Phosphorylation protects sperm-specific histones H1 and H2B from proteolysis after fertilization

At intermediate stages of male pronucleus formation, sperm-derived chromati n is composed of hybrid nucleoprotein particles formed by sperm H1 (SpH1), dimers of sperm H2A-H2B (SpH2A-SpH2B), and a subset of maternal cleavage st age (CS) histone variants. At this stage in vivo, the CS histone variants a re poly(ADP-ribosylated), while SpH2B and SpH1 are phosphorylated. We have postulated previously that the final steps of sperm chromatin remodeling in volve a cysteine-protease (SpH-protease) that degrades sperm histones in a specific manner, leaving the maternal CS histone variants unaffected. More recently we have reported that the protection of CS histones from degradati on is determined by the poly(ADP-ribose) moiety of these proteins. Because of the selectivity displayed by the SpH-protease; the coexistence of a subs et of SpH together with CS histone variants at intermediate stages of male pronucleus remodeling remains intriguing. Consequently, we have investigate d the phosphorylation state of SpH1 and SpH2B in relation to the possible p rotection of these proteins from proteolytic degradation. Histones H1 and H 2B were purified from sperm, phosphorylated in vitro using the recombinant a-subunit of casein kinase 2, and then used as substrates in the standard a ssay of the SpH-protease. The phosphorylated forms of SpH1 and SpH2B were f ound to remain unaltered, while the nonphosphorylated forms were degraded. On the basis of this result, we postulate a novel role for the phosphorylat ion of SpH1 and SpH2B that occurs in vivo after fertilization, namely to pr otect these histones against degradation at intermediate stages of male chr omatin remodeling. (C) 1999 Wiley-Liss, Inc.