Unit 6: Gene Expression and Regulation Notes

Unit 6: Gene Expression and Regulation includes the following lessons:

Lesson 1: DNA & RNA

Aim: Explore the molecular characteristics and functions of DNA and RNA that are fundamental to heredity.

Think-Pair-Share: The Genetic Code is universal among all organisms, suggesting a common evolutionary origin. This code is composed of sequences of nucleotides that determine the traits passed from parents to offspring.

RNA Structure and Function

Learning Objectives:

• Describe structures involved in hereditary information transmission and their roles in cell function.

• Identify key characteristics of DNA and RNA that define them as hereditary materials.

RNA nucleotides are monomers of RNA comprised of:

• 5 Carbon Ribose Sugar (numbered 1-5 for identification)

• Phosphate attached to the 5’ ribose carbon, forming the backbone of the RNA strand.

• Nitrogenous bases: Adenine (A), Cytosine (C), Guanine (G), and Uracil (U); the presence of Uracil is a significant difference from DNA.

The 3' OH group is critical for nucleotide synthesis; the presence of a 2' OH in RNA makes it less stable, which is crucial for its transient functions in the cell, contrasting with DNA's stability for long-term storage of genetic information.

Base Pairing and Bonding

• Purines: Adenine (A), Guanine (G) - characterized by their double-ring structure, allowing for complex pairing.

• Pyrimidines: Cytosine (C), Uracil (U), Thymine (T) - feature a single-ring structure.

Base pairing rules in transcription:

• Adenine (A) pairs with Uracil (U) in RNA transcription, while Cytosine (C) pairs with Guanine (G).

Hydrogen Bonding: Synonymous pairs exhibit either 2 hydrogen bonds (A-T) or 3 hydrogen bonds (C-G), which contribute to the stability of the DNA structure.

DNA Structure and History

1. DNA Structure: DNA is structured as a double-stranded helix featuring a sugar-phosphate backbone that provides stability and structural integrity to the molecule.

2. Antiparallel Strands: One strand runs in the 5’ to 3’ direction while the complementary strand runs 3’ to 5’, a feature essential for replication and transcription processes.

3. DNA History: Watson and Crick's model was pivotal, integrating findings from Rosalind Franklin (who provided critical data on DNA's helical shape) and Erwin Chargaff (who introduced base pairing rules).

4. Distinction between DNA and RNA:

   • RNA: Ribonucleic acid, typically single-stranded with A-U and C-G pairing.

   • DNA: Deoxyribonucleic acid, double-stranded with A-T and C-G pairing.

DNA in Prokaryotic and Eukaryotic Cells

• Eukaryotic DNA: Resides within the nucleus and is organized into linear chromosomes, each containing numerous genes.

• Prokaryotic DNA: Found in the nucleoid region, consisting of circular chromosomes; prokaryotes often also contain plasmids—small, circular DNA strands that can confer advantages such as antibiotic resistance.

Plasmids

Plasmids replicate independently from chromosomal DNA. Their manipulation in laboratory settings is a cornerstone of recombinant DNA technology, facilitating gene cloning and gene expression when reintroduced into bacterial cells.

Key Differences

Chargaff's Rule: The observation that A=T and C=G underlines the complementary nature of the DNA structure.

Responses to environmental cues determine when genes are expressed, which is essential for cellular functions and specialization in multicellular organisms.

Lesson 2: DNA Replication

Aim: Understand the intricate mechanisms of DNA replication essential for cellular division.

Key Concepts:

• Replication fork: The junction where the double-stranded DNA is unwound, allowing new strands to be synthesized.

• Okazaki fragments: Short DNA sequences synthesized on the lagging strand, later joined together.

• Leading and Lagging Strands: The leading strand is synthesized continuously, while the lagging strand is synthesized in fragments.

• Role of helicase: Unwinds the double helix, facilitating access for replication machinery.

• DNA polymerase: Synthesizes new DNA strands by adding nucleotides complementary to the template strand.

DNA Replication Models

1. Conservative Model: Suggests that parental strands remain intact and serve as templates for new strands.

2. Semi-Conservative Model: Each daughter DNA molecule contains one original (parental) strand and one new strand; this was confirmed by Meselson and Stahl's experiments.

3. Dispersive Model: Proposes that parent DNA is dispersed among daughter strands.

Steps in DNA Replication

1. Initiation: Begins at specific points called origins of replication, recognized and bound by various initiation proteins.

2. Unwinding: Helicase unwinds the double-stranded DNA; single-strand binding proteins prevent the strands from re-annealing.

3. Primer Addition: Primase synthesizes RNA primers, which are necessary for the initiation of DNA synthesis by DNA polymerase.

4. Antiparallel Elongation: DNA polymerase III synthesizes new strands by adding nucleotides in a 5’ to 3’ direction. The leading strand is synthesized continuously, while the lagging strand is synthesized in short fragments.

5. Ligase Function: DNA ligase joins Okazaki fragments together, ensuring continuous DNA strands.

6. Proofreading and Repair: DNA polymerase has proofreading capabilities to ensure correct nucleotide pairing; mismatch repair enzymes handle post-replication errors.

Lesson 3: Transcription

Aim: Grasp the processes of gene transcription and how proteins are synthesized based on DNA schemes.

Key Processes:

The Central Dogma of Molecular Biology: DNA transcribes to RNA, which then translates to protein.

Transcription Steps

1. Initiation: RNA polymerase binds to the promoter region, a specific sequence that indicates where transcription should begin.

2. Elongation: RNA polymerase synthesizes RNA, elongating it by adding RNA nucleotides complementary to the DNA template strand (the RNA synthesis proceeds in a 5’ to 3’ direction).

3. Termination: The enzyme eventually reaches a termination sequence, concluding RNA synthesis; the pre-mRNA is then modified by adding a 5' cap, a poly-A tail, and splicing out introns to form mature mRNA.

   1. 5’cap: A modified guanine nucleotide added to the 5' end of pre-mRNA, facilitating mRNA stability, export from the nucleus, and recognition by the ribosome during translation.

   2. Poly-A tail: A long sequence of adenine nucleotides added to the 3' end of the pre-mRNA, which enhances stability and regulates the lifespan of the mRNA in the cytoplasm.

   3. Splicing: The process of removing introns and ligating exons together in the pre-mRNA, resulting in a continuous coding sequence that is crucial for proper translation into proteins.

Types of RNA

1. mRNA: Carries genetic information from DNA to ribosomes, serving as a template for translation.

2. tRNA: Transfers specific amino acids to the ribosome during translation, having an anticodon that is complementary to mRNA codons.

3. rRNA: Together with proteins, forms ribosomes, the cellular machinery responsible for protein synthesis.

Lesson 4: Translation

Aim: Learn the essential mechanisms of protein synthesis through translation at the ribosome.

Three Stages of Translation:

1. Initiation: The process begins with the binding of mRNA and tRNA to the ribosome, marking the start codon.

2. Elongation: Comprises codon recognition (tRNA binding to mRNA), peptide bond formation (joining amino acids), and translocation (the movement of the ribosome along the mRNA).

3. Termination: Triggered by a stop codon, resulting in the release of the complete polypeptide chain.

Components of Translation

• Ribosome: Composed of large and small subunits, consisting of two major sites: A site (amino acid site), P site (polypeptide site), and E site (exit site) for tRNA.

• tRNA Structure: Contains an anticodon complementary to the mRNA codon and carries its respective amino acid, facilitating the translation process.

Lesson 5: Gene Regulation

Aim: Comprehend the principles of gene regulation in both prokaryotic and eukaryotic organisms, emphasizing operon functionality in bacterial gene control.

Bacterial Gene Expression

• Operons: Groups of genes regulated together, essential for controlling gene expression (including promoter and operator regions along with structural genes).

• Feedback Mechanisms: Operons can be repressible (e.g., in the synthesis of tryptophan) or inducible (such as in lactose metabolism), allowing bacteria to adapt metabolically to environmental changes.

  • lac operon: An example of an inducible operon, where the presence of lactose in the environment activates the transcription of genes needed for lactose metabolism by removing the repressor from the operator site.

  • tryptophan operon: An example of a repressible operon, where the presence of tryptophan in the environment inhibits the transcription of genes involved in its synthesis, thereby regulating its own production.

Eukaryotic Gene Expression

• Regulation at Multiple Levels: This includes chromatin remodeling, transcriptional regulation, mRNA processing, and post-translational modifications.

  • histone acetylation, a key modification that influences gene expression by altering chromatin structure, making DNA more or less accessible for transcription.

• Epigenetic Inheritance: Involves heritable changes in gene expression that do not alter the underlying DNA sequence, influenced by environmental factors and experiences.

Lesson 6: Mutations & Biotechnology

Aim: Investigate the effects of mutations on phenotypes and explore various biotechnology applications.

Types of Mutations

1. Point Mutations: Affect a single nucleotide, categorized as silent (no change), missense (change of one amino acid), or nonsense (introduces a premature stop codon).

2. Frameshift Mutations: Result from insertions or deletions of nucleotides that alter the original reading frame, potentially resulting in completely different proteins.

3. Large Scale Mutations: Affect entire chromosomal structures through nondisjunction events or translocations that can lead to diseases or new traits.

Biotechnology Applications

• Restriction Enzymes: Vital tools for genetic engineering; they cut DNA at specific sequences, enabling the removal or addition of genes.

• DNA Cloning and Sequencing: Techniques for amplifying specific DNA segments and determining their nucleotide sequences, crucial for numerous genetic analyses.

• PCR for Amplification: Polymerase Chain Reaction (PCR) is a powerful method used to exponentially amplify a specific DNA sequence, facilitating further analysis and experimentation in genetics, biotechnology, and forensic science.