Molecular Basis of Inheritance: RNA Types, Gene Function, and the Genetic Code

The study of the molecular basis of inheritance involves a comprehensive look at the structure and discovery of nucleic acids, specifically DNA and RNA.
Key topics in this lecture include:
- Structure of Nucleic Acid (DNA Discovery, Watson and Crick Model)
- Packaging of DNA
- Search for Genetic Material
- DNA Replication (Direction, Substrates, and Enzymes)
- Transcription and Translation (Prokaryotes vs. Eukaryotes)
- Types of RNA (mRNA, rRNA, tRNA, snRNA, scRNA)
- Genetic Code and Gene Expression
- Specialized topics: Human Genome Project and DNA Fingerprinting

Messenger RNA (mRNA):
- Carries the genetic code for protein synthesis.
- In Eukaryotes: Initially formed as Heterogeneous nuclear RNA (hnRNA) which undergoes processing/modification to become mRNA.
- In Prokaryotes: Formed directly as mRNA without intermediate processing.
Ribosomal RNA (rRNA):
- Synthesized in the nucleolus in Eukaryotes.
- Composition: rRNA combines with proteins to form Ribosome subunits.
- These subunits move out from the nucleus into the cytoplasm for protein synthesis.
Transfer RNA (tRNA):
- Known as the "Adapter molecule" or Soluble RNA (sRNA).
- Functions: Reads the code on the mRNA and brings the corresponding amino acid to the site of protein synthesis.
- Structure: Basic structure is Clover Leaf (secondary structure), but the functional folding is a 3D inverted L-shape (tertiary structure).
Small Nuclear RNA (snRNA):
- Located inside the nucleus.
- Primary function: Involved in the processing of RNA (e.g., splicing).
Small Cytoplasmic RNA (scRNA):
- Located in the cytoplasm.
- Component of the Signal Recognition Particle (SRP) which helps bring ribosomes to the Endoplasmic Reticulum (ER).

Abundance (Amount): rRNA ( $80-85\%$ ) > tRNA ( $10-15\%$ ) > mRNA ( $3-5\%$ ).
Stability: rRNA > tRNA > mRNA > hnRNA. Ribosomal RNA is the most stable, whereas mRNA (especially hnRNA) is the least stable.
Size: rRNA has specific sizes (e.g., $28S, 18S, 5.8S, 5S$ ). The smallest RNA is tRNA ( $75-90$ nucleotides long).
Variability (Types): mRNA is the most diverse type because it depends on the variety of proteins/amino acids required for synthesis.

Prokaryotes (Bacteria):
- Context: No separation between the cytosol and the nucleus.
- Transcription and Translation occur in the cytoplasm.
- Coupling: Translation can begin much before the mRNA is fully transcribed. This unique synchronization is called "Coupled Transcription-Translation."
- Processing: mRNA does not require processing to become active.
- RNA Polymerase: Only one type of RNA polymerase transcribes all types of RNA.
Eukaryotes:
- Separation: Transcription occurs in the nucleus; Translation occurs in the cytoplasm.
- Division of Labor: At least three types of RNA Polymerase are used:
  - RNA Polymerase I: Transcribes rRNAs ( $28S, 18S, 5.8S$ ).
  - RNA Polymerase II: Transcribes mRNA (initially hnRNA).
  - RNA Polymerase III: Transcribes tRNA and $5S$ rRNA.
- Complexity: Involves splicing (removal of introns), capping, and tailing.
- Cistron Type: Eukaryotic genes are Monocistronic.

Definition: The gene is the functional unit of inheritance, historically difficult to define only by sequence.
Gene vs. Cistron:
- A Gene is a sequence on DNA coding for any type of RNA (tRNA, rRNA, mRNA).
- A Cistron is a gene sequence that specifically codes for a polypeptide.
- Logic: "All cistrons are genes, but all genes are not cistrons" (since some genes code for non-translated RNAs like tRNA).
Polycistronic (Prokaryotes): One transcription unit/mRNA contains many genes/cistrons under a common promoter and terminator.
Monocistronic (Eukaryotes): One transcription unit codes for a single cistron.
Split Genes: Eukaryotic genes are interrupted. Coding sequences are called Exons, and non-coding sequences are called Introns.
- Presence of Introns is considered "Reminiscent of Antiquity" (remnants of ancient evolutionary features).
- Archaebacteria also possess introns, a feature shared with Eukaryotes.

Core Principle: While Replication and Transcription utilize base-pairing complementarity, Translation requires converting a nucleic acid sequence into a polypeptide sequence.
Triplet Nature:
- George Gamow (Physicist) proposed the triplet code. Logic: With only 4 bases ( $A, U, G, C$ ), a single-base code would yield only $4^1 = 4$ amino acids; a double-base code yields $4^2 = 16$ ; a triple-base code yields $4^3 = 64$ . This is sufficient to cover the $20$ essential amino acids.
Key Codons:
- Initiator Codon: AUG (codes for Methionine).
- Stop Codons: UAA (Ochre), UAG (Amber), and UGA (Opal). These terminate synthesis and do not have corresponding tRNAs.
- Tryptophan is coded by a single codon: UGG.
Deciphering Scientists:
- Har Gobind Khorana: Synthesized RNA molecules with defined combinations of bases (homopolymers and copolymers).
- Marshall Nirenberg: Used a cell-free system for protein synthesis to decipher the code (main contribution to the codon table).
- Severo Ochoa: Used Polynucleotide phosphorylase to synthesize RNA template-independently.

Q1: Where does DNA Replication occur in E. coli?
- Answer: Cytoplasm (specifically the nucleoid area) after/during fission.
Q2: What is the substrate for DNA Polymerase?
- Answer: dNTPs ( $dATP, dGTP, dTTP, dCTP$ ).
Q3: What is the direction of DNA Replication?
- Answer: Always $5' \rightarrow 3'$ .
Q4: Is the origin (Ori) located on plasmids or nucleoids?
- Answer: Both. Both Eukaryotes and Prokaryotes have defined origins.
Q5: Which feature is similar in Eukaryotes and Prokaryotes regarding replication?
- Answer: The direction of DNA replication ( $5' \rightarrow 3'$ ).
Q6: Is it true that DNA Polymerase helps open DNA at the origin?
- Answer: No; Helicase/other factors are typically involved; DNA polymerase primary role is synthesis.
Q7: Which RNA is synthesized in the nucleolus?
- Answer: rRNA (specifically $28S, 18S, 5.8S$ ).
Q8: What is charged tRNA?
- Answer: A tRNA linked to its specific amino acid at the $3'$ end.
Q9: Which RNA polymerase transcribes snRNA?
- Answer: RNA Pol III.
Q10: Where is snRNA located?
- Answer: In the nucleus (e.g., yeast nucleus).
Q11: Which molecule is present only in the nucleus and not the cytoplasm?
- Answer: hnRNA (it must be processed into mRNA before leaving the nucleus).
Q12: Recognition of the Promoter is done by?
- Answer: Sigma Factor ( $\sigma$ ) in Prokaryotes along with RNA polymerase.