Genetics Chapter on Gene Expression and Splicing

Course Information

• Course: BMS 141
• Lecture Number: 9
• Title: Post-transcription modification, Genetic codon
• Instructor: Dr. Lames Dawood
• Program: Medicine and Surgery
• Year: Fall 2024
• University: Galala University, Powered by Arizona State University

Intended Learning Outcomes

• Posttranscriptional modifications of mRNA
• Posttranscriptional modifications of rRNA
• Posttranscriptional modifications of tRNA
• Define genetic codon
• Enumerate different types of mutations
• Explain the effect of different mutations on phenotypes

Post-transcriptional Processing of mRNA

In Prokaryotes

• Little modification occurs in mRNA
• The primary transcript is immediately available for translation
• Can be mono-cistronic (one polypeptide chain) or poly-cistronic (many polypeptide chains)
• tRNA and rRNA primary transcripts are broken into smaller units

In Eukaryotes

• The primary transcript undergoes several modifications in the nucleus
• Capping of 5' end by GTP (followed by methylation, e.g., 7-methyl GTP)
• Removal of introns and splicing of exons
• Addition of a poly(A) tail at the 3' end

Capping of 5' end

• Enzyme: guanyltransferase
• Source of guanine: GTP
• Methylation follows in the cytosol (7-methyl GTP).
• Enzyme responsible for methylation: guanine-7-methyltransferase
• Source of methyl group: S-adenosylmethionine (SAM) (Active methionine)

Function of Cap

• Protects mRNA against 5' exonucleases
• Helps transport of mRNA to the cytoplasm
• Helps recognition of mRNA by ribosomes and initiation of translation
• Eukaryotic mRNA lacking the cap are not efficiently translated

Removal of Introns and Splicing of Exons

• Coding regions of DNA are called exons
• Exons are separated by non-coding regions called introns
• Pre-mRNA contains exons and introns
• Introns are removed, and exons are spliced together
• Histone mRNA does not contain introns
• snRNPs (snurps), which are part of spliceosomes (snRNA+protein), determine the splicing site

Alternative Splicing

• Leads to different protein products from the same gene
• Example: different muscle cell types produce tissue-specific tropomyosin proteins from same gene
• Different mRNAs can be generated from one primary transcript during mRNA processing.
• Example of alternative splicing: pre-mRNA for calcitonin produces different mRNAs in the thyroid gland and neurons (leading to different hormones)

Removal of Introns and Splicing of Exons - Clinical Significance

• Produces proteins with different activities
• Alternative splicing is involved in 60% of human genetic coding
• Provides a mechanism for producing a diverse set of proteins from a limited set of genes

Addition of Poly(A) Tail at 3' end

• Most eukaryotic mRNAs have a chain of 40-200 adenine nucleotides at the 3' end (poly(A) tail)
• This tail is not transcribed from the DNA
• Added after transcription by the enzyme, polyadenylate polymerase, using ATP.
• The mRNA is cleaved downstream of the consensus sequence (AAUAAA) and the poly(A) tail is added to the new 3' end.

Significance of Poly A tail

• Stabilizes mRNA
• Facilitates the exit of mRNA from the nucleus

Eukaryotic rRNA Processing

• Primary rRNA transcript (45S RNA) contains 18S, 5.8S and 28S rRNA
• RNA polymerase I transcribes rRNA genes to produce a single 45S precursor pre-rRNA
• Cleaving and processing of pre-rRNA in the nucleus yield 18S, 5.8S and 28S rRNA
• 5S rRNA gene is transcribed by RNA polymerase III

Synthesis of tRNAs

• Eukaryotic tRNA genes transcribed by RNA polymerase III
• pre-tRNA processing:
  • Removal of the extra 5' sequence (leader sequence)
  • Replacement of UU nucleotides with CCA at the 3' end
  • Modification of some bases (e.g., methylation)
  • Removal of a short intron in the anticodon loop

Genetic Code and Mutations

• Genome: All DNA in a cell
• Chromosome: Contains many genes
• Gene: A DNA sequence that specifies a polypeptide
• Genetic codon: A triplet of nucleotides that codes for an amino acid
• Information is based on a triplet code (codons)
• 64 possible codons (4^3)
• 61 codons specify amino acids
• 3 codons are stop codons (UAA, UAG, UGA)
• AUG is the initiator codon
• The ordering and grouping of nucleotides is the reading frame

Characteristics of the Genetic Code

• Specificity: Each codon codes for only one specific amino acid
• Universality: All organisms use the same genetic code (except mitochondria)
• Degeneracy: A given amino acid may have more than one codon (e.g., Leucine has 6 codons)
• Reading frame: Codons are read from the initiation codon as a continuous sequence of 3 bases

The Wobble Hypothesis

• Interaction between codon and anticodon needs to be exact at first two positions. Conventional base-pairing rules apply.
• The third base is not in close apposition and can allow non-traditional base-pairing (wobble).
• This allows codons for the same amino-acid to be recognized by multiple tRNAs/anticodons.

DNA Mutations

• Mutation: Permanent change in a gene's nucleotide sequence
• Mutagen: Physical or chemical agent that causes mutation
• Mutagenesis: Process of producing a mutation (spontaneous or induced)

Single Base Mutations (Point Mutations)

• Transition mutation: Changing a purine to another purine (A ↔ G) or a pyrimidine to another pyrimidine (C ↔ T).
• Transversion mutation: Changing a purine to a pyrimidine (A ↔ T or G ↔ C) or vice versa.
• Silent mutation: The resulting codon still codes for the same amino acid
• Missense mutation: The resulting codon codes for a different amino acid
• Nonsense mutation: The resulting codon codes for termination of the peptide chain
• Sense mutation: The terminating codon is changed to one that codes for an amino acid

Addition or Deletion of Nucleotides

• Addition or deletion of one or two bases results in a "frame-shift" mutation
• Addition or deletion of three nucleotides results in addition or deletion of one amino acid.

In-Class Assessment Questions

• Functions of poly(A) tail in mRNA (except)

• Functions of 5' mRNA capping (choose one)

• Systemic lupus erythromatosus results from (choose one option)

• Processing of tRNA (except)

Test your knowledge on gene expression, alternative splicing, and the roles of various RNA types in eukaryotic cells. This quiz covers essential topics such as mutations, the significance of poly(A) tails, and the complexities of splicing mechanisms. Perfect for students studying genetics and molecular biology.