Transcription, RNA Processing, and Translation: Study Notes

Transcription, RNA Processing, and Translation

Overview of Transcription

Transcription is the process by which RNA polymerases synthesize an RNA copy of the instructions stored in DNA. This process is essential for gene expression and occurs in both prokaryotes and eukaryotes, with some key differences.

• RNA polymerases use ribonucleoside triphosphates (NTPs) to build RNA.

• Only one strand of DNA serves as the template strand; the other is the non-template (coding) strand, which matches the sequence of the mRNA (except U replaces T).

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

• Unlike DNA polymerases, RNA polymerases do not require a primer to begin transcription.

• Bacteria have one RNA polymerase; eukaryotes have at least three distinct types.

Initiation of Transcription in Bacteria

Transcription initiation is the first phase, requiring specific proteins and DNA sequences.

• RNA polymerase cannot initiate transcription alone; it requires a sigma protein to bind first.

• The combination of RNA polymerase and sigma forms a holoenzyme.

• Sigma recognizes promoters, which are DNA sequences where transcription begins.

• Bacterial promoters are typically 40–50 base pairs long and include the -10 box (TATAAT sequence) and -35 box (TTGACA sequence).

• Promoter orientation determines which DNA strand is used as the template and the direction of RNA polymerase movement.

Events Inside the Holoenzyme

Once the holoenzyme is formed, transcription begins with the following steps:

• RNA polymerase opens the DNA double helix, creating a transcription bubble.

• The template strand is threaded through the RNA polymerase active site.

• Incoming NTPs enter the enzyme and pair with complementary DNA bases, initiating polymerization.

Elongation and Termination in Bacteria

During elongation, RNA polymerase reads the DNA template and adds nucleotides to the 3' end of the RNA. Termination occurs when a specific signal is transcribed.

• Termination signal codes for RNA that forms a hairpin structure, causing RNA polymerase to separate from the RNA transcript.

Transcription in Eukaryotes

Eukaryotic transcription differs from bacterial transcription in several ways:

• Three distinct RNA polymerases are present.

• Promoters are larger and more diverse, including the TATA box.

• General transcription factors recognize promoters instead of sigma proteins.

• Termination involves a poly(A) signal and cleavage of the RNA downstream, rather than a hairpin.

• Transcription occurs in the nucleus; translation occurs in the cytoplasm.

RNA Processing in Eukaryotes

Primary Transcript and RNA Processing

In bacteria, transcription produces fully functional RNAs. In eukaryotes, the initial product is an immature primary transcript (pre-mRNA) that must undergo processing before translation.

• Primary transcripts require RNA processing to become mature mRNA.

Discovery of Split Eukaryotic Genes

Protein-coding genes in eukaryotes contain noncoding DNA segments called introns, which are not present in the final mRNA. Exons are the coding regions retained in mature mRNA.

• Introns are removed during RNA processing.

• Exons are joined together to form the final mRNA.

RNA Splicing

RNA splicing removes introns from primary transcripts and joins exons. This process is catalyzed by small nuclear ribonucleoproteins (snRNPs), which form a spliceosome.

• Splicing allows different mRNAs and proteins to be produced from a single gene (alternative splicing).

• Four steps to splicing:

  1. snRNPs bind to exon–intron and intron–exon boundaries and to an A near the end of the intron.

  2. Other snRNPs join to form a spliceosome.

  3. The intron forms a lariat structure (stem plus loop).

  4. The lariat is cut out, exons are linked, and the intron is degraded.

Adding Caps and Tails to Transcripts

Pre-mRNAs are processed by two additional events to become mature mRNAs:

• 5' cap: A modified guanine nucleotide added to the 5' end, enabling ribosome binding and protecting from degradation.

• Poly(A) tail: 100–250 adenine nucleotides added to the 3' end after cleavage, needed for translation and protection from degradation.

• After splicing and addition of the cap and tail, the product is a mature mRNA, which contains untranslated regions (UTRs) at both ends.

Key Terms and Concepts

• Transcription bubble: The region where the DNA double helix is unwound for RNA synthesis.

• Promoter: DNA sequence where RNA polymerase binds to initiate transcription.

• Spliceosome: Complex of snRNPs that catalyzes RNA splicing.

• Introns: Noncoding regions removed from pre-mRNA.

• Exons: Coding regions retained in mature mRNA.

• 5' cap and poly(A) tail: Modifications that protect mRNA and facilitate translation.

Relevant Equations

Direction of RNA synthesis:

$\text{RNA synthesis proceeds in the 5' \rightarrow 3' direction}$

Promoter sequences (bacterial):

$\text{-10 box: TATAAT}$

$\text{-35 box: TTGACA}$

Example: Alternative splicing allows a single gene to code for multiple proteins, increasing genetic diversity.