Previously, it was found that a novel class of neutral fucosylated glycosphingolipids (GSLs) is required for male fertility. These lipids contain very long-chain (C26 -C32) polyunsaturated (4 -6 double bonds) fatty acid residues (VLC-PUFAs). To assess the role of these complex GSLs in spermatogenesis, we have now investigated with which of the testicular cell types these lipids are associated. During postnatal development, complex glycosylated and simple VLC-PUFA sphingolipids were first detectable at day 15, when the most advanced germ cells are pachytene spermatocytes. Their synthesis is most likely driven by ceramide synthase-3. This enzyme is encoded by the Cers3/ Lass3 gene (longevity assurance genes), and out of six members of this gene family, only Cers3 mRNA expression was limited to germ cells, where it was up-regulated more than 700-fold during postnatal testicular maturation. Increasing levels of neutral complex VLC-PUFA GSLs also correlated with the progression of spermatogenesis in a series of male sterile mutants with arrests at different stages of spermatogenesis. Remarkably, fucosylation of the complex VLC-PUFA GSLs was not essential for spermatogenesis, as fucosylation-deficient mice produced nonfucosylated versions of the complex testicular VLC-PUFA GSLs, had complete spermatogenesis, and were fertile. Nevertheless, sterile Galgt1 ؊/؊ mice, with a defective meiotic cytokinesis and a subsequent block in spermiogenesis, lacked complex but contained simple VLC-PUFA GSLs, as well as VLC-PUFA ceramides and sphingomyelins, indicating that the latter lipids are not sufficient for completion of spermatogenesis. Thus, our data imply that both glycans and the particular acyl chains of germinal sphingolipids are relevant for proper completion of meiosis.The testis is composed of two functional compartments as follows: (i) the seminiferous tubules, containing developing germ cells and supporting Sertoli cells, and (ii) the steroidogenic Leydig cells in the interstitium (Fig. 1A) (1, 2). In mature testis, the seminiferous tubules are separated by a blood-testis barrier (BTB) 5 into a basal and an adluminal compartment. Tight junctions contribute to the establishment of the BTB, which is made up of adjacent Sertoli cells and physically segregates post-meiotic germ cells from nutrients and biomolecules in the systemic circulation (3, 4).The development of the male germ cells, taking place within the seminiferous tubules, is a complex and highly regulated process (2). During spermatogenesis, testicular stem cells (undifferentiated spermatogonia) give rise to a lineage of cells that multiply by mitosis (proliferative spermatogonia). These cells differentiate to go through the meiotic division (spermatocytes) and become haploid germ cells (spermatids), which transform into spermatozoa. It is during the meiotic prophase that leptotene spermatocytes transit the BTB. The post-meiotic development (spermiogenesis) involves a dramatic change of nuclear shape, chromatin condensation, the loss of most cell organelles, a...

Modification of glycoproteins by the attachment of fucose residues is widely distributed in nature. The importance of fucosylation has recently been underlined by identification of the monogenetic inherited human disease "congenital disorder of glycosylation IIc," also termed "leukocyte adhesion deficiency II." Due to defective Golgi GDP-fucose transporter (SLC35C1) activity, patients show a hypofucosylation of glycoproteins and present clinically with mental and growth retardation, persistent leukocytosis, and severe infections. To investigate effects induced by the loss of fucosylated structures in different organs, we generated a mouse model for the disease by inactivating the Golgi GDP-transporter gene (Slc35c1). Lectin binding studies revealed a tremendous reduction of fucosylated glycoconjugates in tissues and isolated cells from Slc35c1 ؊/؊ mice. Fucose treatment of cells from different organs led to partial normalization of the fucosylation state of glycoproteins, thereby indicating an alternative GDP-fucose transport mechanism. Slc35c1-deficient mice presented with severe growth retardation, elevated postnatal mortality rate, dilatation of lung alveoles, and hypocellular lymph nodes. In vitro and in vivo leukocyte adhesion and rolling assays revealed a severe impairment of P-, E-, and L-selectin ligand function. The diversity of these phenotypic aspects demonstrates the broad general impact of fucosylation in the mammalian organism.The covalent attachment of oligosaccharide moieties to newly synthesized proteins comprises one of the most frequent but also complex forms of co-and posttranslational protein modifications, which has been found in nearly all living organisms (1). Glycan structures affect the physicochemical properties and the function of proteins in a variety of biological processes, including folding, solubility, sorting, proteolytic stability, and receptor-ligand interactions. In mammalian organisms, the biosynthesis of the oligosaccharide chains requires a broad spectrum of glycosyltransferases, glycosidases, transport proteins, and 13 different monosaccharides (2-4).Due to its variability of binding types (␣-1,2-, ␣-1,3-, ␣-1,4-, and ␣-1,6-fucosylation have been described), the monosaccharide fucose plays an important role in the microheterogeneity of oligosaccharide structures (5). Fucose residues are predominantly linked to peripheral parts of N-, O-, and lipid-linked oligosaccharides, thereby building cap structures, which have been observed in many surface-localized and secreted proteins,

Leukocyte adhesion deficiency II (LAD II), also known as congenital disorder of glycosylation IIc (CDG-IIc), is a human disease in which a defective GDP-fucose transporter (SLC35C1) causes developmental defects and an immunodeficiency that is based on the lack of fucosylated selectin ligands. Since the study of in vivo leukocyte trafficking in patients with LAD II is experimentally limited, we analyzed this process in mice deficient for Slc35c1. We found that E-, Land nd P-selectin-dependent leukocyte rolling in cremaster muscle venules was virtually absent. This was accompanied by a strong but not complete decrease in firm leuko-cyte adhesion. Moreover, neutrophil migration to the inflamed peritoneum was strongly reduced by 89%. Previous reports showed surprisingly normal lym-phocyte functions in LAD II, which indicated sufficient lymphocyte trafficking to secondary lymphoid organs. We now found that while lymphocyte homing to lymph nodes was reduced to 1% to 2% in Slc35c1 / mice, trafficking to the spleen was completely normal. In accordance with this, we found a defect in the hu-moral response to a T cell-dependent an-tigen in lymph nodes but not in the spleen. Taken together, Slc35c1 / mice show strongly defective leukocyte trafficking but normal lymphocyte homing to the spleen, which may explain normal lym-phocyte functions in LAD II. (Blood. 2008;