Involvement of protein kinase A in the phosphorylation of spermatidal protein TP2 and its effect on DNA condensation.
Rat spermatidal protein TP2 is rich in serine residues and has several potential sites for phosphorylation by different protein kinases. Recombinant TP2 is phosphorylated upon incubation in vitro with salt extract of testicular sonication resistant nuclei (SRN) (representing elongating and elongated spermatids). The major phosphorylation sites were localized to the C-terminal, V8 protease-derived, fragment (residues 87-114). Phosphorylation experiments with the wild type and different site-specific mutants of TP2 revealed that serine 109 and threonine 101 are the phosphorylation sites. Phosphorylation of the C-terminal fragment of TP2 was also demonstrated in vivo. Phosphorylation was not stimulated by either protein kinase C activators or cGMP but was inhibited by protein kinase A inhibitor (PKI) peptide, showing the involvement of protein kinase A in the phosphorylation of TP2. Phosphorylation of TP2 greatly reduced its DNA condensation property. TP2 when complexed with DNA was not a good substrate for phosphorylation by PKA. Dephosphorylation of the DNA-TP2 complex by calf intestinal alkaline phosphatase restored the DNA condensation property to a level equivalent to that observed with TP2. The physiological significance of the phosphorylation-dephosphorylation cycle is discussed with reference to the two-domain model of TP2.
Meetei
Amom Ruhikanta
AR
Department of Biochemistry, Indian Institute of Science, Bangalore 560 012, India.
Ullas
Kolathur S
KS
Vasupradha
V
V
Rao
Manchanahalli R Satyanarayana
MR
D000595:Amino Acid Sequence; D000818:Animals; D002467:Cell Nucleus; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism* / Q000494:pharmacology; D002942:Circular Dichroism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D010450:Endopeptidases / Q000378:metabolism; D007700:Kinetics; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D051381:Rats; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013087:Spermatids / Q000201:enzymology*; D013737:Testis / Q000378:metabolism
D002868:Chromosomal Proteins, Non-Histone; D011994:Recombinant Proteins; C053886:spermatid transition proteins; D004247:DNA; D017868:Cyclic AMP-Dependent Protein Kinases; D010450:Endopeptidases