Week%209-21 - This week office hour: Wed 9/23 5:00-6:00 pm...

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Unformatted text preview: This week office hour: Wed 9/23 5:00-6:00 pm Thur 9/24 5:00-6:00 pm RRI 3rd FL breakroom (across from 301) Fewer or more than one centromere is highly detrimental Differences in centromeres of eukaryotes The eukaryotic mitotic cell cycle • During the Gap phases (G1 and G2) checkpoints ensure that all is well with the DNA and segregation machinery, before allowing DNA replication (S phase) and mitosis (M) to commence. Various levels of chromatin compaction during cell cycle Chromosome Cohesion and Condensation by SMC Proteins • SMC proteins: the Structural Maintenance of Chromosomes proteins S Phase • Individual replication origins initiate replication and produce two replication forks per origin. • During replication cohesin proteins assemble around the new DNA molecules, holding them together (cohesion). • The fully replicated DNA molecules are called sister chromatids. M Phase • Sister chromatids separate by the end of M pahse. Mitosis • Sister chromatids are segregated to daughter cells. • Checkpoints regulate various steps, especially the metaphase to anaphase transition to ensure accuracy. Meiosis • After DNA replication, but before chromosome segregation, homologous (maternal and paternal) chromosomes pair and recombine. Two rounds of chromosome segregation follow. In meiosis I, homologous chromosomes segregate. In meiosis II, sister chromatids segregate (just like in mitosis). • • • Chromatin structure Outline • nucleosome structure and assembly • chromatin structure and regulation Reading assignments: Molecular Biology of the Gene (Watson, 6th ed) chapter 7 p157-192 Bacterial DNA is negatively supercoiled • DNA gyrase introduces the negative supercoil to compact DNA in prokaryotes. • Coated by polyamines and some basic proteins. Eukaryotic chromatin is compacted by histones in negative supercoils • Histones are major chromosomal proteins in enkaryotes. • There are five major histones: H1, H2A, H2B, H3 and H4. • They are small, highly basic, very abundant proteins, and are among the most conserved proteins in evolution. Histones are small, basic proteins • Most of histone-DNA interactions are mediated by Hbonds between the protein and the oxygen atoms in the phosphodiester backbone. Nucleosomes are the building blocks of eukaryotic chromosomes • 147 bp of DNA wrapped around a histone core 1.65 times. • Between each nucleosome is a region of DNA called the linker, which varies in size, typically 20-60 bp. • Compact DNA sixfold. Analysis of nucleosomal structure with Micrococcal Nuclease • micrococcal nuclease (MNase) is a non-specific double-strand endonuclease that cleaves preferentially in regions lacking DNA binding proteins. The core histones share a common structural feature (Motif) The histone core has eight subunits Histone H1 tightens the nucleosomal structure 10-nm fiber 30-nm fiber Compact DNA by 40-fold Two models for the 30-nm chromatin fiber • The solenoid forms a helical structure with a short linker DNA. • The zigzag has a longer linker. Further compaction of chromatin • 30 nm filaments are further organized into loops of 40-90 kb. • The nuclear scaffold (TOPO II, SMC proteins) stabilize the loops. Compaction of chromatin Histone variants functionally distinguish DNA regions • CENP-A replaces H3 in centromeric region nucleosomes, allows binds to the kinetochore. Nucleosome obstructs access to DNA of sequence-specific binding proteins • DNA binding proteins generally prefer to bind naked DNA. Nucleosomes can be moved to enable binding site accessibility Nucleosome remodeling proteins use the energy of ATP to alter the association of the DNA and nucleosome particle. Nucleosome positioning in vivo • Nucleosomes are sometimes positioned or excluded by DNA binding proteins to maintain access to a region of DNA, such as promoter or replication origin, or to prevent the access to certain DNA sequence. Histone tails make contacts with the DNA • The histone tails interact with the DNA to stabilize the nucleosome. Modification of histone tails Regulation of chromatin function by modification of histone tails • In general, acetylation is associated with gene expression and phosphorylation of H3 with chromosome condensation. Chromatin remodeling complexes and modifying enzymes cooperate in regulating gene expression • DNA binding proteins recruit remodeling complexes and modifying enzymes to chromatin. • The function of one complex can often facilitate the interaction of the other. Nucleosome assembly occurs during DNA replication • Old histones are reused with H3-H4 tetramers remaining randomly associated with one of the two daughter strands of DNA. • Old H2A-H2B dimers are displaced and compete with new dimers for assembly into nucleosomes. Chromatin assembly factors help assemble the new nucleosome • Also called histone chaperones. • Negatively charged, form complexes with the nucleosome subunits to prevent them from interacting with DNA nonproductively. Inheritance of parental H3-H4 tetramers facilitates inheritance of chromatin states Midterm Friday (9/25) • Attend your registered section. • Exam based on lectures. • Room assignment posted on Blackboard. •Bring your ID. •Use a pen to answer the questions. ...
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This note was uploaded on 10/20/2009 for the course BISC 320L taught by Professor Baker,aparicio during the Spring '07 term at USC.

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