Transcription, RNA Processing, and Translation (Chapter 17, Part 1): Study Notes

Transcription: The Synthesis of RNA from DNA

Overview of Transcription

Transcription is the process by which RNA is synthesized from a DNA template. This process is essential for gene expression and occurs in both prokaryotic and eukaryotic cells.

• RNA polymerases synthesize an RNA version of the instructions stored in DNA.

• RNA polymerases use ribonucleoside triphosphates (NTPs) as substrates.

• 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).

Equation for RNA synthesis:

$\text{(DNA template)} + \text{NTPs} \xrightarrow{\text{RNA polymerase}} \text{RNA transcript}$

Comparison: DNA Polymerase vs. RNA Polymerase

• Both perform template-directed synthesis in the 5' → 3' direction.

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

• Bacteria have one RNA polymerase; eukaryotes have at least three types (RNA polymerase I, II, III).

Initiation of Transcription in Bacteria

Promoters and Sigma Factor

Transcription initiation requires specific DNA sequences called promoters and a protein called sigma factor in bacteria.

• Initiation is the first phase of transcription.

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

• Sigma recognizes promoters (specific DNA sequences where transcription begins).

Bacterial Promoter Structure

• Promoters are 40–50 base pairs long.

• Key regions:

  • -10 box: TATAAT sequence, ~10 bases upstream of the transcription start site.

  • -35 box: TTGACA sequence, ~35 bases upstream.

• Downstream: direction RNA polymerase moves (toward the gene).

• Upstream: opposite direction.

Events Inside the Holoenzyme

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

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

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

• Incoming NTPs pair with complementary DNA bases, and polymerization begins.

Elongation and Termination in Bacteria

Elongation

During elongation, RNA polymerase moves along the DNA template, adding nucleotides to the 3' end of the growing RNA molecule.

• Nucleotides are added to the 3' end of the RNA.

Termination

Termination occurs when RNA polymerase transcribes a specific sequence called the transcription-termination signal.

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

Transcription in Eukaryotes

Key Differences from Bacteria

• Three RNA polymerases (I, II, III).

• Promoters are larger and more diverse, often containing a TATA box.

• General transcription factors (not sigma) recognize promoters.

• At termination, a poly(A) signal is transcribed, and the RNA downstream is cut.

• 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 or pre-mRNA.

• Primary transcripts must undergo RNA processing before translation.

The Discovery of Split Eukaryotic Genes

• Protein-coding genes contain noncoding DNA (introns).

• Experiments showed that stretches of DNA are not present in mature mRNA.

• Introns are removed during processing; exons remain in the final mRNA.

RNA Splicing

Splicing removes introns from the primary transcript and joins exons together.

• 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).

Steps of Splicing:

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

2. Other snRNPs join to form the spliceosome.

3. The intron forms a lariat (loop) structure with A as the connecting point.

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:

  • 5' cap: Modified guanine nucleotide added to the 5' end, helps ribosomes bind and protects from degradation.

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

• After splicing and addition of the cap and tail, the product is a mature mRNA.

• Mature mRNAs contain untranslated regions (UTRs) at both ends.

Summary Table: Key Differences in Transcription and RNA Processing

Feature	Bacteria	Eukaryotes
RNA Polymerases	One	Three (I, II, III)
Promoter Elements	-10 and -35 boxes	TATA box and others
Initiation Factors	Sigma protein	General transcription factors
RNA Processing	None (RNA is functional after transcription)	5' cap, splicing, poly(A) tail
Location	Cytoplasm	Nucleus (transcription), cytoplasm (translation)

Example: In eukaryotes, the gene for β-globin contains three exons and two introns. After transcription, the introns are removed by splicing, and the mature mRNA is exported to the cytoplasm for translation.