Genes

Chapter 16 Genes Part 1

1. The Central Dogma

  • The central concept of molecular biology that describes the flow of genetic information within a biological system.

  • Process involves three key molecules: DNA, RNA, and proteins.

  • Key processes:

    • Replication: The process where DNA can copy itself.

    • Transcription: The conversion of DNA into RNA.

    • Translation: The process whereby RNA is used to synthesize proteins.

  • Important note: RNA can be reverse transcribed back into DNA, however, proteins cannot revert back to RNA.

2. RNA

  • Defined as the intermediate between DNA and protein.

  • Structurally differs from DNA in the following aspects:

    • It is single-stranded.

    • Contains uracil (U) instead of thymine (T).

    • Contains ribose sugar instead of deoxyribose.

Types of RNA

  • mRNA (messenger RNA):

    • Carries the genetic code from DNA to ribosomes for protein synthesis.

    • Synthesized during the process of transcription.

  • tRNA (transfer RNA):

    • Responsible for transporting amino acids to the ribosomes during translation based on the sequence in mRNA.

  • rRNA (ribosomal RNA):

    • Major structural and catalytic component of ribosomes, where translation occurs.

3. Transcription

  • This is the second step of the Central Dogma.

  • Defined as the synthesis of RNA using a DNA template via RNA-dependent RNA polymerases.

  • Key enzyme:

    • RNA Polymerase: The enzyme responsible for synthesizing mRNA from a DNA template.

Types of RNA Polymerases

  • RNAP I: Synthesizes rRNA.

  • RNAP II: Synthesizes mRNA and some small RNA molecules.

  • RNAP III: Synthesizes tRNA and some small RNA molecules.

Mechanism of Transcription

  • Only one DNA strand is transcribed, occurring in the 5’ to 3’ direction.

    • Thus, the template DNA strand is read in the 3’ to 5’ direction.

  • Terminologies:

    • Upstream: Relative to the 5’ end of the RNA transcript.

    • Downstream: Relative to the 3’ end of the RNA transcript.

  • The mRNA is synthesized based on the coding strand (non-template strand) of DNA.

  • Steps of transcription:

    1. Initiation:

    • Begins when RNA Polymerase binds to the promoter region of the gene.

    1. Elongation:

    • The actual synthesis of the mRNA strand.

    1. Termination:

    • The process is signified by specific sequences (terminators) that signal the end of transcription.

4. Gene Structure

  • Promoter:

    • A regulatory sequence situated upstream of the gene that serves as the binding site for RNA polymerase.

    • Often referred to as the ‘TATA box’ due to its characteristic sequence rich in thymine (T) and adenine (A), typically represented as TATAAA, located 25-35 base pairs upstream of the gene.

  • Exons:

    • Coding regions of a gene that are retained and transcribed into mRNA.

  • Introns:

    • Non-coding regions between exons, which are spliced out during mRNA processing.

  • Terminator:

    • A sequence that indicates the conclusion of transcription, instructing RNA polymerase to stop.

  • Regulatory elements/enhancers:

    • Additional sequences that control gene expression control when, where, and how much of the gene is expressed.

Initiation of Transcription

  • Requires a promoter for RNA polymerase to bind; otherwise, RNAP cannot initiate transcription.

  • Transcription Factors (TF’s):

    • Proteins that facilitate and regulate RNAP binding to the promoter.

    • Types include:

    • Repressors: Prevent transcription when bound to the promoter.

    • Activators: Allow transcription when bound to the promoter.

  • Importance of regulating transcription:

    • Cells must flexibly regulate protein synthesis based on contextual/environmental factors.

    • Examples of regulation in response to:

    • Cell division and growth signals.

    • Apoptotic signals.

Transcription Process Overview

  • RNA polymerase binds to the promoter, opens, and unwinds the DNA.

  • The resulting mRNA strand grows in the 5’ to 3’ direction, complementary to the DNA template strand.

  • Only one RNA strand is produced, thus maintaining a single-stranded structure.

  • Termination in Prokaryotes:

    • Terminator sequence is transcribed, leading to the release of both the RNA strand and RNA polymerase.

  • Termination in Eukaryotes:

    • More complex; requires specific sequences to delineate the stop point of transcription, typically 100 bases from the gene’s end.

5. The Genetic Code

  • A gene is defined as a sequence of nucleotides that encodes an mRNA molecule.

  • The information within a gene for protein synthesis is referred to as the genetic code.

  • The genetic code is based on codons:

    • Groups of three nucleotide bases that specify the addition of a particular amino acid or a stop signal in a polypeptide chain.

  • Codon characteristics:

    • Universally present across organisms.

    • Read in sequences of three bases on mRNA, always in a 5’ to 3’ direction.

  • Why three bases per codon?

    • Options:

    1. With four nucleotides (A, U, G, C) and three bases per codon, the total possible combinations are $64$ (i.e., $4^3$).

    2. This is sufficient to code for the 20 commonly occurring amino acids in proteins.

Reading Frame

  • The reading frame is established as codons are sequentially read, modulating protein synthesis.

  • Frame Shift Mutation:

    • Errors or shifts in the reading frame can lead to mutations, generating entirely different amino acid sequences.

6. Translation

  • The third and final step of the Central Dogma.

  • Translation is the process wherein the information carried by mRNA is utilized for protein synthesis.

Differences Between Prokaryotic and Eukaryotic Translation

  • In prokaryotes, transcription and translation occur simultaneously in the cytoplasm.

  • In eukaryotes, these processes are separated:

    • Transcription occurs in the nucleus.

    • Translation occurs in the cytoplasm.

Ribosomes

  • Composed of rRNA and proteins, ribosomes are the essential machinery of translation.

  • Subunits:

    • Contain a large and a small subunit that function together.

    • Ribosomes catalyze the translation process.

Peptide Bonds and Polypeptides

  • Ribosomes facilitate the formation of peptide bonds linking amino acids to form a polypeptide chain.

  • The unique sequence of codons in the mRNA transcript determines the polypeptide structure.

  • Monomer: Amino acids

  • Polymer: Polypeptide chains

  • tRNA (Transfer RNA):

    • Carries specific amino acids to the translation site based on mRNA codons.

    • Exhibits complex, folded 3D structures necessary for proper function.

    • Each tRNA possesses an anticodon sequence that pairs complementary with mRNA codons.

Degeneracy of the Genetic Code

  • Although there are 64 codon combinations, there are fewer tRNA types since some tRNAs can correspond to multiple codons.

  • The wobble hypothesis explains how variability in the third nucleotide of a codon accommodates this overlap.

Ribosome Sites

  • The ribosome consists of three sites:

    • A site (Aminoacyl site): Where tRNA with the corresponding amino acid first binds.

    • P site (Peptidyl site): Where the tRNA carries the polypeptide chain.

    • E site (Exit site): Where completed tRNA exits the ribosome.

Stages of Translation

  1. Initiation:

    • The beginning of polypeptide formation, requiring energy in the form of GTP.

    • The initiator tRNA recognizes the start codon (AUG), which codes for Methionine (Met), written as tRNA^Met.

  2. Elongation:

    • The addition of amino acids to the growing polypeptide chain, facilitated by the binding of the next corresponding tRNA to the A site, requiring GTP energy.

    • Peptide bond formation occurs spontaneously.

  3. Termination:

    • Signaled by a stop codon (UAA, UGA, UAG), resulting in the release of the polypeptide, mRNA, and the last tRNA from the ribosome through the action of a release factor.

Polyribosomes

  • A single mRNA transcript can be translated by multiple ribosomes simultaneously; typically, around 20 ribosomes can coordinate with each mRNA.

7. Post-transcriptional Modifications

  • In eukaryotes, the precursor mRNA formed in the nucleus undergoes modifications before translation.

  • 5’ Capping:

    • Enzymatic addition of a 5’ cap on the mRNA, essential for ribosome binding and stability against degradation while promoting export from the nucleus.

  • 3’ Polyadenylation Signal:

    • A polyadenylation signal added at the 3’ end results in a poly-A tail (series of adenines) that enhances mRNA stability.

  • The mRNA transcript often exceeds the size of the resultant mature mRNA due to intron presence, necessitating:

    • Splicing:

      • Involves the removal of introns and ligation of exons via spliceosome activity, finalizing the mature mRNA transcript.

8. Protein Sorting

  • Observations through electron micrographs of eukaryotic cells during translation reveal two ribosome populations:

    • Free Ribosomes:

    • Suspended in the cytosol and primarily synthesize proteins that remain in the cytosol.

    • Bound Ribosomes:

    • Attached to the cytosolic side of the Endoplasmic Reticulum (ER), producing proteins that integrate into the endomembrane system or are secreted from the cell.

    • Mutations

      • May occur as accidents during DNA replication or may be induced by DNA-damaging chemicals/toxins 

      Some terms regarding Mutations

      • Mutagen - DNA-damage inducers

      • Carcinogens - mutagens that are correlated with cancer 

      • Mutational hot spots - some DNA regions that are more prone to mutations (trinucleotide repeats are an example - Huntington's and Fragile X) 

      Mutations that are most likely to lead to genetic change include…

      -Point Mutation (single base change) 

      - Missense mutation - single base changes; changes amino acid 

      - Nonsense mutation - single base change; encodes a stop codon

      - Silent mutation (no change) - single base change; no change in AA

      - Frameshift mutations - shifting of reading frame 

      - Transposons (transposable elements) - “jumping genes” that can jump around in the genome and insert themselves somewhere else

      - Mutations that result in the substitution of one base for another 

      Recall: Sickle cell anemia 

      • Due to a mutation in the protein hemoglobin 

      • A missense mutation occurs at the 6th codon in the hemoglobin beta chain

      • In the DNA, a T is replaced with an A, which leads to valine being translated instead of glutamic acid

      • The resulting hemoglobin is sticky, which prompts it to crystallize easily, forming sickled red blood cells.