Nucleosome formation with the testis-specific histone H3 variant, H3t, by human nucleosome assembly proteins in vitro.
Five non-allelic histone H3 variants, H3.1, H3.2, H3.3, H3t and CENP-A, have been identified in mammals. H3t is robustly expressed in the testis, and thus was assigned as the testis-specific H3 variant. However, recent proteomics and tissue-specific RT-PCR experiments revealed a small amount of H3t expression in somatic cells. In the present study, we purified human H3t as a recombinant protein, and showed that H3t/H4 forms nucleosomes with H2A/H2B by the salt-dialysis method, like the conventional H3.1/H4. We found that H3t/H4 is not efficiently incorporated into the nucleosome by human Nap1 (hNap1), due to its defective H3t/H4 deposition on DNA. In contrast, human Nap2 (hNap2), a paralog of hNap1, promotes nucleosome assembly with H3t/H4. Mutational analyses revealed that the Ala111 residue, which is conserved among H3.1, H3.2 and H3.3, but not in H3t, is the essential residue for the hNap1-mediated nucleosome assembly. These results suggest that H3t may be incorporated into chromatin by a specific chaperone-mediated pathway.
Tachiwana
Hiroaki
H
Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, 3-4-1 Okubo, Shinjuku-ku, Tokyo 169-8555, Japan.
Osakabe
Akihisa
A
Kimura
Hiroshi
H
Kurumizaka
Hitoshi
H
D018797:Cell Cycle Proteins / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D004252:DNA Mutational Analysis; D014644:Genetic Variation; D006657:Histones / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008297:Male; D009687:Nuclear Proteins / Q000378:metabolism*; D056490:Nucleosome Assembly Protein 1; D009707:Nucleosomes / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D013737:Testis / Q000378:metabolism
D018797:Cell Cycle Proteins; D006657:Histones; C533557:NAP1L1 protein, human; C104356:NAP1L4 protein, human; D009687:Nuclear Proteins; D056490:Nucleosome Assembly Protein 1; D009707:Nucleosomes; D020033:Protein Isoforms; D004247:DNA