Phosphorylation at clustered -Ser-Pro-X-Lys/Arg- motifs in sperm-specific histones H1 and H2B.

Abstract

Sea urchin sperm‐specific histones H1 and H2B have distinctive N‐terminal, and in the case of H1 also C‐terminal, domains containing repeats of a basic motif (‐Ser‐Pro‐Lys/Arg‐Lys/Arg‐ or a closely related sequence). The histones in spermatids (the precursors of sperm) are phosphorylated, and the unphosphorylated histones of mature sperm are rephosphorylated upon fertilization. These changes correlate with finely tuned changes in chromatin packing in the nucleus, and the domains responsible are evidently the N‐terminal domains. We show that in spermatids there are six tandemly repeated phosphorylation sites in the N‐terminal domain of H1 (a typical cAMP dependent protein kinase site is not phosphorylated) and that H2B is phosphorylated in the N‐terminal domain at two or three sites in the case of H2B1 and four sites in H2B2. The consensus sequence for phosphorylation is ‐Ser‐Pro‐X‐Lys/Arg‐, where X is Thr, Gln, Lys or Arg. There is an additional phosphorylated site in the C‐terminal domain of H1 but most (or possibly all) copies of the consensus motif, which are here dispersed, are not phosphorylated. The negative charge introduced upon phosphorylation would be expected to weaken or abolish electrostatic interaction with DNA of this motif, which also occurs, and is phosphorylated, in somatic H1s.

DNA‐binding sperm proteins with oligo‐arginine clusters function as potent activators for egg CK‐II

“…The crude histone preparation from spermatozoa of the sea urchin, Hemicentrotus pulcherrimus, was obtained by a modification of the method originally described by Hill et al [13]. Briefly, the spermatozoa (about 30 g) were homogenized, using a Potter type homogenizer, in 15 mM Tris-HC1 buffer (pH 7.4) containing 60 mM KCI, 15 mM NaCI, 0.15 mM spermine, 0.5 mM spermidine, 0.2 mM PMSE, 15 mM 2-mercaptoethanol and 0.34 M sucrose.…”

A 180-kDa Protein Kinase Seems to Be Responsible for the Phosphorylation of Prothymosin α Observed in Proliferating Cells

“…Although no biological significance can at present be inferred from this result, it should be noted that these proteins are not substrates of CK-2 (29) and that the pattern of phosphorylation of ProT␣ by the ProT␣K does not bear any resemblance to that reported for kinases which phosphorylate H2B (40,41). In addition, the kinases that phosphorylate this histone are cAMP or cGMPdependent (40); by contrast, signal transduction experiments performed in our laboratory 2 indicate that ProT␣ is not directly phosphorylated by protein kinase A or C, and its phosphorylation in proliferating splenic lymphocytes seems to be dependent on protein kinase C activity. The present data thus suggest that the purified 180-kDa ProT␣K does not fall into any of the protein kinase categories described to date (41).…”

Fibroblasts transformed by combinations of ras, myc and mutant p53 exhibit increased phosphorylation of histone H1 that is independent of metastatic potential

“…Phosphorylation of H 1 alters its interaction with chromatin and may be important in the transcriptional state of genes to which it is bound [15,16]. We observed in cells transformed by combinations of ras, myc and mutant p53 that there is an increased amount of the phosphorylated H1 subtypes c-pHlb, pHlb and c-pHlc.…”

“…H1 subtypes differing in primary structure include Hla, Hlb, Hlc, Hld, Hle, Hit and H1 ° [12][13][14]. Posttranslational modifcation adds to heterogeneity in the H 1 s. Phosphorylation of H1 on serine and threonine in their amino and carboxyl terminal tails occurs in vivo and alters their interaction with DNA [11,15,16]. H 1 phosphorylation destabilizes higher order chromatin structure which is thought to allow accessory factors to participate in replication, mitotic condensation, and gene activation [16].…”