Gene Expression & Transcription in Eukaryotes

Gene Expression & Transcription in Eukaryotes

• Gene expression is the process of converting a gene's information into a functional molecule (protein or RNA).
• Gene expression is controlled at various points in the protein synthesis sequence.
• The human genome contains an estimated 30,000 to 120,000 genes.
• Only a small fraction of these genes are used in any particular cell at a given time.
• Genes are composed of several functional components involved in gene expression.
• Eukaryotic genes contain coding sequences (exons) interrupted by noncoding sequences (introns).
• The average gene consists of 7-10 exons spanning 10-16 kb of DNA.
• Genes must have exons, start signals, stop signals, and regulatory control elements.
• Broad functional units are the promoter region and the coding region.

Objectives for Gene Expression and Eukaryotic Transcription

• Gene Structure
• Gene expression
• Types of genes
• Post-transcriptional modifications: RNA splicing, 3' polyadenylation, 5' capping

Genes

• Genes are units of inheritance determining phenotypes.
• Each gene has instructions for a specific protein.
• DNA serves as the genetic blueprint for cellular proteins.

Gene Structure

• Each gene has functional components involved in various aspects of gene expression.
• Most eukaryotic genes include coding sequences (exons), separated by non-coding segments (introns).
• The average gene spans 10-16 kb and contains 7-10 exons.
• Each gene must contain exons, start signals, stop signals, and regulatory elements.
• Genes have a promoter region and a coding region.

Gene Expression

• Gene expression involves converting genetic information into cellular structures and functions.
• This occurs via producing a functional molecule (protein or RNA).
• Gene expression is regulated at different stages.

Central Dogma

• DNA → RNA → Protein
• Transcription: DNA is transcribed into RNA.
• Translation: RNA is translated into protein.

Types of Genes

• Constitutive (Housekeeping) genes: Expressed constantly at a predictable rate; not subject to regulation.
  • Examples: enzymes in glycolysis
• Regulated genes: Expressed only under specific conditions.
  • Examples: insulin gene in pancreas cells

Prokaryotes vs. Eukaryotes

• Prokaryotic transcription and translation happen in the same compartment.
• Eukaryotic transcription occurs in the nucleus; translation occurs in the cytoplasm.

Eukaryotic Promoter

• Promoter regions of DNA initiate transcription of a specific gene.
• Promoters are located near gene start sites upstream of the gene.
• Promoters are typically 100-1,000 base-pairs long.
  • Contains conserved promoter elements (e.g., CAAT box, TATA box, GC box, CAP site).

Eukaryotic Gene Promoter Sequences

• RNA polymerase II recognizes specific sequences (e.g., CAAT box, TATA box).
• The 45-55 bases within these sequences are upstream of the transcription start site.

Enhancer Sequences

• Regulatory DNA sequences that bind transcription factors.
• Enhancers can either enhance or repress the likelihood of transcription.
• Enhancers are not always near the gene they affect.

Silencers

• Transcription factors called repressors bind to regions called silencers.
• Silencers diminish transcription.

Transcription (Synthesis of RNA)

• Transcription copies DNA into RNA to carry information for protein synthesis.
• DNA's information is encoded for protein synthesis.

Messenger RNA (mRNA)

• mRNA carries genetic information from DNA to ribosomes for protein synthesis.
• mRNA is a type of RNA necessary for protein production.

Reverse Transcription

• Synthesis of DNA from an RNA template catalyzed by reverse transcriptase.
• Retroviruses use RNA as their genetic material for DNA synthesis within host cells.
• Examples include HIV and Hepatitis C.

Transcription in Eukaryotes

• Takes place in two steps:
  • 1. RNA polymerase enzymes perform the process based on Watson-Crick base pairing (A pairs with U).
  • 2. The RNA polymerase is responsible for producing an RNA copy of one DNA strand, which is then converted into the protein.

RNA Polymerase

• RNA Polymerases I, II, and III transcribe rRNA, mRNA, and tRNA, respectively.
• RNA Polymerase II is the primary enzyme involved in mRNA synthesis from a DNA template.
• RNA polymerase enzymes consist of multiple protein subunits.
• RNA polymerase II is more susceptible to ?-amanitin than RNA polymerases I and III.

Transcription Factors

• Polymerases interact with promoters via transcription factors.
• Transcription factors recognize and initiate transcription.
• Some transcription factors interact with specific recognition sequences within coding regions of DNA.
• Specific transcription factors (TFIIIA/C for RNA polymerase III) bind to specific recognition sequences within the coding region.

The General Transcription Factors Needed for Transcription Initiation by Eukaryotic RNA Polymerase II

• Specific factors are needed for successful transcription initiation.
• Key transcription factors include TFIID, TFIIB, TFIIF, TFIIE and TFIIH.

RNA Polymerase II

• Crucial for regulating mRNA synthesis.
• Consists of multiple core peptides.
• Core subunits, such as RPB1 and RPB2, have homologous sequences with bacterial RNA polymerase, indicating similar functionalities.
• RPB1 has a DNA-binding site; RPB2 binds to NTP (nucleotide triphosphate).

CTD

• Essential for transcription initiation by RNA polymerase II.
• It interacts with other proteins in the transcription machinery.
• It projects up to 50nm away from the enzyme's main structure.
• Only unphosphorylated CTD can trigger transcription initiation.

TATA Box

• A region within promoters, containing the consensus sequence TATAAA.
• Indicates the beginning of transcription.
• Used by RNA polymerase II.

General Transcription Factors TFIID

• Essential for many transcription initiation functions.
• TFIID is needed for transcription initiation.

Transcription Details (Continued)

• One DNA strand in a gene becomes a template strand.
• Transcription uses only one of the two DNA strands, the template strand.
• The RNA transcript formed during transcription is a copy of the template strand and not the coding strand.
• RNA polymerase catalyzes the process.
• Transcription ends when RNA polymerase hits a "stop" signal.
• DNA reforms into a double helix shape.

Post-transcriptional Modifications/Processing of mRNA

• The pre-messenger RNA undergoes three major functional modifications in the cell nucleus.
  • 1. 5' capping
  • 2. 3' polyadenylation
  • 3. RNA splicing.

Capping

• Addition of 7-methylguanosine (m7G) to the 5' end of the pre-mRNA.
• Stabilizes and protects the molecule.
• Protects the 5' end from ribonuclease.

Polyadenylation

• Adding adenine residues (about 250) to the 3' end of the pre-mRNA.
• Poly(A) tail protects the 3' end.

Splicing

• Introns, not needed for protein synthesis, are removed from pre-mRNA.
• The exons are joined together.
• Creation of the mature mRNA.

The Structure of a Eukaryotic Gene

• Shows how exons and introns are organized.
• Indicates promoter, enhancer, and terminator areas.

Synthesis of ribosomal RNA (rRNA) and Ribosome assembly

• rRNA is synthesized and then processes, and ribosomes assemble in the cellular nucleolus.

Synthesis of transfer RNA (tRNA)

• RNA polymerase III synthesizes tRNA.
• Special nucleotides are modified after transcription (example: pseudouridine).
• Modified and variable bases exist in tRNAs.

Explore the intricate mechanisms of gene expression and transcription in eukaryotic cells. This quiz covers essential concepts such as gene structure, regulatory components, and types of genes. Delve into the processes like RNA splicing and modifications that influence gene activity.