Requirement of Seminolipid in Spermatogenesis Revealed by UDP-galactose:Ceramide Galactosyltransferase-deficient Mice

Abstract

Although seminolipid has long been suspected to play an essential role in spermatogenesis because of its uniquely abundant and temporally regulated expression in the spermatocytes, direct experimental evidence has been lacking. We have tested the hypothesis by examining the testis of the UDP-galactose:ceramide galactosyltransferase-deficient mouse, which is incapable of synthesizing seminolipid. Spermatogenesis in homozygous affected males is arrested at the late pachytene stage and the spermatogenic cells degenerate through the apoptotic process. This stage closely follows the phase of rapid seminolipid synthesis in the wild-type mouse. These observations not only provide the first experimental evidence that seminolipid is indeed essential for normal spermatogenesis but also support the broader concept that cell surface glycolipids are important in cellular differentiation and cell-to-cell interaction.Seminolipid (3-sulfogalactosyl-1-alkyl-2-acyl-sn-glycerol) is the principal glycolipid in spermatozoa of mammals comprising, for example, approximately 3% of total lipids and more than 90% of total glycolipids in boar spermatozoa (1-3). During spermatogenesis, seminolipid is synthesized rapidly in the early phase of spermatocyte development and maintained in subsequent germ cell stages (4 -6). This developmentally regulated rapid synthesis suggested a specific and possibly essential function of seminolipid in spermatogenesis (7) but experimental evidence has been lacking. Firm evidence in support of the speculation would have important bearing to the general concept that cell surface glycoconjugates are important in cellular differentiation, and cell-to-cell interaction (8).Seminolipid is synthesized by sulfation of its precursor, galactosylalkylacylglycerol (GalEAG) 1 . GalEAG is synthesized by UDPgalactose:ceramide galactosyltransferase (CGT, EC 2.4.1.62), which, besides GalEAG, also synthesizes the major myelin galactolipid, galactosylceramide (GalCer), galactosylsphingosine (psychosine), and galactosyldiacylglycerol (GalAAG) (9, 10). The CGTdeficient mice recently generated by gene-targeting do not synthesize any of these products and subsequent derivatives of the products (11)(12)(13)(14). Thus, the CGT-deficient mouse is an ideal experimental model to examine the consequences of lack of seminolipid to spermatogenesis. This report describes the first definitive evidence that deficient seminolipid biosynthesis indeed causes devastating disruption of the normal spermatogenetic process. EXPERIMENTAL PROCEDURESMice-The mice heterozygous for the disrupted Cgt gene (11) were originally supplied by Dr. B. Popko and maintained by backcrossing to C57BL/6N. Genotype was determined according to Coetzee et al. (11). WBB6F1 Kit W/W-v and WBB6F1 Mg f Sl/Sl-d mutant mice were purchased from Japan SLC, Inc., and C57BL/6N inbred mice were purchased from CLEA Japan, Inc. Isolation of Testicular Germ Cells-Testicular germ cells were isolated from decapsulated testes of sexually mature male C57BL/6N mice (15).RT-PCR Analysis-RNA...

Paranodal junction formation and spermatogenesis require sulfoglycolipids

“…CGT-knockout mice are completely devoid of GalCer and sulfatide in the brain (14,15) and of GalEAG and seminolipid in the testis (16). CGT-deficient mice manifest neurological disorders caused by myelin dysfunction (14,15) and the complete inhibition of spermatogenesis at the late-pachytene spermatocyte stage (16). These results provide strong support for the view that the intermediate products, GalCer and GalEAG, and͞or the end products, sulfatide and seminolipid, are indispensable for myelin function and spermatogenesis.…”

“…This single enzyme, CGT, is responsible for the biosynthesis of GalCer and GalEAG, which are the precursors of sulfatide and seminolipid, respectively (14)(15)(16). CGT-knockout mice are completely devoid of GalCer and sulfatide in the brain (14,15) and of GalEAG and seminolipid in the testis (16).…”

“…This single enzyme, CGT, is responsible for the biosynthesis of GalCer and GalEAG, which are the precursors of sulfatide and seminolipid, respectively (14)(15)(16). CGT-knockout mice are completely devoid of GalCer and sulfatide in the brain (14,15) and of GalEAG and seminolipid in the testis (16). CGT-deficient mice manifest neurological disorders caused by myelin dysfunction (14,15) and the complete inhibition of spermatogenesis at the late-pachytene spermatocyte stage (16).…”

“…These results provide strong support for the view that the intermediate products, GalCer and GalEAG, and͞or the end products, sulfatide and seminolipid, are indispensable for myelin function and spermatogenesis. However, the creation of sulfotransferase-deficient mice would be highly desirable to understand completely the functions of the precursor galactolipids and their sulfated products (14)(15)(16).…”

Sphingolipids and membrane biology as determined from genetic models

“…In ceramide galactosyl transferase mutants, spermatogenesis in the homozygous males is arrested at the late pachytene stage, and the spermatogenic cells degenerate through apoptosis [136].…”