Human sperm tall fibrous sheath undergoes phosphorylation during its development

Abstract

The phosphorylation of the human sperm tail fibrous sheath as a maturational step during its development is reported for the first time. This was demonstrated using GDA-J/F3 and RT97 monoclonal antibodies (MoAb) which recognize the fibrous sheath. In indirect immunofluorescence microscopy of frozen sections of human adult testes, the two antibodies reacted with the assembled fibrous sheath only, but the numbers of sperm tails stained with RT97 were consistently lower than those treated with GDA-J/F3. Furthermore, by using double indirect immunofluorescence, although the majority of spermatozoa were doubly stained with the two MoAbs, some GDA-J/F3-positive sperm tails were negative with RT97. In epididymal and ejaculated spermatozoa, the two antibodies stained all the tails. This indicated that the ontogenic appearance of the GDA-J/F3 epitope precedes that of RT97. In Western blotting and/or indirect immunofluorescence of spermatozoa, treatment of samples with alkaline phosphatase abolished the reactivity of RT97 while that of GDA-J/F3 MoAb was not affected. This finding indicated that the RT97 but not the GDA-J/F3 epitope was phosphorylated. Together, these results therefore reveal that during tail morphogenesis, the fibrous sheath undergoes phosphorylation as part of its structural maturation. Screening of sperm cell precursors recovered from oligozoospermic donors showed reaction of some abnormal germ cells with GDA-J/F3 MoAb but not with RT97, suggesting the possible failure of phosphorylation of the fibrous sheath protein in these cells. The significance of these findings is discussed together with the biological importance of phosphorylation to the fibrous sheath.