BBMB 405 1st Edition Lecture 36 Outline of Last Lecture XVII Chapter 31 The control of gene expression in prokaryotes A Introduction B Many DNA binding proteins recognize specific DNA sequences C Prokaryotic DNA binding proteins bind specifically to regulatory sites in operons Outline of Current Lecture XVII Chapter 31 The control of gene expression in prokaryotes D Gene expression can be controlled at posttranscriptional levels XVIII Chapter 32 The control of gene expression in eukaryotes A Eukaryotic vs Prokaryote transcriptional regulation B Eukaryotic DNA is organized into chromatin C The control of gene expression can require chromatin remodeling Current Lecture XVII Chapter 31 The control of gene expression in prokaryotes D Gene expression can be controlled at posttranscriptional levels 1 Attenuation Co translational control of transcription a Attenuation turn off gene because metabolite is at high enough concentrations that don t need be synthesized example trp b High concentrations of Trp present in cell terminator sequences added and translation stopped c Low concentrations of Trp present in cell then ribosome pauses and something prevents stop codon from forming so Trp can be translated d Mutually exclusive share some of the same genes e Terminator and alternate stem loop are mutually exclusive 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 f At high levels of tryptophan after completion of leader peptide the third and fourth elements create a stem loop which is a transcription terminator signal At low levels of tryptophan when leader peptide is not fully completed the second and third elements create an alternate stem loop and transcription continues Can you guess what types of genes these leaders are from Many amino acid biosynthesis operons contain attenuators a Threonine operon b Phenylalanine operon c Histidine operon XVIII Chapter 32 The control of gene expression in eukaryotes A Eukaryotic vs Prokaryotic transcriptional regulation 1 Commonalities DNA binding proteins that activate or repress genes 2 Differences Eukaryotes Prokaryotes DNA packaged in chromatin DNA relatively accessible Large genomes Compact genomes Generally no operons Genes often in operons Multiple transcription factors per gene Single factor can control several genes Regulatory DNA sequences often distal from promoter Operator sequences are close to promoter Cell type specific regulation multicellular Generally nutrient specific regulation B Eukaryotic DNA is organized into chromatin 1 Eukaryotic DNA is organized and compacted a Nucleosome is fundamental unit of organization Octomeric 8 histone protein subunits 147 bp DNA wrapped around Sites of chemical modification for epigenetic regulation b Nucleosomes organize into chromatin fiber Left handed superhelix Greatly compacts long strands of DNA DNA accessibility can regulate transcription c Higher order structures form full chromosomes 2 Four types of histones make up nucleosome core a Highly homologous and adopt very similar 3D conformations b Octomer contains H3 2 H4 2 tetramer and two H2A H2B dimers c Histones are highly basic rich in Arg and Lys 3 Assembly of nucleosome core histones H3 H4 H3 H4 dimer H3 H4 dimer H3 H4 tetramer H2A H2B H2A H2B dimer Two H2A H2B dimer two H3 H4 tetramer Histone Octamer 4 Nucleosomes organize DNA a 147 bp of DNA wraps around histones in left handed solenoid arrangement b Nucleosomes store negative supercoils DNA is under wound easier to unwind during transcription and replication 5 Nucleosomes are packaged into chromatin fibers a Left handed superhelix b Beads on string condense through internuclosome interactions tetranucleosome are two stacks of nucleosomes connected by straight linker DNA c Histones H1 and or H5 are linkers not part of nucleosome core located in interior of fiber 6 Chromatin structure strongly impacts transcription a DNase I cleavage of chromatin b Non transcribed DNA is densely packed and inaccessible to nuclease and DNA binding proteins c Actively transcribed DNA is packaged less densely and can be susceptible to cleavage by nuclease d Chromatin structure must be relaxed to enable gene expression 7 Hypersensitive sites to DNA I high levels of degradation when sites being expressed reveal what conditions gene is being expressed C The control of Gene expression can require chromatin remodeling 1 Chromatin remodeling dictates transcription activation at specific sites yeast transcription factor GAL4 a Controls gene involved in galactose metabolism b DNA recognition sequence 5 CGG N 11CCG 3 c Contains two zinc finger DNA binding domains only contact 5 CGG 3 sequences in major grooves d About 4000 potential binding sites in yeast genome 2 Chromatin immunoprecipitation ChIP can reveal binding sites for transcription factors a ChIP experiments for GAL4 revealed only 10 out of 4000 potential binding sites were bound by GAL4 in yeast grown in galactose media b 99 of sites are blocked presumably by chromatin structure 3 Mechanisms for altering chromatin structure a Covalent modification of histone N terminal tails histone acetyltransferase transfers Acetyl from acetyl CoA onto N terminal tails they no longer has positive charge and don t bind to DNA backbone as well b Movement restructuring or removal of nucleosomes chromatin remodeling complex uses remodeling protein and ATP to remove histones 4 Histone acetylation alters interactions with DNA a Loosens interaction between histone and DNA b Recruits bromodomain acetyllysine binding domain containing proteins 5 Chromatin remodeling complexes or engines 6 Histones can be modified in multiple ways 7 Transcription factors a Enhance or decrease transcription initiation General transcription factors promote low levels of transcription Activators increase initiation from specific sites Repressors decrease initiation at specific sites b Multiple domains DNA binding domains specific recognition of cis regulatory elements Activation domains interact with transcription machinery often indirectly through bridged interactions or remodel chromatin c Multiple eukaryotic transcription factors work together to control transcription Can exhibit functional redundancy modular interactions and synergistic activity These features lead to combinatorial control 8 Major groove recognition by DNA binding protein major groove is wide and shallow also can make more