Regulation of metabolic pathways Bacterial control of gene

Regulation of metabolic pathways

Bacterial control of gene expression Operon: Operon cluster of related genes with on/off switch Three Parts: 1. Promoter – where RNA polymerase attaches 2. Operator – “on/off”, controls access of RNA poly 3. Genes – code for related enzymes in a pathway

Regulatory gene: gene produces repressor protein that binds to operator to block RNA poly

Repressible Operon (ON OFF) Inducible Operon (OFF ON)

Repressible Operon Normally ON Anabolic (build organic molecules) Organic molecule product acts as corepressor binds to repressor to activate it Operon is turned OFF Eg. trp operon

trp operon

Inducible Operon Normally OFF Catabolic (break down food for energy) Repressor is active inducer binds to and inactivates repressor Operon is turned ON Eg. lac operon

lac operon

• Typical human cell: only 20% of genes expressed at any given time • Different cell types (with identical genomes) turn on different genes to carry out specific functions • Differences between cell types is due to differential gene expression

Eukaryotic gene expression regulated at different stages

Chromatin Structure: • Tightly bound DNA less accessible for transcription • DNA methylation: methyl groups added to DNA; tightly packed; transcription • Histone acetylation: acetyl groups added to histones; loosened; transcription

Transcription Initiation: • Control elements bind transcription factors • Enhances gene expression

Transcription Initiation Complex Enhancer regions bound to promoter region by activators

Regulation of m. RNA: • micro RNAs (mi. RNAs) and small interfering RNAs (si. RNAs) can bind to m. RNA and degrade it or block translation

Embryonic Development: Zygote Organism 1. Cell Division: large # identical cells through mitosis 2. Cell Differentiation: cells become specialized in structure & function 3. Morphogenesis: “creation of form” – organism’s shape

Determination: irreversible series of events that lead to cell differentiation

• Cytoplasmic determinants: maternal substances in egg distributed unevenly in early cells of embryo

• Induction: cells triggered to differentiate • Cell-Cell Signals: molecules produced by one cell influences neighboring cells – Eg. Growth factors

Pattern formation: setting up the body plan (head, tail, L/R, back, front)

Morphogens: substances that establish an embryo’s axes

Homeotic genes: master control genes that control pattern formation (eg. Hox genes)

Control of Cell Cycle: 1. Proto-oncogene = stimulates cell division 2. Tumor-suppressor gene = inhibits cell division • Mutations in these genes can lead to cancer

Proto-oncogene Oncogene

Genes involved in cancer: • Ras gene: stimulates cell cycle (protooncogene) – Mutations of ras occurs in 30% of cancers • p 53 gene: tumor-suppresor gene – Functions: halt cell cycle for DNA repair, turn on DNA repair, activate apoptosis (cell death) – Mutations of p 53 in 50+% of cancers

• Cancer results when mutations accumulate (5 -7 changes in DNA) • Active oncogenes + loss of tumorsuppressor genes • The longer we live, the more likely that cancer might develop

Bacteria vs. Viruses Bacteria • Prokaryotic cell • Most are free-living (some parasitic) • Relatively large size • Antibiotics used to kill bacteria Virus • Not a living cell (genes packaged in protein shell) • Intracellular parasite • 1/1000 size of bacteria • Vaccines used to prevent viral infection • Antiviral treatment

Viruses • Very small (<ribosomes) • Components = nucleic acid + capsid – Nucleic acid: DNA or RNA (double or single-stranded) – Capsid: protein shell – Some viruses also have viral envelopes that surround capsid • Limited host range (eg. human cold virus infects upper respiratory tract) • Reproduce within host cells

Simplified viral replicative cycle

Bacteriophage • Virus that infects bacterial cells

Lytic Cycle of T 4 Phage

Bacteriophage Reproduction • Lytic Cycle: – Use host machinery to make copies of virus – Death of host cell by rupturing it (lysis) lysis – Virulent phages replicate by this method • Lysogenic Cycle: – Phage DNA incorporated into host DNA and replicated along with it – Phage DNA = prophage • Temperate Phage: uses both methods of replication

Lytic Cycle vs. Lysogenic Cycle

Animal viruses have a membranous envelope • Host membrane forms around exiting virus • Difficult for host immune system to detect virus

Retrovirus • RNA virus that uses reverse transcriptase (RNA DNA) • Newly made viral DNA inserted into chromosome of host • Host transcribes viral DNA (= provirus) provirus to make new virus parts • Example: HIV (Human Immunodeficiency Virus)

HIV = Retrovirus

HIV ◦ Infects white blood cells ◦ HIV+: provirus (DNA inserted) ◦ AIDS: active viral reproduction

Vaccines • Weakened virus or part of pathogen that triggers immune system response

Emerging viruses = mutation of existing viruses

Viroids • Small, circular RNA molecules that infect plants • Cause errors in regulatory systems that control plant growth • Eg. coconut palms in Philippines

Prions • Misfolded, infectious proteins that cause misfolding of normal proteins • Eg. mad cow disease (BSE), Creutzfeldt. Jakob disease (humans), scrapie (sheep)

Diseases caused by prions • Prions act slowly – incubation period of at least 10 years before symptoms develop • Prions are virtually indestructible (cannot be denatured by heating) • No known cure for prion diseases

Tools of Genetic Engineering S Restriction enzymes (restriction endonucleases): used to cut strands of DNA at specific locations (restriction sites) S Restriction Fragments: have at least 1 sticky end (singlestranded end) S DNA ligase: joins DNA fragments S Cloning vector: carries the DNA sequence to be cloned (eg. bacterial plasmid)