Xenopus Nucleoplasmin (Np) and N1 are equally abundant chaperones found in oocytes and eggs. Np participates in nucleosome assembly and exchange reactions involving histone H2A/H2B dimers and DNA; whilst N1 functions as a chaperone specific for the H3/H4 tetramer. Therefore, N1 and Np may function together to promote nucleosome assembly in oocytes. DNA replication and transcription are sensitive to the local packing of nucleosomes, which may be modulated by histone acetylation/deacetylation, phosphorylation, DNA methylation, or a combination thereof. Hence, nucleosome assembly and exchange factors may play a critical role in these cellular processes. We propose to study the roles of Np and N1 in nucleosome assembly and in the decompaction of sperm chromatin after fertilization, by obtaining the first high resolution crystal structures of these chaperones, alone and complexed with their cognate histories. This will reveal the molecular architecture and function of the Np and N1-families, while providing insights into the extreme thermal and chemical stability of the Np pentamer. It was suggested previously that N1 and Np function sequentially to deliver core histones to DNA during nucleosome assembly. We have demonstrated that Np forms a decamer in solution which can be reconstituted with separately purified histories H2A/H2B and H3/H4, to form stable Np decamer-octamer complexes in the absence of DNA. Hence, the Np decamer with 522 point group symmetry may provide a docking ring for 10 H2A/H2B dimers and 5 H3/H4 tetramers, to form an Np decamer-octamer complex which utilizes their common 2-fold symmetry. We further suggest that N1 may establish an N1 dimeric cap on H3/H4 tetramers in cells, which then docks with Np decamer-dimer complexes to form a larger Histone Octamer co-Chaperone (HOC) complex. Overall, our data suggest a new paradigm for nucleosome assembly, in which pre-formed histone octamer-chaperone complexes and DNA are brought together in a concerted mechanism. To understand this process, the structures of the Np decamer-octamer and putative HOC complexes will be determined by electron cryo-microscopy and X-ray crystallography, in combination with biochemical studies of nucleosome assembly.
| Platonova, Olga; Akey, Ildikó V; Head, James F et al. (2011) Crystal structure and function of human nucleoplasmin (npm2): a histone chaperone in oocytes and embryos. Biochemistry 50:8078-89 |
