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Control of Gene Expression

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Control of Gene Expression Control of! Gene Expression Heterochromatin vs Euchromatin • heterochromatin • euchromatin – tight coils of DNA – loose coils of DNA Regulatory Elements in Gene Expression • promoter (TATA) • transcription factors • enhancers/activators • repressors Prok vs Euk ! DNA arrangement 1. 1x circular DNA 1. multiple, linear chs 2. 1x origin of 2. multiple origins replication 3. no operons 3. operon control 4. many introns 4. no introns 5. modifications (*methylation) 6. transposons Prok vs Euk ! Transcription & Translation 1. tcn & tln in cytoplasm 1. tcn (nucleus); tln (cytoplasm) 2. no post-transcriptional 2. post-transcriptional modifications modifications (methylation, 5’cap & 3’tail) 3. no introns 3. introns spliced out 4. 70S ribosome 4. 80S ribosome • RNA polymerase • aminoacyl synthase (adds aa to tRNA) • transcription factors • peptidyl transferase (makes peptide bonds) lac Operon • inducible – always “OFF” (negative control) • turned ON by inducer (lactose); inactivates the repressor lac Operon - cast of characters • promotor TATA box, transcription factors, RNA polymerase • lacI – gene for repressor • lacZ, lacY, and lacA - genes for metabolizing lactose (galactosidase, permease, transacetylase) • inducer - lactose sugar trp Operon • repressible – always “ON” (positive control) • turned OFF by negative feedback - product tryptophan binds to repressor lac Operon model *grocery shopping • repressor – credit card bill • inducer – paycheck • gene A – taking money out of an ATM • gene B – driving to a supermarket • gene C – pushing a shopping cart lac operon model cont’ *Control of Shopping • turned “OFF” credit card bill (repressor) is high • turned “ON” monthly paycheck (inducer) arrives • household bills (glucose) are high, very little appetite (cAMP) is formed, and therefore no desire for food • low appetite (low cAMP-CAP), no grocery-shopping lac Operon - cast of characters • promotor TATA box, transcription factors, RNA polymerase • lacI – gene for repressor • lacZ, lacY, and lacA - genes for metabolizing lactose (galactosidase, permease, transacetylase) • inducer - lactose sugar cAMP-CRP complex (glucose or no glucose) • low glucose = high cAMP; CRP binds to lac gene, operon turned “ON” • high glucose = low cAMP; CRP cannot bind; operon turned “OFF” trp Operon • repressor – activated when tryptophan binds, turns transcription OFF DNA Cloning Transgenic organisms Mutant plants Big, juicy tomatoes… Bug-resistant plants… Mutant Fish glow-in-the-dark zebrafish Mutant Mammals hairless mice - test skin diseases obese mice - test diets Mutant Mouse mouse with human ear grown on its back Mutant Fly normal fly legs on its head Replication/Transcription/ Translation Mutations Base substitution (i.e. point mutations): • silent (same-sense) – no aa change • missense – aa changed – conservative – aa change is not essential *nonpolar to nonpolar • nonsense – aa changed to a stop codon Frame-shift – insertion or deletion changes reading frame Beadle & Tatum Expt Neurospora fungi exposed to X-ray radiation; mutants grew only if nutrients are added DNA Gene 1 Gene 2 Gene 3 | | | | mRNA mRNA 1 mRNA 2 mRNA 3 | | | | enzyme 1 2 3 | | | | substrate A à B à C à D Beadle & Tatum Expt —1 gene, 1 enzyme Biochemical Pathways Gene B is mutated. What compound is needed to get final product 5? (1, 2, 3, 4, or 5?) Mutant Pathway Problem (a) •5 mutants cannot synthesize compound G to grow •Order of compounds A to E in the pathway? Mutant Pathway Problem (b) • compounds used latest in the pathway will support the growth of the most mutants .
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