Comprehensive Transcription and Translation Notes

Transcription Factors and RNA Polymerase

  • RNA polymerase, the primary enzyme, attaches to the promoter.
  • Transcription factors facilitate the binding of RNA polymerase to the promoter.

DNA Strands

  • RNA polymerase opens and separates the DNA strands.
  • Template Strand: The strand used as a guide for RNA synthesis.
  • Non-Template Strand: The strand that is largely ignored during transcription.

mRNA Modification

  • The initial RNA transcript undergoes modification.

5' Cap

  • A modified guanine molecule is added to the 5' end.
  • Protects the transcript from degradation by cytoplasmic enzymes (nucleases).
  • Enzymes in the cytoplasm do not recognize the modified end as nucleic acid, preventing its destruction.

3' Poly-A Tail

  • A long sequence of adenine (A) bases is added to the 3' end.

  • Slows down degradation by nuclease enzymes but does not prevent it entirely.

  • A longer tail allows the transcript to persist longer in the cytoplasm, leading to more protein production.

  • A shorter tail results in a briefer persistence and less protein production.

  • The cell can regulate protein production by adjusting the length of the poly-A tail.

  • Making proteins is energy-intensive, so regulation is crucial.

Exon Splicing (Intron Removal)

  • Non-coding sections (introns) are removed from the RNA transcript.

  • The remaining coding sections (exons) are spliced back together.

  • The book doesn't cover the mechanism, but several enzymes are involved.

  • Introns: Intervening regions that are removed.

  • Exons: Expressed regions that are retained and spliced together.

  • Typically, about 90% of the transcript is removed during splicing.

  • There is variable splicing, where exons can be reordered, producing different proteins from the same transcript; for example, exon ABCD can be put back in the order of BCAD.

  • The human genome project revealed that a single gene can produce multiple proteins through variable splicing.

  • There are about 30,000 genes in the human genome, but they can make about 125,000 different proteins.

  • Introns are only called introns if they are removed; if left in, they are considered part of the exon.

  • This process occurs post-transcriptionally but pre-translationally, while still in the nucleus.

  • Once modified, the mRNA transcript leaves the nucleus for translation in the cytoplasm.

mRNA Export and Ribosome Binding

  • The modified 5' cap helps the mRNA bind with transporters in the nuclear envelope and facilitates export.
  • The cap also helps the mRNA find a ribosome in the cytoplasm.

Translation Players

  • mRNA: The modified transcript.
  • Ribosomes
  • tRNA
  • rRNA

Untranslated Regions (UTRs)

  • The region between the 5' cap and the start codon is called the leader sequence or untranslated region (UTR).
  • This region is transcribed but not translated.
  • The leader sequence aids in exiting the nucleus and finding the ribosome.

Codons

  • The ribosome translates the mRNA three bases (a codon) at a time.
  • Start Codon: AUG is typically the start codon.
  • Genes may have start codons other than AUG but not frequently enough to change the definition of AUG representing the start codon.
  • Everything before the start codon is part of the leader sequence.

tRNA

  • tRNA is a piece of RNA that is folded into hairpin loops held together by complementary base pairing.

  • One end of the tRNA binds an amino acid (peptide binding site), and the other end has an anticodon.

  • The anticodon is a three-base sequence complementary to a codon on the mRNA.

  • The tRNA with the anticodon UAC always carries the amino acid methionine (Met).

Codon Table and Amino Acids

  • There are 20 common amino acids that make up all proteins.
  • The codon table lists all codons and their corresponding amino acids.
  • Some amino acids are coded for by multiple codons (redundancy).
  • AUG codes for methionine and is the start codon.
  • UAA, UAG, and UGA are stop codons (do not need to be memorized).
  • DNA sequence corresponds to RNA sequence which corresponds to amino acid sequence which then codes for the protein.
  • There are 64 possible codons but only 20 amino acids, resulting in redundancy.
  • The third base in a codon is often variable (wobble), allowing for redundancy.

Protein Synthesis

  • The ribosome has two subunits: one large, one small.
  • The ribosome clamps into the start codon along with the first tRNA.
  • The ribosome consists of proteins and ribosomal RNA (rRNA).
  • First assumption of chromosomes was DNA only had four bases in it and the proteins were doing the actual inheritance information storage work, turns out this was the opposite.
  • First assumption breaking down the ribosome they found it contained a lot of protein, so they assumed that the protein was doing the work and the nucleic acid was forming the scaffolding, this was also the opposite.
  • Chromatin is about 50% protein (histones, which DNA is wrapped around) and 50% DNA. Proteins make up the scaffolding in this instance.
  • Combination of different macro molecules (carbohydrates, lipids, nucleic acids, and proteins) will form combination molecules (lipoproteins, glycolipids, etc.).

Ribosome Binding Sites

  • The ribosome has two binding sites to accommodate two tRNAs at a time.
  • The ribosome moves down the mRNA three bases at a time.
  • As the ribosome moves, tRNAs are brought in, and the previous one is released.
  • The ribosome breaks the bond between the first tRNA and its amino acid and binds the amino acid to the second tRNA.
  • Recycling enzymes grab and attach the amino acid back on to the right tRNA for it to be recycled back into the system for use.

Termination

  • When the ribosome reaches a stop codon, a release factor binds instead of a tRNA.
  • The release factor causes the ribosome to disengage, and the protein is released.
  • The process occurs in the cytoplasm for small simple proteins with a free-floating ribosome.
  • More complex proteins use ribosomes bound to the rough ER.
  • Chaperone proteins in the rough ER aid protein folding and modification.
  • Termination happens when the tRNA with methionine and anticodon binds to the start codon with the ribosome at the binding site, ribosome starts moving down one codon at a time, new tRNAs come in and old ones are kicked out, tRNA with old codon now has a amino acid binded to the new tRNA codon.

Translation Overview

  • Initiation: tRNA with methionine and anticodon binds to the start codon along with the ribosome.
  • Elongation: The ribosome moves down one codon at a time which allows for new tRNA to come in. The old tRNA is kicked out. The amino acids are broken off the old tRNA and bound to the new tRNA codon.
  • Termination: A release factor protein binds to the stop codon, and everything disengages and the protein folds up.

RNA Types

  • tRNA (transfer RNA): Transfers amino acids to the mRNA and ribosome.
  • mRNA (messenger RNA): Carries the instructions to make the protein.
  • rRNA (ribosomal RNA): A component of the ribosome.

Key Terms

  • Start codon
  • Reading frame
  • Wobble

Operons

  • Bacterial cells need multiple proteins for one activity, they will put them right next to each other with no dead space in between them.
  • Bacterial cells use one long mRNA to be transcripted at the same time.
  • Arrangement of single promoter followed by the sequences you need to make the final protein only appears in bacteria called an operon.
  • Eukaryotes have a lot of space so they can bring things together from other times. Bacteria does not have enough DNA to work with so they are as efficient as possible.
  • Some operons are always on while some are always off and need to be turned on/off.

Chromatin Packaging

  • How tightly the chromatin (DNA and histone proteins) is packaged can affect gene expression.
  • Tightly packed regions (heterochromatin) are not transcribed.
  • Loosely packed regions (euchromatin) are accessible for transcription.
  • Chromatin packaging is a way of turning genes on and off/making something available.

Methylation

  • Addition of methyl groups (CH3) to DNA prevents transcription factors from binding.
  • Methylation can control turning genes on and off for long periods without altering the DNA sequence.
  • Chromatin packaging and methylation are epigenetic mechanisms.
  • Epigenetics is the study of changes outside of the DNA sequence that can affect gene expression and persist across generations.

Post-Transcriptional Regulation

  • Alternative splicing
  • RNA export (holding RNA back in the nucleus)

MicroRNA

  • MicroRNAs can bind to mRNA and block ribosomes, preventing translation.

Protein Modification

  • The endoplasmic reticulum (ER) and Golgi apparatus modify proteins for better function.

Mutations

  • A mutation is a change in the DNA that affects the protein.
  • Change to the DNA that effects the protein (amino acid sequence is altered).
  • Mutations are a natural part of life and can lead to variety.
  • Mutations can be spontaneous or caused by chemicals, UV radiation, or other factors.
  • Type: most common
  • Mutant: less common version

Point Mutations

  • A point mutation is a change in a single nucleotide.

Types of Point Mutations

  • Base Substitution: One base is changed for another (e.g., A to G).
  • Insertion: A base is added.
  • Deletion: A base is removed.

Effects of Point Mutations

  • Substitution tends to change one amino acid.
  • Insertion/deletion changes the reading frame; changing every amino acid downstream.

Silent Mutation

  • No change to the amino acid sequence due to wobble.

Missense Mutation

  • One amino acid is changed which can cause a different function (sickle cell anemia caused by one single amino acid being different).

Nonsense Mutation

  • A premature stop codon is introduced.

Frameshift Mutation

  • This occurs with insertions and deletions which will cause for there to be change in downstream acids.

Significance of Mutations

  • They contribute to variation.
  • Duplication of the FOXP2 (language) gene in humans and Neanderthals is associated with the ability to speak.