Novel EukaryoticDouble Histone Fold Motifs SharingHigh Sequence Identity with Histones H3 and H4 in Single-Domain Proteinsand in Proteins Featuring Remarkable Histone Fold Multiplets

Histone proteins play a central role in chromatin organization. In eukaryotes, the fundamental units of DNA packaging─the nucleosomal cores─are assembled from histone dimers. The double histone fold (DHF) refers to a protein architecture in which two adjacent regions, each containing a histone fold, associate to form a histone pseudodimer. In the present study, by targeted sequence searches in protein databases and subsequent structural and phylogenetic investigations, we identified a large number of DHF proteins featuring a high or very high degree of identity with the amino acid sequences of both histones H3 and H4, which constitute a new class of eukaryotic DHF proteins. Strikingly─somehow in analogy with recently identified proteins encoded in some giant viruses─we found, as well, triplets of various kinds (i.e., proteins showing regions of homology with histones H3, histone H4, and an additional histone fold). We were also able to evidence the existence of unprecedented quadruplets encompassing two distinct DHF domains, as well as multiplets that include not only region of homology to nucleosome core histones, but also to the linker histone H1. Focus was put on the evolutionary scenarios for the origin of the newly identified proteins, as well as on the conservation of residues relevant for dimerization and DNA binding. Implications of our findings in fundamental areas of biochemistry are illustrated, and perspectives for future research directions are discussed.