Characterization of cAMP‐dependent protein kinase and its endogenous substrate proteins in ram testicular, cauda epididymal, and ejaculated spermatozoa

Abstract

Mammalian spermatozoa have been shown to possess cAMP-dependent protein kinase (A-PK) and endogenous substrate proteins for this enzyme. A study of the kinase system was undertaken to determine changes that may be associated with sperm maturation by comparing immature testicular with mature cauda epididymal and ejaculated spermatozoa. Absolute activity levels of A-PK, stimulated over a concentration range of 10(-9) to 10(-5) M, was significantly greater in testicular than ejaculated spermatozoa. At an optimal cAMP concentration (10(-6) M), testicular spermatozoa had significantly greater amounts of cAMP-dependent protein kinase activity than did cauda or ejaculated spermatozoa. Electrophoretic analysis and autoradiography of NP-40-soluble protein extracts revealed the presence of two substrate proteins (Mr = 62,000 and 44,000) in all three types of spermatozoa. In addition, a phosphoprotein (Mr = 20,000) was detected in mature cauda and ejaculated but not immature testicular spermatozoa. The phosphorylation of these substrate proteins was both dose and time dependent. Examination of cyclic AMP phosphodiesterase activity revealed significantly higher levels in testicular than ejaculated spermatozoa. These results indicate marked alterations in cAMP-modulated protein phosphorylation and dephosphorylation systems in ram spermatozoa during epididymal maturation.

Differential localization of two isoforms of the regulatory subunit RIIα of cAMP‐dependent protein kinase in human sperm: Biochemical and cytochemical study

“…been performed intensively. CAMP-dependent protein kinase activities, CAMP-dependent protein kinases and phosphoproteins have been found in detergent extracts and sonicates of mammalian sperm (Hoskins et al, 1972;Garbers et al, 1973;Brandt and Hoskins, 1980;Tash et al, 1984;Wooten et al, 1987;Noland et al, 1987). The characterization and localization of CAMPdependent protein kinases in rat and bovine sperm have been performed by Horowitz et al (1984Horowitz et al ( , 1988Horowitz et al ( , 1989 and Paupard et al (1988).…”

Evidence for presence of hyaluronan binding protein on spermatozoa and its possible involvement in sperm function

“…The studies on involvement of the phosphorylated form of hyaluronan binding protein on sperm function is of further interest since we have recently reported the in vivo phosphorylation of this protein at tyrosine residues and, at the same time, the phosphorylation of spermatozoa proteins during essential sperm functions are well established (Brandt and Hoskins, 1980;Hayashi et al, 1987;Wooten et al, 1987;Komatsu, 1990;Majumdar et al, 1990).…”

Influence of epididymal maturation on cyclic AMP levels in hamster spermatozoa

“…As the only known role for cAMP in eukaryotic cells is the stimulation of protein phosphorylation via activation of protein kinase A (Rosen, Erlichman & Rubin, 1975), changes in the activity of this enzyme could thus affect the acquisition of motility in the epididymis. Although protein kinase activity increases during epididymal transit in the bull (Hoskins et af., 1974), levels of this enzyme have been reported both to rise (Pariset etal., 1985) and to fall (Wooten, Voglmayr & Wrenn, 1987) during epididymal maturation in the ram, and to remain unchanged in the rat (Atherton et al, 1985). However, two phosphorylated proteins of 20 and 16 kD were found exclusively in ram sperm from the cauda epididymis (Wooten et af., 1987).…”