Chapter 17: Transcription and Translation – From DNA to Protein

Chapter 17: Transcription and Translation

Overview

This chapter explores how genetic information encoded in DNA is used to synthesize RNA and proteins. The processes of transcription and translation are central to gene expression in all living cells.

17.1 An Overview of Transcription

RNA Synthesis from DNA

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

• RNA polymerase uses ribonucleoside triphosphates (NTPs) as substrates (in contrast to dNTPs in DNA synthesis).

• RNA is synthesized by matching complementary bases to one strand of DNA, called the template strand.

• The other DNA strand is the non-template (coding) strand, which matches the sequence of the mRNA (except U replaces T).

Transcription Is the Synthesis of RNA from a DNA Template

• RNA polymerase catalyzes the formation of phosphodiester bonds between ribonucleotides, using the DNA template to specify the sequence.

• The reaction can be summarized as: $\text{(NMP)}_n + \text{NTP} \rightarrow \text{(NMP)}_{n+1} + PP_i$

Initiation: How Does Transcription Begin in Bacteria?

• Initiation is the first phase of transcription.

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

• Together, RNA polymerase and sigma form a holoenzyme.

• Sigma recognizes specific DNA sequences called promoters, where transcription begins.

• RNA polymerase is the core enzyme that synthesizes RNA.

Events Inside the Holoenzyme

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

• Sigma binds in only one orientation, determining which DNA strand is used as the template and the direction of RNA synthesis.

Termination in Bacteria

• Transcription ends when an RNA hairpin forms, causing the RNA polymerase to dissociate from the DNA template.

Transcription in Eukaryotes

• Eukaryotes have three distinct RNA polymerases (I, II, III).

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

• General transcription factors (not sigma) recognize promoters.

• Termination involves a poly(A) signal rather than a hairpin; the RNA downstream is cut.

• Transcription occurs in the nucleus, while translation occurs in the cytoplasm.

17.2 RNA Processing in Eukaryotes

• In bacteria, transcription produces functional RNAs directly.

• In eukaryotes, the initial RNA product is an immature primary transcript or pre-mRNA.

• Primary transcripts must undergo RNA processing before translation.

Discovery of Introns

• Introns are non-coding sequences removed from the final mRNA.

• Exons are coding sequences that remain in the mature mRNA.

RNA Splicing

• Primary RNA transcripts contain both exons and introns.

• Introns are removed by splicing, catalyzed by small nuclear ribonucleoproteins (snRNPs).

• snRNPs form a complex called a spliceosome.

• Splicing allows for the production of different mRNAs and proteins from a single gene (alternative splicing).

Adding Caps and Tails to Transcripts

• Pre-mRNAs are processed by two additional events:

  • 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, necessary for translation and stability.

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

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

17.3 An Introduction to Translation

• The sequence of mRNA bases is converted into an amino acid sequence (protein).

• Translation involves ribosomes, mRNA, and tRNAs.

An Overview of Translation

• In bacteria, translation can begin before transcription is complete (coupled transcription and translation).

• Multiple ribosomes can translate a single mRNA simultaneously, forming a polyribosome.

• In eukaryotes, transcription and translation are separated by the nuclear envelope.

How Does mRNA Specify Amino Acids?

• Two hypotheses:

  1. mRNA codons and amino acids interact directly.

  2. Crick's adapter hypothesis: an adapter molecule (tRNA) holds the amino acid and interacts with the codon.

17.4 The Structure and Function of Transfer RNA (tRNA)

• Transfer RNA (tRNA) acts as the adapter molecule in translation.

• An aminoacyl tRNA is a tRNA linked to its specific amino acid.

• Amino acids are transferred from tRNAs to the growing polypeptide chain.

How Are Amino Acids Attached to tRNAs?

• Aminoacyl-tRNA synthetases "charge" tRNAs by catalyzing the addition of amino acids.

• There are 20 different synthetases, one for each amino acid.

• For each amino acid, there is one or more corresponding tRNAs.

How Many tRNAs Are There?

• There are 61 codons for amino acids but only about 40 tRNAs in most cells.

• Wobble pairing allows the third position of the anticodon to form nonstandard base pairs, enabling one tRNA to recognize multiple codons.

17.5 Ribosome Structure and Function in Translation

• Ribosomes are composed of many proteins and ribosomal RNA (rRNA).

• They consist of two subunits:

  1. The small subunit holds the mRNA in place.

  2. The large subunit is where peptide bonds form.

• During translation, three tRNAs line up within the ribosome at the A, P, and E sites.

Ribosome Structure and Function in Translation

• Protein synthesis occurs in a three-step sequence:

  1. An aminoacyl tRNA enters the A site and remains if there is a codon-anticodon match.

  2. A peptide bond forms between the amino acid on the A-site tRNA and the polypeptide on the P-site tRNA.

  3. The ribosome moves down the mRNA by one codon; the tRNA in the E site exits, and the A site is available for another tRNA.

• The protein grows by one amino acid with each cycle.

• Amino acids are always added to the carboxyl end (C-terminus) of the polypeptide.

• Translation has three phases: Initiation, Elongation, and Termination.

Initiation of Translation

• Requires binding of the initiator tRNA to the start codon on mRNA and assembly of the ribosome.

Elongation of the Polypeptide Chain

• Three steps repeat at each codon:

  1. Arrival of the aminoacyl tRNA at the A site.

  2. Peptide bond formation between amino acids.

  3. Translocation of the ribosome along the mRNA.

Termination of Translation

• Occurs when a release factor binds to a stop codon encountered by the ribosome, releasing the completed polypeptide.

Major Steps of Gene Expression in a Eukaryotic Cell

1. Transcription (in the nucleus): DNA is transcribed to pre-mRNA.

2. RNA processing (in the nucleus): pre-mRNA is processed to mature mRNA (splicing, capping, polyadenylation).

3. Translation (in the cytoplasm): mRNA is translated by ribosomes to synthesize proteins.

4. Post-translational modification: Proteins may be modified after translation (e.g., phosphorylation, folding).

Summary Table: Key Differences in Transcription and Translation between Bacteria and Eukaryotes

Feature	Bacteria	Eukaryotes
RNA Polymerases	One	Three (I, II, III)
Promoter Recognition	Sigma protein	General transcription factors
Termination Signal	Hairpin loop	Poly(A) signal
RNA Processing	None (mRNA is ready for translation)	Splicing, 5' cap, poly(A) tail
Location of Transcription & Translation	Cytoplasm (can be coupled)	Transcription in nucleus, translation in cytoplasm

Key Terms

• Transcription: Synthesis of RNA from a DNA template.

• Translation: Synthesis of a polypeptide using the information in mRNA.

• RNA polymerase: Enzyme that synthesizes RNA.

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

• tRNA: Adapter molecule that brings amino acids to the ribosome during translation.

• Ribosome: Molecular machine that synthesizes proteins by translating mRNA.

• Spliceosome: Complex that removes introns from pre-mRNA.

• Polyribosome: Multiple ribosomes translating a single mRNA simultaneously.