N-Acetylglucosaminyl Transferase Regulates the Expression of the Sulfoglucuronyl Glycolipids in Specific Cell Types in Cerebellum during Development

Abstract

In the adult cerebellum, sulfoglucuronyl glycolipids (SGGLs) are specifically localized in Purkinje cells and their dendrites in the molecular layer. Other major cell types such as granule neurons and glial cells lack SGGLs. To explain the cell specific localization and the known biphasic expression of SGGLs, enzymic activities of four glycosyltransferases involved in the biosynthesis of SGGLs were studied in murine cerebellar mutants, in distinct cellular layers of rat cerebellum, and in isolated granule neurons during development. The enzymes studied were lactosylceramide: N-acetylglucosaminyl transferase (GlcNAc-Tr), lactotriaosylceramide:galactosyltransferase, neolactotetraosylceramide:glucuronyltransferase, and glucuronylglycolipid:sulfotransferase. In the cerebellum of Purkinje cell-deficient mutants, such as (pcd/pcd) and lurcher (Lc/؉) where Purkinje cells are lost, GlcNAc-Tr was absent, but the other three glycosyltransferase were not severely affected. This indicated that the latter three enzymes were localized in other cell types, such as in mature granule neurons and glial cells, in addition to that in Purkinje cells, and the lack of SGGLs in these mutants was due to absence of GlcNAc-Tr. Analyses of the enzymes in the specific micro-dissected cellular layers also showed that Purkinje cell layer and molecular layer (where Purkinje cell dendrites are localized) contained all four enzymes. However, granule neurons and glial cells in the white matter lacked GlcNAc-Tr, but expressed the other three enzymes. It was concluded that the absence of SGGLs in adult granule neurons and glial cells was due to specific deficiency of the GlcNAc-Tr. Although adult granule neurons lacked GlcNAc-Tr and therefore SGGLs, isolated granule neurons from the neonatal cerebellum contained all four enzymes necessary for the synthesis of SGGLs. With development, the activity of GlcNAc-Tr in the isolated granule neurons declined but the other enzymes were not as affected, indicating that immature granule neurons were capable of synthesizing SGGLs and with maturation the synthesis was down-regulated. This also explains the biphasic expression of SGGLs in the developing cerebellum.

Molecular Cloning and Characterization of UDP-GlcNAc:Lactosylceramide β1,3-N-Acetylglucosaminyltransferase (β3Gn-T5), an Essential Enzyme for the Expression of HNK-1 and Lewis X Epitopes on Glycolipids

“…4) The expression level of the ␤3Gn-T5 transcript in various cultured cancer cells was well correlated with the Lc 3 Cer synthesizing activity. 5) The changes in the ␤3Gn-T5 transcript level during HL-60 differentiation and during rat brain development were consistent with those of the Lc 3 Cer synthesizing activity as reported by others (15,20,35,46).…”

“…In particular, the expression level of two SGGLs, SGGL-1 and SGGL-2, carrying the HNK-1 epitope is determined by the Lc 3 Cer synthase (15, 19 -22, 46). The change in the level of ␤3Gn-T5 transcript in developing rat brain almost paralleled that in Lc 3 Cer synthesizing activity reported previously (15,20,46). This strongly indicated that ␤3Gn-T5 is the Lc 3 Cer synthase.…”

“…Lc 3 Cer synthase has been detected in many tissues, including the developing rat brain (14,15,20,46), hematopoietic cells (35,36,(47)(48)(49), and colorectal tissues (50 -52). In all tissues examined, the Lc 3 Cer synthase was identified to be a key enzyme in the extension of the carbohydrate chain on neolactoseries GSLs.…”

“…The marked reduction in the ␤3Gn-T5 activity for the longer polylactosamine chain would suggest that GSLs with a long polylactosamine chain, such as nLc 6 Cer, are not good substrates for ␤3Gn-T5. In previous studies (15,20,46), the activity of ␤3Gn-T toward LacCer and nLc 4 Cer was measured using tissue homogenates of rat brain during development. Both activities showed not only very similar profiles of change during the development, but almost the same level.…”

Combinatorial Ganglioside Biosynthesis

“…In adult cerebellum, it is restricted to Purkinje cells (8). The biological function of this molecular diversity is unclear, but the vital role of glycosphingolipids has been demonstrated by the fact that their complete deficiency in mice leads to early embryonic lethality (see below).…”