Biochem 8-28-09

-------------------------------BIOCHEM 8-2809-----------------------------------------------------------------Releasing factor binds to stop codon Spliceosomes primarily which snRNA? Lac operons consist of: promoter, initiator, proteins Definition of some word: I forget I.

II.

III.

IV.

Immediate and delayed response to environment (transcription) a. Transcriptional regulation of genes b. mRNA stability c. alternate splicing/ alternate transcripts Long term responses a. Gene rearrangement b. Gene amplification c. Mutations/recombination Definitions a. Gene is expressed: transcribed b. Levels of expression = amount of mRNA c. Constitutive expression: always transcribed d. Regulated expression: modulated transcription (up or down) Regulattion of gene transcription a. Major mechanisom for regulating b. Lac operon: coordinated unit of gene expression i. Regulator genes ii. Operator sitex iii. Structural genes 1. CHART 2. Organization: promoter region, operator, z,y a gene 3. Proteins: z: Beta galactosidase, Y: permease A: transacetylase 4. Function: lactose -> glucose + galactose _> atp c. E.Coli can use glucose or lactors i. Prefers to use glucose as a source of carbon and energy ii. Can use lactose due to ability to synthesize B galactidase from the lac operon iii. Converts lactose to glucose and galactose d. Absence of lactose i. Repressor bound to operator site prevents transcription of z,y,a 1. 4x106 as strong as to random sites on DNA e. Lactose present i. Repressor-inducer complex doesn’t bind DNA ii. B galactiosidase, permease, and transacetylase transcribed f. Inducer i. When glucose is absent and lactose present, some lactose converts to 1,6 allolactose (inducer) ii. Inducer binds to the repressor and inactivates repressor iii. Induder = alpha-1,6 linked galactose + glucose iv. Allows RNA polymerase to bind to the promoter

g. Catabolite repression: if both galactorse and glucose present i. Levels of glucose are important 1. Alter cAMP levels 2. Repressor and inducer bind in the absence of glucose to RNA polymerase to transcribe Z, Y, A h. Carbolite Repression i. Absence of glucose leads to increase in cAMP ii. cAMP binds catabolite activator protein (CAP; cAMP response protein = CRP) iii. Polymerase stabilized; transcription stimulated x50 iv. Lactose and glucose present 1. cAMP levels low since glucose present 2. CAP doesn’t bind to polymerase 3. Very little or no transcription of lac operon genes v. Lactose but no glucose 1. Inducer binds repressor preventing binding to promoter 2. cAMP levels high since no glucose 3. Cap binds to & stabilizes polyhmerase 4. Transcription of lac genes occurs i. Tryptophan operon i. Post transcriptional regulation ii. Transcription/translation tightly coupled in bacteria, thus translation occurs as soon as Shine-Delgarno sequence transcribed iii. Encodes 5 enzymes that convert chorismate into tryptophan iv. Trp mRNA contains short open reading frame 1. Contains 14 aa peptide + untranslaged attenuator sequence v. Tryptophan never use in transcription, practically 1. Procedes attenuator region: can have 2 confirmations vi. High [Trp} 1. If high levels available, the ribosome passed quickly and the termination turn forms 2. Don’t remember region; remember 2 different conformations vii. Low [Trp] 1. When trp levels low, ribosomes stall at trp codons, allowing the alternative turn to fomr, preventing the formation of the termination turn j. Post transcriptional regulation i. Other attenuated operons include: threonine, phenylalanine, and histidine ii. Can’t exist in euk: transcription/translation separated k. SUMMARY; i. Several types of mechanisms can regulate gene expression ii. In absence of lactose, repressor binds to lac operon iii. Inducer iv. Catabolite v. Attenuation

------------------------------------------EUKARYOTIC GENE EXPRESSION------------------------------------------I.

Eukaryotic gene expression a. Larger genomes require greater complexity of regulation b. Different cell types requires increased levels of regulation c. Genes are NOT organized in operons d. Transcription and translation are NOT coupled (mRNA modificiation ETC) e. Tissue specific Gene expression i. Negative or positive regulation by reg. molecule can alter gene exp. ii. Most euk. Genes not express are unless “turned on” iii. Program in ind. Cells to control. Exp. Of specific genes iv. Development requires program of gene exp;. 1. Genes inovled in dev. Are ofthen transcription factors 2. Cells can signal each other and modify gen exp. Depending on position (positional information) v. Most occurs at level of transcription f. Structure of euk. Gene and products i. Upstream enhancers (can be downstream) 1. Proteins combine with it and loop around to interact with polymerase ii. Promoter iii. TATA box iv. 5’ UTR to 3’ UTR -> 1. Initial transcript still in nucleaus: 5’ cap and 3’ poly-A tail -> a. Introns removed -> i. Final mRNA in cytoplasm g. Regulation of gene expression i. Mostly at the transcriptional level 1. DNA -> a. Transcriptional control 2. Primary RNA transcript -> a. RNA processing control 3. mRNA travels through nuclear pore a. Translation control 4. Protein a. Protein activity control h. Althering Genes available for transcription i. Chromatin remodeling ii. Methylation of DNA 1. Form 5 methylcytosine located in GC islands (GC-rich region) 2. Globin gene highly methylated in non-erythroid cells 3. Important in fetal development iii. Gene rearrangement

1. iv. Gene 1. 2.

Ig genes amplification Not normally physiological mechanism Occurs in response to stimuli a. DHFR amplification in response to methyl substiture