Sperm chromatin

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sperm chromatin structure

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Sperm chromatin

  1. 1. Mohammad Salehi Ph.DEmbryologist
  2. 2. Chromatin structure in somaticcell• DNA• Proteins• Histones: rich of lys & arg• H1, H2A, H2B, H3, H4• Non Histones• Acidic proteins• Hertons• Enzymes• Topoisomerase ,• Polymerases• Helicase• Neutral proteins• Scarcely RNA
  3. 3. DNA structure Nucleotide Base (A,T,C,G) Pentose sugar (deoxyribose) Phosphoric acidB DNA
  4. 4. Organization of chromatinstructure• First level• Nucleosome: cylindrical shape ( diameter: 10nm , height: 6nm)• Core particle contains an octamer of 2 eachof the core histones (H2A, H2B, H3 and H4)• 146 bp of DNA wrapped 1.75 turns• 10 nm fiber: Nucleosome + linker DNA (0-50bp)
  5. 5. Nucleosome
  6. 6. nucleosome- building blocks of chromosomesH2A: redH2B: yellowH3: purpleH4: greenHistones are small, positively-charged proteins
  7. 7. The nucleosome has an approximate twofold axis of symmetry
  8. 8. Higher-order chromatin structureH binds to linker DNA at one end ofThe nucleosome and the central DNA helix
  9. 9. The core Histone N-terminal tails are required for theformation of the 30-nm fiberThe tail of H2A, H3 and H4 interact with adjacent nucleosome
  10. 10. Organization of chromatinstructure• Second level• 30 nm fiber• Solenoid model• Zigzag (crossed linker)model• Nucleomer (super bead)model• Third level• Super coil
  11. 11. The addition of H1 leads to more compact nucleosomal DNAWithout H1
  12. 12. 30-nm fiberSuperhelix, 6 nucleosome per turn, supported by EM and X-ray studiesBased on zigzag pattern upon H1 addition, requires linker DNA to pass throughcentral axis,
  13. 13. Higher compaction of DNA involves large loops ofnucleosomal DNANuclear scaffold (Topo II, SMC)
  14. 14. How histones chaperones facilitate the assembly of nucleosomeduring DNA replication(sliding clamp)
  15. 15. Chromatin organization in spermDuring spermatogenesis spermatogonia undergomodification to produce spermatozoa that include:• Mitosis (spermatocytogenesis)• Meiosis• Spermiogenesis• Golgi phase• Cap phase• Acrosomal phase (chromatin condensation)• Maturation phase
  16. 16. Causes of condensation in spermchromatin• Reduction of size• Facilitate of sperm penetration into oocyte• Protection of genome from physical,chemical and biological damages• Reprogram of paternal genome• Coordination of cell cycle between spermand oocyte
  17. 17. Protein replacement inspermiogenesisTransitionalProteinsSomaticHistonesTesticularHistonesProtaminesIncrease basic property
  18. 18. Testicular Histones• Absence of H1• H2A• H2AZ• H2AX• H3 and H4 (hyperacetylation)• 15-20% remains in mature sperm
  19. 19. Mechanism of replacement• Histones acetylation(H4)• H2AX Phosphorylation• Ubiquitination
  20. 20. Transitional proteinsType of protein MW (KDa) Amino acidlengthPropertiesHPI1(TP1) 6 54 Rich of arg, lys, hisDNA destabilizationand promote of repairHPI2(TP2) 13 137 Rich of cys, arg, lysBond to CG andcessation transcriptionalactivityHPS1 N/A 69 P2 precursorHPS2 N/A 66 P2 precursor
  21. 21. ProtaminesType ofProtamineMW (Da) Amino acidlengthPropertiesHP1 6692.2 50 Rich of arg,cysHP2 7652.4 57 Rich of hisHP3 N/A 54 Rich of arg,cys, hisHP4 N/A 58 Rich of arg,cys, his
  22. 22. Amino acid sequence ofP1&P2Human P2 and P1Bovine P1Anchoring domainsHis, Arg, Arg sequences
  23. 23. P2 family: zinc finger proteins• Contain cys2/his2motif• Inhibition oftranscription at the endof spermiogenesis• Stabilization of spermchromatin by zinc• P2 family possiblyattached to majorgroove of DNA
  24. 24. Comparison P1 and P2• P1/P2 ratio= 0.98 0.12• P1 rich of cys• P2 rich of his and few cys compare to P1• P2 is more basic• P2 has more affinity to DNA
  25. 25. Mechanism of condensation• In bull and fish P1 attached to major grooveof DNA• Anchoring domains attached to major groove• After the synthesis of protamine, and before itsdeposition on DNA, the serine and threonineresidues in protamine are phosphorylated• After attachment, this residuesdephosphorylated• Formation of intra and inter protamine disulfidebond
  26. 26. Mechanism of condensation• In humans and other mammalian whichhave two or more protamine there arecontradictory evidence:• P1 & P2 lie down in major groove• P1 & P2 attached to both minor and majorgroove• P1 & P2 attached to external surface of theDNA chain
  27. 27. Evaluation of nuclear maturity• Chromatin decondensation induced in vitro• DNA staining after chromatin denaturation• Aniline blue staining• Assessment of thiol- disulfide status of nuclei• Ultra structural examination• Biochemical analysis• Comet assay• TUNEL assay• Sperm chromatin dispersion test (SCD test)

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