Biological Functions of Membrane Glycolipids

(Update Issue: March.19, 2007) 
 Glycolipids are present ubiquitously in cell surface membranes. The most common glycolipids in mammals are a class of sphingolipids, glycosphingolipids (GSLs). Major GSLs including gangliosides are derived from glucosylceramide (GlcCer) that is synthesized by a specific glycosyltransferase, ceramide glucosyltransferase (GlcT-1, UGCG). Galactosylceramide (GalCer) formed by ceramide galactosyltransferase (CGT) is also present as a dominant glycolipid in myelin sheath. The class of glyceroglycolipids is found in particular tissues: galactosylalkylacylglyceride (seminolipid) in the testis, and phosphatidyglucose in human cord blood cells. Glucosylated cholesterol is found in mammalian cells (Table 1).
Table I Roles of monoglycosylated lipids in mammals: almost all lipids are glycosylated.
Glycolipid Distribution Synthetic enzyme Role
GlcCer ubiquitous UGCG (human, mouse, rat) precursor for GSLs biosynthesis, negative regulator for ceramide, axonal elongation of neuron, multidrug resistance
GalCer myelin, kidney, CGT (human, mouse, rat) insulative function of myelin
ÉøGalCer unknown unknown ligand for NKT cell
FucCer colon cancer, lung cancer unknown unknown
Cholesteryl glucoside human skin fibroblast unknown heat shock response
Galactosyldiacyl glycerol sperm (seminolipid), intestine, the nervous system CGT (the same as GalCer synthase) sperm development
phosphatidylglucoside cord blood cell, HL60 unknown unknown
NKT: natural Killer T
Phosphatidylinositol is not included.

Table II Gene targeted mice: glycolipid functions at the whole animal level
Gene disrupted Biochemistry General
GlcCer synthase (UGCG) loss of all GSLs except GalCer embryonic lethality at E7.5, enhancemant of apoptosis in ectodermal layer
GalCer synthase (CGT) loss of GalCer and appearance of GlcCer in myelin, loss of seminolipid in sperm ataxia, loss of insulative function of myelin, sterility in male
Sulfatide synthase (CST) loss of sulfatide in myelin and of sulfated seminolipid in sperm similarity to CGT KO mouse
GM2/GD2 (GalNAc-T) synthase loss of higher gangliosides (only GM3, GD3 and GT3) normal development, defect in nerve regeneration
GD3 synthase loss of higher gangliosides (only monosialo gangliosides) normal development defect in nerve regeneration
GM2/GD3 double KO loss of gangliosides except GM3 normal development, lethality to sound stress
UDCG:UDP-Glc Ceramide Glucosyltransferase
CGT: Ceramide Galactosyltransferase
CST: Cerebroside Sulfotransferase
Recent cell biological studies show that GSLs in cell membranes are preferentially distributed into lipid domains, so-called rafts (Fig. 1, see also “Glyco Word” by Higashi). Raft-like lipid microdomains can be isolated biochemically as a low density Triton X-100 insoluble fraction. The fraction isolated contains sphingolipids (GSLs and sphingomyelin) and cholesterol. Importantly, the glycolipid-enriched membrane domain also includes proteins related to signal transduction such as src-family kinases. The lipid domains are suggested to play roles in cell-cell adhesion and receptor-mediated signal transduction. A glycolipid-enriched lipid micodomain is also involved in target for host pathogens (Vibrio cholera, O157, HIY) and their toxin bindings (Fig. 1).

A cell line deficient in an entire group of GSLs was established from B16 mouse melanoma. This mutant cell line is defective in a GlcCer synthase activity. In vitro studies on glycolipid function using the mutant cells show that the glycolipid biosynthesis is not essential for cell survival and proliferation, at least in the melanoma cells. Sphingomyelin as the sole sphingolipid on the plasma membrane can act as a substitute. Similarly, GSLs are not a critical component for membrane domain formation. These results from in vitro experiments indicate that GSLs apparently do not have house-keeping functions when studied at the single cell level. Thus, GSL research is now focusing on multicellular systems or whole animals. Up to now, however, there is no case report of human genetic disease associated with glycolipid synthetic enzyme. Therefore, the physiological functions of GSLs remain unclear in humans.

To solve the riddle of glycolipid functions, a method to eliminate a gene of glycolipid glycosyltransferase in mice has been developed using a homologous recombination system and several knockout mice have been generated (Table II). Surprisingly, mutant mice deficient in complex gangliosides are born normally, showing no apparent abnormalities. Even a double knockout mouse of GD3 synthase and GM2/GD2 synthase, expressing only GM3 ganglioside, results in normal development. Studies with these mutant mice prove that complex gangliosides are not involved in cellular differentiation but rather in the maintenance (homeostasis) and regeneration of nervous system tissues (see “Glyco Word” by Furukawa). Very recently, Kawai et al. have shown that the double mutant is highly sensitive to sound stimulus and displays a sudden death phenotype

In 1999, a mutant mouse for GlcCer synthase (UGCG) was generated. Since the mutant mouse is deficient in all GSLs except for GalCer, it gives us valuable information on the general functions of GSLs. The mice die at around embryonic day 7.5, indicating clear evidence of the essential role of GSL biosynthesis during multicellular development. The mutant mice exhibit an elevated apoptosis in the ectoderm. However, it is difficult to explain clearly why ablation of the UGCG gene causes the enhancement of apoptotic cell death because UGCG has at least two distinct functions one of which is a negative regulatory factor for intracellular ceramide content through ceramide glycosylation. The other is a positive regulatory role of the glycolipid, GlcCer in cell growth. Moreover, it is not evident if there is any relationship between the embryonic lethality and glycolipid functions in raft-like membrane microdomains.

GalCer knockout mouse was first generated in 1996. In contrast to the UGCG mutant, the CGT knockout mouse is born normally and exhibits no obvious abnormalities in myelin structure, although GalCer and sulfatide, both enriched in myelin, are lacking. However, the mouse shows severe generalized tremor and mild ataxia, proving that the GSLs play a role in the insulative function of myelin. The same mouse shows that CGT is also essential for sperm development.

As shown above, glycolipid research is now regarded as a potent and promising research target of the post-genomic era.
Yoshio Hirabayashi and Soh Osuga (Brain Science Institute, The Institute of Physical and Chemical Research (RIKEN))
References (1) Hirabayashi Y, Ichikawa S,: Roles of glycolipids and sphingolipids in biological membrane: The Frontiers in Molecular Biology Series (Eds., Fukuda, M., Hindsgaul, O, IRL Press at Oxford Press) (1999) pp220-248
(2) Yamashita T, Wada R, Sasaki T, Deng C, Bierfreund U, Sandhoff K, Proia RL Proc Natl Acad Sci U S A96, 9142-9147, 1999
(3) Kobayashi T, Hirabayashi Y, Glycoconjugates J. 17, 163-171, 2000
(4) Kawai H, Allende M, Wada R, Kono, M, Sango K, Deng C, Miyakawa T, Crawley, J, Werth N, Bierfreund U, Sandhoff K, Proia R, J. Biol. Chem. 276, 6885-6888, 2001
Jun. 15, 2002

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