Fig 1. UNC-85 mutations restore the sterility of H2B 3KO C. elegans.

(A) C. elegans H2B genes and proteins. (B) Localization of the endogenously GFP::tagged HIS-58 on chromosomes during metaphase in wild-type (WT) one-cell embryos. Scale bar, 5 μm. (C) Loss of H2B decreased brood size. N = 20–53, data are presented as mean ± SD (error bars). Statistical significance compared with the control with a matching color code is based on Student’s t-test, n.s., not significant, **p< 0.01, ***p< 0.001. Three biological replicates calculated the fertile adult percentage. (D) Schematic of the C. elegans UNC-85 and ASFL-1 proteins. The amino acid changes (cas1067, cas1068, and cas1100) and the deletion allele of unc-85 (cas1571) or asfl-1 (cas1222) are indicated. The scale bar represents 20 amino acids. (E) Multi-sequence alignment of C. elegans (Ce) UNC-85 with ASFL-1 and its homologs in human (Hs), mouse (Mm), Drosophila (Dm), and Saccharomyces (Sc). The sequence alignment shows the protein sequence of UNC-85 histone chaperone domain flanking (± 20 aa) around the mutation isolated from the EMS screen, and the histone chaperone domain is highly acidic, which favors its interaction with histone. The red asterisk highlights the mutation isolated from the EMS screen. An asterisk (*) indicates positions 100% conserved in the alignment; a colon (:) indicates conservation between groups of strongly similar properties; a period (.) indicates conservation between groups of weakly similar properties. Hydrophobic residues are colored in blue, positive charge residues in yellow, negative charge residues in magenta, and polar residues in green. (F) Fertile adult percentage and brood size of the indicated strains. N = 10–70, values are presented as mean ± SD (error bars). Statistical significance compared with the control with a matching color code is based on Student’s t-test, n.s., not significant, *p< 0.05, **p< 0.01, ***p< 0.001. Three biological replicates calculated the fertile adult percentage.