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Chromatin Structure & Genome Organization

Chromatin Structure & Genome Organization. Overview of Chromosome Structure. Nucleosomes ~200 bp DNA in 120 Å diameter coil 3.4 Å /bp x 200 = 680 Å 680/120 = 5X compaction 30nm fiber Coil of nucleosomes w/ 6/turn 1200bp/120 Å vs 4080 Å = 34X compaction Chromatin loops

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Chromatin Structure & Genome Organization

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  1. Chromatin Structure & Genome Organization

  2. Overview of Chromosome Structure • Nucleosomes • ~200 bp DNA in 120Å diameter coil • 3.4Å/bp x 200 = 680Å • 680/120 = 5X compaction • 30nm fiber • Coil of nucleosomes w/ 6/turn • 1200bp/120Å vs 4080Å = 34X compaction • Chromatin loops • 30nm fiber is looped into 15-100Kbp loops • 7-8 loops form rosette w/ bases of loops attached to central core of scaffold proteins • 300Å/rosette, 800Kbp/rosette = 800kbp/300Å vs 2720000Å9000X compaction

  3. Nucleosomes • Core of 8 proteins, 2 of each • H2A • H2B • H3 • H4 • 166 bp of DNA • ~40bp between successive nucleosomes

  4. Nucleosomes • DNA associated with individual nucleosomes determined by DNAse I digestion

  5. Nucleosome • Each nucleosome contains 1 linker histone – H1 • Linker histones can vary with cell types and differentiation states

  6. Acetylation Phosphorylation Histone Post-translational Regulatory Modifications

  7. Histones!Meet the Histones!They're a basic protein family. There areJust four subtypes.They're conserved evolutionarily. Two eachOf each histoneDo their thing.FormingJust like beads upon a string. H1Binds the complex.And it inhibits new transcriptionAnd the creationOf new mRNA. The Histone Song (To the tune of "Flintstones")With Accompaniment

  8. Transcriptional Regulation by Histone Modification • Histone acetyltransferase is component of transcriptional activation complexes

  9. H1 Promotes Nucleosome Compaction • H1 function thought to be regulated by phosphorylation on N and C terminal tails -H1 +H1

  10. Heterochromatin vs Euchromatin • Constitutive heterochromatin • Never transcribed DNA • Facultative heterochromatin • DNA regions that can be switched between heterochromatin and euchromatin states • X inactivation • Chromo domain proteins • Bind to DNA keeping in heterochromatin state • Polycomb protein family in Drosophila

  11. Heterochromatin vs Euchromatin • Modifications to histones and DNA direct formation of heterochromatin • Deacetylation of H4 • Methylation of H3 • Methylation of CpG islands

  12. Loops & Chromosome Scaffolds • Chromosome scaffold • Proteins at base of loops • Potentially regulate formation of loops and/or regulate global gene expression • Components • DNA topoisomerase II • SMC proteins • HMGA • DNA elements • S/MARS • Sequences of DNA to which scaffold proteins bind • LCR • DNA elements known to regulate expression of large regions of DNA containing multiple genes • Boundary elements • scs elements flank puffs in Drosophila polytene chromosomes

  13. Chromosome Loops

  14. Chromosome Structure: The Metaphase Chromosome

  15. DNA sequence Repetitive DNA elements Constitutive heterochromatin Protein components CENP-A, B & C Centromeres

  16. Centromeres • Repetitive DNA of mammalian centromeres •  satellite DNA • 171 bp unit • Several specific unit sequences • Multiple repeats of these units form arrays

  17. Centromeres • Centromere of S. cerevisiae • CEN DNA is 125bp element • 3 regions - CDE I, II, & III • CDE's bound by special histone and other proteins and direct attachment of microtubule

  18. Telomeres

  19. Genome Organization • Sequence composition • Unique sequences • Genes • Low copy # elements • rRNA, 5S, tRNA genes • High copy # repetitive DNA • Retroelements • LINEs • SINEs • Simple Sequence DNA • Minisatellites (VNTR) • Microsatelites

  20. Retrotransposons, Retroelements & Transposons

  21. 23 – 9bp Repeats Mouse Satellite DNA Repeating unit given by restriction enzyme digestion 4 Imperfect Repeating Units

  22. VNTRs in DNA Fingerprinting

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