Review of L19-20-DNA TRANSCRIPTION & TRANSLATION

THE NATURE OF GENES

  • Beadle and Tatum's Work

    • Demonstrated that genes specify enzymes.

    • Studied Neurospora mutants unable to synthesize arginine, revealing absence of specific enzymes.

    • Proposed the "one gene/one polypeptide" hypothesis.

  • Central Dogma of Molecular Biology

    • Information flow in cells: DNA → RNA → Protein.

    • DNA strand copied to mRNA is the template (antisense) strand; the other is the coding (sense) strand.

    • Transcription: RNA copy of DNA is created.

    • Translation: Uses RNA information to synthesize proteins.

    • Transfer RNA (tRNA) acts as an adapter to connect mRNA information to amino acid sequences.

    • RNA has multiple roles in gene expression.

THE GENETIC CODE

  • Structure and Function of the Code

    • Read in groups of three nucleotides (codons).

    • Nonoverlapping, established by Crick and Brenner.

    • A codon consisting of 3 nucleotides leads to 64 possible codons.

  • Codon Functions

    • Three codons signal "stop"; one codon signals "start" and also encodes methionine (AUG).

    • 61 codons encode 20 different amino acids.

    • The code is degenerate (many amino acids have multiple codons) but specific (each codon specifies only one amino acid).

  • Universality of the Code

    • Practically universal, with exceptions in mitochondrial and protist genomes where stop codons can function as amino acids.

PROKARYOTIC TRANSCRIPTION

  • Overview of Prokaryotic RNA Polymerase

    • Contains single RNA polymerase.

    • Forms: core polymerase (synthesizes mRNA) and holoenzyme (core + σ factor for accurate initiation).

    • Initiation occurs at promoters, which are crucial for proper RNA polymerase binding.

  • Process of Transcription

    • Initiation: Requires start site and promoter located upstream.

    • Binding of the holoenzyme to the –35 region positions RNA polymerase.

    • Elongation: Addition of nucleotides in the 5′-to-3′ direction within the transcription bubble (contains RNA polymerase, unwound DNA, and growing mRNA).

    • Termination: Specific sites where termination occurs through double-stranded hairpin formation, causing polymerase to pause.

    • Coupling of Transcription and Translation: Translation can start while mRNA is still being transcribed.

EUKARYOTIC TRANSCRIPTION

  • RNA Polymerases in Eukaryotes

    • Three forms: RNA polymerase I (rRNA), polymerase II (mRNA, some snRNAs), and polymerase III (tRNA).

    • Core promoter and general transcription factors essential for recruitment of RNA Pol II to form initiation complex.

  • Transcription Process

    • Complex formation includes binding factors for RNA Pol II.

    • Capping occurs around 20 nucleotides in, with polymerase possibly pausing needing elongation factors for resumption.

    • Carboxy terminal domain (CTD) interacts with elongation factors and modifying enzymes.

    • Termination involves polyadenylation where poly-A polymerase adds adenine residues to the 3′ end of the mRNA transcript.

    • Primary transcripts undergo modifications to yield mature mRNAs (addition of a 5′ cap and 3′ poly-A tail, removal of introns).

EUKARYOTIC PRE-MRNA SPLICING

  • Splicing Overview

    • Eukaryotic genes feature coding (exons) and noncoding (introns) sequences.

    • Introns are removed via splicing by spliceosomes.

  • Splicing Mechanism

    • snRNPs recognize intron-exon junctions to recruit spliceosomes.

    • The spliceosome joins exons, resulting in multiple potential transcripts from one gene.

THE STRUCTURE OF tRNA AND RIBOSOMES

  • tRNA Functionality

    • Aminoacyl-tRNA synthetases attach specific amino acids to tRNA.

    • The tRNA charging reaction couples amino acids to the 3′ end of tRNA.

  • Ribosomal Structure

    • Ribosome comprises multiple tRNA-binding sites: A site for initial binding, P site for peptide bond formation, E site for tRNA release.

    • Ribosomes facilitate both decoding (reading mRNA) and enzymatic functions (forming peptide bonds).

THE PROCESS OF TRANSLATION

  • Translation Phases

    • Initiation: Requires accessory factors; in prokaryotes, the ribosome-binding sequence of mRNA is essential; in eukaryotes, the 5′ cap is utilized.

    • Elongation: New amino acids added as the ribosome traverses mRNA.

    • Termination: Recognizes stop codons through termination factors; proteins may be targeted to the endoplasmic reticulum (ER).

SUMMARIZING GENE EXPRESSION

MUTATION: ALTERED GENES

  • Types of Mutations

    • Point mutations lead to single-nucleotide variations (SNVs).

    • Base substitutions can either have no effect (silent) or alter encoded amino acids (missense, nonsense).

  • Indels and Structural Variations

    • Insertions/deletions (indels) affect 1-50 bp, causing frameshifts if not multiples of 3, potentially leading to shortened proteins.

    • Triplet repeat expansions linked to neurodegenerative disorders.

    • Structural variations (SV) result from chromosomal mutations (additions, deletions, inversions, translocations).

  • Role of Mutations

    • Mutations are vital as starting points for evolution.

    • Human mutation rates approximately measure at 70 new mutations per generation.