GEN 3022 1st Edition Lecture 22 Outline of Last Lecture I Introduction a Eukaryotes and gene regulation b Necessity of gene regulation II Transcriptional Factors a General Transcription factors b Regulatory Transcription factors i Control elements ii Activators and repressors iii Enhancers and silencers c Combinatorial control i Common factors that contribute to combinatorial control d Structural features of regulatory transcription factors i Domains ii Motifs e Variation in regulatory transcription factor functions i Three main functions f Steroid receptors i Action of steroid hormones These notes represent a detailed interpretation of the professor s lecture GradeBuddy is best used as a supplement to your own notes not as a substitute ii Response of the cell to steroid hormones 1 Glucocorticoids 2 Gonadocorticoids g CREB protein i Mechanism ii cAMP response element CRE III Chromatin remodeling and histones a Purpose b Structure of chromatin i Closed conformation ii Open conformation iii Nucleosomes c ATP dependent chromatin remodeling d Histones i 5 histone genes ii Humane histone genes iii Specialized chromatin e Histone code i Modification of histones in mammals ii Protein recognition and binding IV DNA methylation a DNA methylation in eukaryotes b CpG islands i Housekeeping genes ii Tissue specific genes c Heritability V Insulators a Purpose b Mechanism Outline of Current Lecture I DNA methylation a Maintenance of methylation b Unmethylated CpG islands c Housekeeping genes II Regions of the lac operon a Disruption of 10 and 35 sequences b Lac repressor binding site c Product of lacI III Functions of the lac operon a Inducer of the lac operon b Production of mRNA transcripts IV Transcription factors a Inducible system b Molecules that induce transcription c Modulation of regulatory transcription factors d DNA binding sites V DNA modulation a Histone acetylation b ATP dependent chromatin remodeling c Insulators d Histone code VI Gene expression in bacteria eukaryotes VII Mutations of the lac operon a Three mutation possibilities b Most likely mutation Current Lecture I DNA methylation a Maintenance of methylation maintenance methylation refers to DNA methylation patterns that are maintained during replication of somatic cells b Unmethylated CpG islands generally when CpG islands are unmethylated it means that chromatin in the promoter region is open allowing access of transcription factors and RNA polymerase c Housekeeping genes housekeeping genes are constitutively expressed II Regions of the lac operon the regions of the lac operon are lacI lacO lacP lacZ lacA and lacY a Disruption of 10 and 35 sequences most likely due to a mutation in lacP because 10 and 35 are promoter sequences b Lac repressor binding site in the absence of lactose the lac repressor is bound to the lacO site III Functions of the lac operon a LacI function the most convincing evidence that the product of lacI can work in trans is a merozygote with only one functional lacI gene is able to regulate both copies of the lac operon b Inducer of the lac operon allolactose is considered the inducer of the lac operon c Production of mRNA transcripts transcription of the lac operon results in the production of only one mRNA transcript IV Transcription factors a Inducible system in an inducible system there is a repressor protein that is bound to DNA in the absence of any other factor b Molecules that induce transcription two molecules that work to induce transcription are a repressor and an inducer An example would be the lac operon repressor and its inducer allolactose c Modulation of regulatory transcription factors regulatory transcription factors can be modulated by the binding of small effector molecules inducer inhibitor protein protein interactions and covalent modifications d DNA binding sites DNA sites that bind regulatory transcription factors are cis acting V DNA modulation a Histone acetylation histone acetylation can loosen chromatin packing because it decreases the positive charge on the R group of lysine which weakens the bond between DNA and the histone proteins b ATP dependent chromatin remodeling complexes these complexes are responsible for the change in location of nucleosomes eviction of nucleosomes from DNA and replacement of standard histones with histone variants c Insulators insulators function by defining the boundaries in chromatin and limiting the extent of chromatin changes that occur ex acetylation They can also block activity of enhancers d Histone code the pattern of histones that are recognized by specific proteins These proteins bind to the DNA to regulate transcription through chromatin remodeling VI Gene expression in bacteria eukaryotes the four levels of gene expression that are regulated in bacteria eukaryotes are transcription RNA processing translation and post translational modifications VII Mutations of the lac operon a strain of E coli produces B galactosidase when lactose is present and does not produce it when lactose is not present This is the wild type phenotype The mutant phenotype does not produce B galactosidase regardless of the presence of lactose Both the mutant and the merozygote must be considered for this scenario a Four mutation possibilities the mutation could occur at any region of the lac operon except lacO A mutation in lacP would result in no expression of the operon under any circumstances because the promoter could not bind to the operon A mutation in lacZ would result in no production of B galactosidase A mutation in lacI would result in no recognition of allolactose which would mean that the repressor is always bound to the operator A mutation in lacY would result in no production of permease and thus no import of lactose into the cell b Most likely mutation the most likely mutation is the mutation of the lacI region because it would result in the repressor always being bound to the operator This mutation would function in trans instead of cis