., around 1, 5, 10, 14, 22, 25, 26) with exon lengths depicted up to roughly 10001000 nucleotides in the example.

  • Intron sizes vary widely between species (human, worm, fly). Intron length distribution spans small to very large values (categories shown such as 100100, 20002000, 50005000, and >30{,}000 nucleotides).

  • Frameshift example (translation impact): A normal translation vs. a frameshift (either +1+1 or −1-1) drastically alters the amino-acid sequence downstream of the mutation. The slide shows representative translations under normal, +1 frameshift, and -1 frameshift conditions, illustrating how frameshifts modify codon interpretation.

  • Each set of 3 nucleotides, known as a codon, corresponds to a specific amino acid or a stop signal, which is crucial for accurate protein synthesis. Therefore, any frameshift mutation within a coding region can lead to an entirely different protein product, potentially resulting in loss of function or gain of a harmful trait.

  • Concept takeaway: Exon–intron architecture and intron length diversity influence gene regulation and evolution; frameshift mutations disrupt the reading frame and change the protein sequence.

  • Introns are - Intervening Sequence

  • Exons -Expressed Sequence

Page 2

  • Three consensus sequences for locating intron/exon boundaries for splicing are presented (note: the exact sequences in the slide text are garbled in the transcript, but the key concept is preserved):

    • 5' splice site (donor) boundary signals

    • Branch point signal within the intron (A residue with a nearby pyrimidine tract)

    • 3' splice site (acceptor) boundary signals

  • Structural markers shown in the slide include: 5' exon boundary, 3' exon boundary, Branch point, 5' splice site, 3' splice site, Left (5) site, Right (3) site, and a pyrimidine-rich region (12Py) before the branch point. These signals together guide intron excision.

  • Practical note: While exact nucleotide strings may vary, splicing relies on a 5' donor site, a branch point A, and a 3' acceptor site.


Page 3

  • snRNPs and the spliceosome components:

    • U1 binds to the 5' splice site (donor).

    • U2 binds to the branch point within the intron.

    • U4, U5, and U6 form a complex with U1 and U2 to create the inactive spliceosome.

  • Activation: The complex rearranges to form an active spliceosome that carries out two transesterification steps to remove the intron and join exons.

Page 4

  • Splicing mechanism overview (intron removal and exon joining):

    • First transesterification: the 2' hydroxyl of the branch-point A attacks the 5' splice site, creating a lariat structure and cleaving the 5' exon.

    • Second transesterification: the 3' splice site is cleaved, and the 5' exon is joined to the 3' exon, yielding the spliced mRNA and the excised intron in a lariat form.

  • Key components during this process: U1, U2, U4, U5, U6 participate in different steps; after splicing, U1 and U4 are released, and the intron lariat along with U2, U5, and U6 is degraded. The snRNPs are recycled for future splicing.

  • Outcome: Exons are covalently linked; intron (as a lariat) is removed.

Page 5

  • mRNA coding and processing regions:

    • Coding segment (open reading frame).

    • Start codon and stop codon mark the beginning and end of translation.

    • 5' Cap and 3' Poly(A) tail are added during processing; the leader (5' untranslated region) and trailer (3' untranslated region) flank the coding sequence.

    • Termination signal at the mRNA level is associated with the stop codon.

  • 3' end processing signals:

    • Polyadenylation signal in the transcript is typically AAUAAAAAUAAA, guiding poly(A) tail addition.

  • Overall, mature mRNA contains: 5' Cap, Leader (5' UTR), coding sequence, Stop codon, Trailer (3' UTR), and Poly(A) tail.

Page 6

  • Coupling of transcription and RNA processing via the CTD of RNA polymerase II:

  • The mechanics of Splicing

    • The C-terminal domain (CTD) of RNA polymerase II coordinates recruitment of processing factors during transcription.

    • Enzymes/factors associated with the CTD include: capping enzymes, initiation factors, elongation factors, splicing factors, termination factors, and polyadenylation factors.

    • Histone mark connection: SET1 and the H3K4me3 mark are associated with active transcription and CTD-mediated recruitment of processing machinery.

    • This coupling ensures that capping, splicing, and 3' end processing occur co-transcriptionally and in a coordinated manner.