11.3 RNA Transcription

RNA Transcription

RNA transcription is the process of creating an RNA copy from a DNA template. It's the first major step in gene expression, bridging the information stored in DNA to the functional molecules (proteins and RNAs) a cell needs. The process follows three stages: initiation, elongation, and termination, all driven by the enzyme RNA polymerase.

Transcription differs between prokaryotes and eukaryotes in both location and complexity. Prokaryotes transcribe in the cytoplasm with a single RNA polymerase, while eukaryotes use three different RNA polymerases in the nucleus and add extra processing steps before the RNA is ready for use.

Process of RNA Synthesis

RNA synthesis always proceeds in the 5' to 3' direction, catalyzed by RNA polymerase. The three stages break down as follows:

Initiation

1. RNA polymerase recognizes and binds to a promoter sequence located upstream of the gene.
2. The enzyme unwinds a short stretch of the DNA double helix, exposing the template strand for reading.

Elongation

1. RNA polymerase reads the template strand in the 3' to 5' direction while building the new RNA strand 5' to 3'.

2. Complementary RNA nucleotides are added one at a time according to base-pairing rules:

   • Adenine (A) on DNA pairs with Uracil (U) in RNA (not thymine)
   • Thymine (T) on DNA pairs with Adenine (A) in RNA
   • Guanine (G) pairs with Cytosine (C), and vice versa

3. A sugar-phosphate backbone forms as nucleotides are linked together. RNA uses ribose sugar rather than the deoxyribose found in DNA.

Termination

1. RNA polymerase encounters a termination signal on the DNA.
2. The newly synthesized RNA strand is released, and RNA polymerase dissociates from the template.

Transcription in Prokaryotes vs. Eukaryotes

Both prokaryotes and eukaryotes use DNA as a template, require RNA polymerase, and follow the same basic initiation → elongation → termination sequence. The key differences are where and how the process happens.

Prokaryotic transcription

• Takes place in the cytoplasm using a single type of RNA polymerase for all RNA synthesis.
• Because there's no nuclear membrane, transcription and translation can happen simultaneously. Ribosomes can begin translating an mRNA while it's still being transcribed.

Eukaryotic transcription

• Takes place in the nucleus using three distinct RNA polymerases, each dedicated to different RNA products:
  • RNA polymerase I → ribosomal RNA (rRNA)
  • RNA polymerase II → messenger RNA (mRNA)
  • RNA polymerase III → transfer RNA (tRNA) and other small RNAs
• Transcription and translation are spatially separated: RNA is made in the nucleus, then exported to the cytoplasm for translation.
• Before export, the RNA undergoes post-transcriptional modifications: a 5' cap is added, introns are spliced out, and a poly-A tail is attached to the 3' end.

Components of the Transcription Process

Promoters

Promoters are specific DNA sequences upstream of a gene that serve as the landing pad for RNA polymerase. They determine both the start point and the direction of transcription.

• In prokaryotes, promoters contain conserved sequences at the -10 and -35 positions (relative to the transcription start site). The sigma ($\sigma$) factor of RNA polymerase recognizes these regions.
• In eukaryotes, a common promoter element is the TATA box, typically located about 25–30 base pairs upstream of the start site. Transcription factors bind the TATA box first, then recruit RNA polymerase II.

RNA Polymerase

This is the central enzyme of transcription. Its roles include:

• Unwinding the DNA double helix to expose the template strand
• Reading the template 3' → 5' and synthesizing RNA 5' → 3'
• Maintaining the transcription bubble (the unwound region) during elongation
• Recognizing promoter sequences (directly in prokaryotes; with help from transcription factors in eukaryotes)

Termination Signals

These are DNA sequences that tell RNA polymerase to stop transcribing and release the RNA.

In prokaryotes, there are two distinct mechanisms:

1. Rho-dependent termination: The Rho ($\rho$) protein binds to the growing RNA and chases RNA polymerase. When the polymerase pauses at a termination site, Rho catches up and forces it to dissociate.
2. Rho-independent (intrinsic) termination: The RNA transcript forms a stem-loop (hairpin) structure from inverted repeat sequences, followed by a run of U residues (from A-T rich DNA). The hairpin causes the polymerase to stall, and the weak A-U base pairs allow the RNA to peel away from the template.

In eukaryotes, termination is less clearly defined. It generally involves recognition of specific sequences downstream of the gene and the action of termination/cleavage factors associated with the RNA polymerase complex.

DNA and RNA Structure

• Nucleotides are the building blocks of both DNA and RNA. Each consists of a sugar (deoxyribose in DNA, ribose in RNA), a phosphate group, and a nitrogenous base.
• Template strand: The DNA strand that RNA polymerase actually reads during transcription. It runs 3' to 5' relative to the direction of RNA synthesis.
• Coding strand (also called the non-template or sense strand): The opposite DNA strand. Its sequence is identical to the RNA transcript, except with T in place of U. This is why gene sequences are often written as the coding strand.
• Base pairing: Hydrogen bonds between complementary bases (A-T or A-U, and G-C) are what ensure the RNA copy is an accurate reflection of the DNA sequence.

Genetic Code and Protein Synthesis

These terms connect transcription to the next step, translation:

• Gene: A segment of DNA that encodes a specific protein or functional RNA molecule.
• Codon: A three-nucleotide sequence in mRNA that specifies a particular amino acid (or a stop signal). For example, AUG codes for methionine and also serves as the start codon.
• Anticodon: A complementary three-nucleotide sequence on tRNA that pairs with a codon during translation, ensuring the correct amino acid is added to the growing polypeptide chain.