Mutations and Transcription

Mutations and Allele Diversity

• Mutations lead to new alleles.

• Example: Mouse fur color changes from brown to black due to a mutation in the melanocortin receptor.

• This process introduces diversity in the alleles.

Types of Mutations

• Point Mutation:

  • Change in one single base. For Example A changes to T.

• Insertions:

  • Addition of a base, shifting everything down.

• Deletions:

  • Removal of a base, shifting everything up.

  • Insertions and deletions can involve a single base or a stretch of nucleotides.

• Chromosome-Level Mutations:

  • Affect parts or entire chromosomes.

  • Include missing or duplicated chromosome pieces.

  • Chromosome segments can be flipped or entire chromosomes added/removed.

Location of Mutations and Their Effects

• Mutations can occur anywhere in the genome.

• Coding Sequences: Affect amino acids and proteins.

• Regulatory Regions: Impact gene expression.

• Non-functional Regions: May have minimal consequences.

Types of Point Mutations

• Original sequence of DNA is transcribed into mRNA, then translated into amino acids.

• Silent Mutation:

  • A change in DNA that results in the same amino acid due to code redundancy.

  • Example: A base change still codes for tyrosine.

• Missense Mutation:

  • A base change that results in a different amino acid being coded.

  • Example: Tryptophan replaced by cysteine.

  • Impact varies depending on the significance of the amino acid change.

• Nonsense Mutation:

  • A base change that results in a stop codon.

  • Causes termination of translation at that position.

  • The impact depends on the stop codon's position; early stop codons are more detrimental.

• Frameshift Mutation:

  • Insertions or deletions shift the reading frame.

  • All amino acids after the mutation are different.

  • Usually problematic, especially if it occurs early in the protein sequence.

Impact on Fitness

• Mutations can have different effects on an organism's fitness.

  • Beneficial, Neutral, Deleterious (harmful).

• Beneficial: Enhance survival and reproduction.

• Neutral: Have no real effect on fitness (e.g., silent mutation).

• Deleterious: Decrease the ability to survive and reproduce.

• Most mutations are neutral or deleterious; beneficial mutations are rare.

Mutations in Non-Coding Regions

• Mutations in regulatory regions (e.g., promoters) can affect gene expression.

• Impacts phenotype via regulatory RNAs.

Chromosome-Level Mutations

• Often serious, arising from issues during meiosis or mitosis.

• Meiosis: Affects gamete formation and the resulting organism.

• Mitosis: Initially affects specific cells and their progeny, but can lead to tumor formation.

• Chromosome number alterations: Polyploidy, Aneuploidy.

Chromosome Structure Changes

• Inversion:

  • A chromosome piece breaks off, flips, and reattaches.

• Translocation:

  • A piece of one chromosome breaks off and attaches to another.

• Deletions:

  • A chromosome chunk breaks off.

• Duplication:

  • Multiple copies of a chromosome.

• Karyotypes from cancer patients often show multiple chromosomal abnormalities.

Impact of Chromosome Mutations

• Chromosome mutations can be beneficial, neutral, or deleterious.

• Most are deleterious because they affect many genes.

• Karyotypes visualize chromosome mutations.

• Aggressive cancers often show more damage in karyotypes due to increased genomic instability.

Central Dogma Review

• Relating genes to RNA and proteins.

• Focusing on transcription (DNA to RNA) and translation (RNA to protein).

Functions of Proteins

• Provide cell shape.

• Control chemical reactions as enzymes.

• Regulate transport of materials across membranes and within the cell.

• Proteins are the workhorses of the cell.

Overview of Transcription and Translation

• Cells build proteins using DNA instructions.

• DNA is transcribed into RNA.

• RNA carries information to ribosomes where proteins are synthesized.

• Translation converts nucleic acid code into amino acid code.

• Transcription occurring along a gene in a frog cell looks like a leaf

• Multiple RNAs can be transcribed from the same DNA simultaneously.

RNA Polymerases

• Enzymes responsible for making RNA.

• Use RNA version of DNA instructions.

• DNA template (red) and RNA template (yellow) with complementary base pairing.

• RNA synthesized in the 5' to 3' direction.

• Uses the 3' OH to add new nucleotides.

Template vs. Non-Template Strand

• One strand of DNA serves as the template, and the other is the non-template (coding) strand.

• RNA has the same sequence as the coding strand (except uracil replaces thymine).

• The noncoding strand is used as the template to generate the desired RNA sequence.

RNA Synthesis

• RNA synthesis is similar to DNA synthesis.

• RNA polymerases perform template-directed synthesis in the 5' to 3' direction.

• RNA polymerases do not need a primer to start.

Transcription Initiation in Bacteria

• Transcription can be broken down into three major parts: Initiation, Elongation, and Termination.

• RNA polymerases need help to initiate transcription.

• Sigma protein (σ) binds to DNA first.

• RNA polymerase works with sigma to form a holoenzyme.

• RNA polymerase is the core enzyme.

• Sigma binds to the promoter region.

• Different sigma proteins interact with different promoters, recognizing certain sequences.

• This process allows organisms to activate genes in response to environmental signals (e.g., temperature, nutrients).

Promoters

• Promoters are 40-50 base pairs long.

• Contain a "minus 10 box" (TATAAT) about 10 nucleotides upstream.

  • Rich in A's and T's to facilitate strand separation.

• Minus 35 box (TTGACA) is also present 35 nucleotides upstream.

• Both boxes are recognized by sigma, aiding in promoter binding and orientation.

Process of Transcription Initiation

• Transcription begins when sigma binds to the -35 and -10 boxes.

• Sigma can only bind in one orientation.

• The promoter orientation dictates which DNA strand is the template and the direction of RNA polymerase movement.

• RNA polymerase opens the DNA double helix just after the promoter region to create a transcription bubble.

• The template strand is threaded through the active site; RNA nucleotides are paired with the template strand.

Elongation Phase

• RNA polymerase moves along the DNA template.

• Synthesizes RNA in the 5' to 3' direction at about 50 nucleotides per second.

Termination Phase in Bacteria

• RNA polymerase transcribes a transcription termination signal.

• The RNA forms a hairpin loop due to complementary base pairing.

• This structure serves as a signal to stop transcription.

• RNA polymerase separates and releases the RNA transcript, which can then be used to make proteins.

Differences in Eukaryotes

• Bacteria have one kind of RNA polymerase; eukaryotes have three.

• Eukaryotes have more diverse promoters.

• TATA box is one important promoter region.

• Eukaryotes use basal transcription factors instead of sigma protein for fine-tuned gene regulation.

• Transcription factors recognize promoters, bind, and dictate where RNA polymerase will initiate transcription.

• Different termination signal: Instead of a hairpin loop, eukaryotes transcribe a long string of adenines (poly-A signal).

• When the cell transcribes the poly-A signal, transcription is terminated, and the transcript is cut off.

Location Differences

• Transcription occurs in the nucleus, and translation occurs in the cytoplasm in eukaryotes.

• In prokaryotes (bacteria), both transcription and translation occur in the cytoplasm.

Impact of Mutations

• Mutating three nucleotides located 10 nucleotides upstream from the transcription start site in bacteria would impede sigma protein binding.

• The -10 box is a key site that sigma needs to recognize to initiate transcription properly.

• If sigma cannot correctly recognize that site, transcription cannot properly initiate.