Fig 5 - uploaded by Makoto Ito
Content may be subject to copyright.
Abnormal membrane formation in cgt-3 RNAi-treated OD70 worms. Membranes can be visualized by strong fl uorescence of an mCherry-tagged PH domain. WT N2 worms RNAi-treated for cgt-3 ( A ), cgt-1 / cgt-2 ( D ), cgt-2 / cgt-3 ( G ) and bre-3 ( J ) are shown. Multinucleated embryos and dead eggs (arrows) are found in the uterus. In OD70 RNAi-treated worms, abnormal eggs with totally disrupted membranes (arrows) are among apparently normal eggs (see the DIC images in the second column and the accompanying epi fl uorescence images in the third column). Asterisk indicates the vulva. Scale bar, 100 μ m.
Source publication
Ceramide glucosyltransferase (Ugcg) [uridine diphosphate (UDP)-glucose:N-acylsphingosine D-glucosyltransferase or UDP-glucose ceramide glucosyltransferase (GlcT): EC 2.4.1.80] catalyzes formation of glucosylceramide (GlcCer) from ceramide and UDP-glucose. There is only one Ugcg gene in the mouse genome, which is essential in embryogenesis and brain...
Contexts in source publication
Context 1
... these affected eggs, no normal cell compartment in early division was formed. In cgt-3 RNAi-treated WT N2 worms, 8% of the worms showed membrane abnormality (n = 80), and in cgt-3 RNAi-treated OD70 worms, 12% of the worms showed this abnormality (n = 32) ( Figure 5A-C: cgt-3 RNAi). Multinucleated cells observed in this study looked similar to the defects produced when chondroitin proteoglycan synthesis is inhibited ( Hwang et al. 2003;Mizuguchi et al. 2003;Izumikawa et al. 2004). ...
Context 2
... cells observed in this study looked similar to the defects produced when chondroitin proteoglycan synthesis is inhibited ( Hwang et al. 2003;Mizuguchi et al. 2003;Izumikawa et al. 2004). The number of abnormal eggs increased to 14% when cgt-2 RNAi was combined with cgt-1 RNAi in N2 worms ( Figure 5D, n = 28) and 10% of cgt-1/cgt-2 double RNAi-treated OD70 worms showed the same defects ( Figure 5E and F, n = 42). When cgt-2/cgt-3 double RNAi Glucosylceramide synthesis in germline and embryos was performed, the abnormality was found in 42% of the N2 worms ( Figure 5G, n = 14) and in 40% of the OD70 worms ( Figure 5H and I, n = 20). ...
Context 3
... cells observed in this study looked similar to the defects produced when chondroitin proteoglycan synthesis is inhibited ( Hwang et al. 2003;Mizuguchi et al. 2003;Izumikawa et al. 2004). The number of abnormal eggs increased to 14% when cgt-2 RNAi was combined with cgt-1 RNAi in N2 worms ( Figure 5D, n = 28) and 10% of cgt-1/cgt-2 double RNAi-treated OD70 worms showed the same defects ( Figure 5E and F, n = 42). When cgt-2/cgt-3 double RNAi Glucosylceramide synthesis in germline and embryos was performed, the abnormality was found in 42% of the N2 worms ( Figure 5G, n = 14) and in 40% of the OD70 worms ( Figure 5H and I, n = 20). ...
Context 4
... number of abnormal eggs increased to 14% when cgt-2 RNAi was combined with cgt-1 RNAi in N2 worms ( Figure 5D, n = 28) and 10% of cgt-1/cgt-2 double RNAi-treated OD70 worms showed the same defects ( Figure 5E and F, n = 42). When cgt-2/cgt-3 double RNAi Glucosylceramide synthesis in germline and embryos was performed, the abnormality was found in 42% of the N2 worms ( Figure 5G, n = 14) and in 40% of the OD70 worms ( Figure 5H and I, n = 20). In dying eggs, the architecture of membranes was malformed, and strong, concentrated fluor- escence was noted (arrows in Figure 5C, F and I). ...
Context 5
... number of abnormal eggs increased to 14% when cgt-2 RNAi was combined with cgt-1 RNAi in N2 worms ( Figure 5D, n = 28) and 10% of cgt-1/cgt-2 double RNAi-treated OD70 worms showed the same defects ( Figure 5E and F, n = 42). When cgt-2/cgt-3 double RNAi Glucosylceramide synthesis in germline and embryos was performed, the abnormality was found in 42% of the N2 worms ( Figure 5G, n = 14) and in 40% of the OD70 worms ( Figure 5H and I, n = 20). In dying eggs, the architecture of membranes was malformed, and strong, concentrated fluor- escence was noted (arrows in Figure 5C, F and I). ...
Context 6
... cgt-2/cgt-3 double RNAi Glucosylceramide synthesis in germline and embryos was performed, the abnormality was found in 42% of the N2 worms ( Figure 5G, n = 14) and in 40% of the OD70 worms ( Figure 5H and I, n = 20). In dying eggs, the architecture of membranes was malformed, and strong, concentrated fluor- escence was noted (arrows in Figure 5C, F and I). Similar experiments were carried out on ER fluorescent worms (WH0327), but no abnormality of ER was found in these worms (data not shown). ...
Context 7
... SAGE experiments have revealed the presence of bre-3, bre-4 and bre-5 gene expression in the germline, but they were not enriched as was cgt-3 ( Wang et al. 2009). If we applied bre-3 RNAi on N2 worms, about 5% of the treated F2 worms showed defects in oocytes and eggs ( Figure 5J, n = 40) and bre-3 RNAi on OD70 worms resulted in the abnormalities in 7% of the worms ( Figure 5K and L, n = 84). In the experiment described in Figure 2, cgt-2 RNAi on tm504 worms resulted in about 15% of the worms showing abnormalities. ...
Context 8
... SAGE experiments have revealed the presence of bre-3, bre-4 and bre-5 gene expression in the germline, but they were not enriched as was cgt-3 ( Wang et al. 2009). If we applied bre-3 RNAi on N2 worms, about 5% of the treated F2 worms showed defects in oocytes and eggs ( Figure 5J, n = 40) and bre-3 RNAi on OD70 worms resulted in the abnormalities in 7% of the worms ( Figure 5K and L, n = 84). In the experiment described in Figure 2, cgt-2 RNAi on tm504 worms resulted in about 15% of the worms showing abnormalities. ...
Similar publications
Background: Gangliosides were found to be associated with Alzheimer's disease (AD). Here we addressed a potential function of γ-secretase (presenilin) dependent cleavage of the amyloid-precursor-protein (APP) in the regulation of ganglioside de novo synthesis. Methods: To identify a potential role of γ-secretase and APP in ganglioside de novo synth...
Altering glycolipid structure by genetic engineering of S. bombicola is a recently started research topic and worthy alternative to the unsuccessful selective feeding strategies conventionally applied to reach this goal. One question to be addressed when expressing heterologous proteins in S. bombicola is the activity of the subsequent biosynthetic...
Steryl glucosides (SG) are abundant steroid conjugates in plant membranes. Beyond structural roles in lipid bilayers, functions
in sugar transport, storage, and/or signalling are predicted. UDP-glucose:sterol glucosyltransferase 80A2 (UGT80A2) and UGT80B1,
which share similarity to fungal counterparts, are implicated in SG synthesis in Arabidopsis...
The UDP-Glc:glycoprotein glucosyltransferase (UGGT) is the sensor of glycoprotein conformations in the glycoprotein folding quality control as it exclusively glucosylates glycoproteins not displaying their native conformations. Monoglucosylated glycoproteins thus formed may interact with the lectin-chaperones calnexin (CNX) and calreticulin (CRT)....
Seeds of oilseed rape (Brassica napus) accumulate high amounts of antinutritive sinapate esters (SE) with sinapoylcholine (sinapine) as major component, accompanied
by sinapoylglucose. These phenolic compounds compromise the use of the protein-rich valuable seed meal. Hence, a substantial
reduction of the SE content is considered essential for esta...
Citations
... Finally, the addition of UDP-glucose to an existing ceramide molecule by ceramide glucosyltransferase (CGT) produces glucosylceramide ( Figure 1A). In C. elegans, there are three ceramide glucosyltransferases, CGT-1, CGT-2, and CGT-3 which are expressed and function in the intestine (Marza et al., 2009;Nomura et al., 2011). ...
... Using mutation or RNAi of each of the genes involved in GlcCer synthesis has probed the biological role of GlcCer in the nematode and has identified an essential role for GlcCer in development as depleting glucosylceramides leads to developmental defects and larval arrest (Lochnit et al., 2005;Laplante and Sabatini, 2009;Nomura et al., 2011;Zhu et al., 2013;Liu and Sabatini, 2020). Specifically, mutations in cgt-3 lead to embryonic defects and reduced brood size. ...
... Specifically, mutations in cgt-3 lead to embryonic defects and reduced brood size. Nomura et al. (2011) showed the cgt-3 (tm504) mutant has abnormal oocytes and eggs that have disrupted membranes and nuclei which accounts for the decreased brood size in this animal. In addition, double mutant cgt-1; cgt-3 arrest at the L1 stage are uncoordinated, and eventually die. ...
Glucosylceramides (GlcCer) are lipids that impact signaling pathways, serve as critical components of cellular membranes, and act as precursors for hundreds of other complex glycolipid species. Abnormal GlcCer metabolism is linked to many diseases, including cancers, diabetes, Gaucher disease, neurological disorders, and skin disorders. A key hurdle to fully understanding the role of GlcCer in disease is the development of methods to accurately detect and quantify these lipid species in a model organism. This will allow for the dissection of the role of this pool in vivo with a focus on all the individual types of GlcCer. In this review, we will discuss the analysis of the GlcCer population specifically in the nematode Caenorhabditis elegans , focusing on the mass spectrometry-based methods available for GlcCer quantification. We will also consider the combination of these approaches with genetic interrogation of GlcCer metabolic genes to define the biological role of these unique lipids. Furthermore, we will explore the implications and obstacles for future research.
... Treatment with RNAi targeting cgt-3 together with cgt-1 or cgt-2 led to gst-4p::GFP inhibition, whereas treatment with double cgt-1 and cgt-2 RNAis had no effect ( Supplementary Fig. 4f). These results are consistent with the major role of CGT-3 in the synthesis of GlcCer in C. elegans 33 . We confirmed the involvement of GlcCer by examining the expression of SKN-1 target genes (Fig. 4c) and the nuclear occupancy of SKN-1::GFP (Fig. 4d) upon cgt-3 knockdown. ...
The contents of numerous membrane lipids change upon ageing. However, it is unknown whether and how any of these changes are causally linked to lifespan regulation. Acyl chains contribute to the functional specificity of membrane lipids. In this study, working with C. elegans , we identified an acyl chain-specific sphingolipid, C22 glucosylceramide, as a longevity metabolite. Germline deficiency, a conserved lifespan-extending paradigm, induces somatic expression of the fatty acid elongase ELO-3, and behenic acid (22:0) generated by ELO-3 is incorporated into glucosylceramide for lifespan regulation. Mechanistically, C22 glucosylceramide is required for the membrane localization of clathrin, a protein that regulates membrane budding. The reduction in C22 glucosylceramide impairs the clathrin-dependent autophagic lysosome reformation, which subsequently leads to TOR activation and longevity suppression. These findings reveal a mechanistic link between membrane lipids and ageing and suggest a model of lifespan regulation by fatty acid-mediated membrane configuration.
... Unlike many other animals, C. elegans has three relevant GT21 ceramide glucosyltransferase genes (cgt-1, cgt-2 and cgt-3); knockout of any three resulted in, e.g., abnormal oocytes and multinucleated embryos. 144 Some of the genetics underlying the following initial stages of the biosynthesis of arthro-series glycosphingolipids is known from Drosophila (i.e., serial transfer of mannose, GlcNAc and GalNAc to form GalNAcb1,4GlcNAcb1,3Manb1,4Glcb1Cer), while the nematode-specific modifications have been partly elucidated by isolation C. elegans bre mutants resistant to one bacterial crystal toxin Cry5B. [145][146][147] The bre-2, bre-3, bre-4 and bre-5 genes encode glycosyltransferases, 48,148 whereby BRE-2 is a GT31 family member and potential GalNAc-modifying b1,3-galactosyltransferase (i.e., the fifth enzyme in the pathway), BRE-3 a GT2 homologue of the Drosophila egghead b1,4-mannosyltransferase (second enzyme), BRE-4 a GT7 family member and proven b1,4-GalNAc transferase (fourth enzyme) and BRE-5, another GT31 family member, homologous to Drosophila Brainiac with b1,3-GlcNAc transferase activity (third enzyme). ...
The nematode worm Caenorhabditis elegans is a widely-used genetic model organism; however, in terms of glycosylation, it is still less than understood. Although the basics of many pathways for the biosynthesis of glycoconjugates in this species are the same as for many eukaryotes, a range of unusual modifications of its N-, O- and lipid-linked glycans are possible. Here, we summarize various aspects of relevant structures, enzymes and functions as well as some open questions for future investigation.
... The much more pronounced downregulation of cgt-2 in ifo-1(kc2) versus nhr-8(hd117) ( Figure 4E) may explain the resulting decrease in d17iso-GlcCer in ifo-1(kc2) but not in nhr-8(hd117) ( Figure 5D). The impaired synthesis of glycosphingolipids could be responsible for the growth defects in ifo-1(kc2) animals in accordance with previous reports, which have shown that the removal of CGT activity induces growth arrest [37,38]. In contrast to previous reports, which have ascribed important functions to CGT-1 and CGT-3, the present study additionally suggests the importance of CGT-2. ...
The intestine is an organ essential to organismal nutrient absorption, metabolic control, barrier function and immunoprotection. The Caenorhabditis elegans intestine consists of 20 cells harboring a dense intermediate filament network positioned below the apical plasma membrane that forms a junction-anchored sheath around the intestinal lumen. This evolutionarily conserved arrangement provides mechanical and overall stress-protection, and it serves as an important model for deciphering the role of intestinal architecture in metazoan biology. We recently reported that the loss-of-function mutation of the intestinal intermediate filament organizer IFO-1 perturbs this architecture, leading to reduced body size and reproduction. Here, we demonstrate that the IFO-1 mutation dramatically affects cholesterol metabolism. Mutants showed an increased sensitivity to cholesterol depletion, reduced cholesterol uptake, and cholesterol transfer to the gonads, which is also observed in worms completely lacking an intermediate filament network. Accordingly, we found striking similarities to transcriptome and lipidome profiles of a nuclear hormone receptor (NHR)-8 mutant. NHR-8 is homologous to mammalian LXR (liver X receptor) that serves as a sterol sensor and transcriptional regulator of lipid metabolism. Remarkably, increasing exogenous cholesterol partially rescues the developmental retardation in IFO-1 mutants. Our results uncover a novel link of the intestinal intermediate filament cytoskeleton to cholesterol metabolism that contributes to compromised growth and reproduction.
... A key step in the synthesis of the complex sphingolipids is the glucosylation of ceramide, which is mediated by ceramide glucosyltransferase (CGT). CGT of C. elegans has been reported to have roles in oocyte formation and early embryonic cell division 47 . A CGT inhibitor, 1-phenyl-2-palmitoylamino-3-morpholino-1-propanol, was able to reduce cyst transformation of Giardia lamblia by 90% 48 . ...
Outbreaks of trichinellosis caused by Trichinella papuae have been reported in South-East Asia. Mebendazole and thiabendazole are the treatments of choice for trichinellosis; however, both drugs result in significant side effects and are less effective for muscle-stage larvae (L1). An alternative therapeutic agent is needed to improve treatment. Information on lipid composition and metabolic pathways may bridge gaps in our knowledge and lead to new antiparasitics. The T. papuae L1 lipidome was analysed using a mass spectrometry-based approach, and 403 lipid components were identified. Eight lipid classes were found and glycerophospholipids were dominant, corresponding to 63% of total lipids, of which the glycerolipid DG (20:1[11Z]/22:4[7Z,10Z,13Z,16Z]/0:0) (iso2) was the most abundant. Overall, 57% of T. papuae lipids were absent in humans; therefore, lipid metabolism may be dissimilar in the two species. Proteins involved T. papuae lipid metabolism were explored using bioinformatics. We found that 4-hydroxybutyrate coenzyme A transferase, uncharacterized protein (A0A0V1MCB5) and ML-domain-containing protein are not present in humans. T. papuae glycerophospholipid metabolic and phosphatidylinositol dephosphorylation processes contain several proteins that are dissimilar to those in humans. These findings provide insights into T. papuae lipid composition and metabolism, which may facilitate the development of novel trichinellosis treatments.
... The generic "ceramide" family is comprised of >50 distinct molecular species, of which about 25 were identified in the current study ( Fig. 4 and Table S1). Pharmacological inhibition of SPT extends life span in both Caenorhabditis elegans 67 and yeast 68 and ceramide glucosyltransferase is involved in oocyte formation and early embryonic cell division in C. elegans 69 . Interestingly, SPT was detected in AM but not in RE 17 . ...
Numerous aquatic invertebrates survive harsh environments by displaying dormancy as encysted embryos. This study aimed at determining whether metabolomics could provide molecular insight to explain the “dormancy syndrome” by highlighting functional pathways and metabolites, hence offering a novel comprehensive molecular view of dormancy. We compared the metabolome of morphologically distinct dormant encysted embryos (resting eggs) and non-dormant embryos (amictic eggs) of a rotifer (Brachionus plicatilis). Metabolome profiling revealed ~5,000 features, 1,079 of which were annotated. Most of the features were represented at significantly higher levels in non-dormant than dormant embryos. A large number of features was assigned to putative functional pathways indicating novel differences between dormant and non-dormant states. These include features associated with glycolysis, the TCA and urea cycles, amino acid, purine and pyrimidine metabolism. Interestingly, ATP, nucleobases, cyclic nucleotides, thymidine and uracil, were not detected in dormant resting eggs, suggesting an impairment of response to environmental and internal cues, cessation of DNA synthesis, transcription and plausibly translation in the dormant embryos. The levels of trehalose or its analogues, with a role in survival under desiccation conditions, were higher in resting eggs. In conclusion, the current study highlights metabolomics as a major analytical tool to functionally compare dormancy across species.
... A variety of neutral and zwitterionic glycolipids have been described in C. elegans (see Figure 4), which are altered in the so-called bre mutants resistant to a Bacillus crystal toxin (Gerdt et al., 1997(Gerdt et al., , 1999Griffitts et al., 2005). Some relevant enzymes required for glycolipid biosynthesis (the CGT glucosyltransferases, the BRE-1/GMD-1 GDP-mannose dehydratase and an N-acetylgalactosaminyltransferase BRE-4) have been characterized in vitro (Kawar et al., 2002;Rhomberg et al., 2006;Nomura et al., 2011). Other classes of glycolipids include phosphoethanolamine glucosylceramides and the ascarosides, which both have signaling functions (Boland et al., 2017;von Reuss, 2018). ...
Caenorhabditis elegans is a genetically well-studied model nematode or “worm”; however, its N-glycomic complexity is actually baffling and still not completely unraveled. Some features of its N-glycans are, to date, unique and include bisecting galactose and up to five fucose residues associated with the asparagine-linked Man2−3GlcNAc2 core; the substitutions include galactosylation of fucose, fucosylation of galactose and methylation of mannose or fucose residues as well as phosphorylcholine on antennal (non-reducing) N-acetylglucosamine. Only some of these modifications are shared with various other nematodes, while others have yet to be detected in any other species. Thus, C. elegans can be used as a model for some aspects of N-glycan function, but its glycome is far from identical to those of other organisms and is actually far from simple. Possibly the challenges of its native environment, which differ from those of parasitic or necromenic species, led to an anatomically simple worm possessing a complex glycome.
... The fructose bisphosphate aldolase is a key enzyme of the glycolytic pathway [79] and its inhibition may contribute to accumulation of sugar molecules on the cuticle surface [30]. Similarly, glucosyl transferase catalyzes the transfer of sugar moieties to a wide range of acceptor molecules [73,112]. RNAi induced in vitro inhibition of both these transcripts reduced the endospore attachment on nematode surface. ...
Background
Southern root-knot nematode Meloidogyne incognita (Kofoid and White, 1919), Chitwood, 1949 is a key pest of agricultural crops. Pasteuria penetrans is a hyperparasitic bacterium capable of suppressing the nematode reproduction, and represents a typical coevolved pathogen-hyperparasite system. Attachment of Pasteuria endospores to the cuticle of second-stage nematode juveniles is the first and pivotal step in the bacterial infection. RNA-Seq was used to understand the early transcriptional response of the root-knot nematode at 8 h post Pasteuria endospore attachment.
Results
A total of 52,485 transcripts were assembled from the high quality (HQ) reads, out of which 582 transcripts were found differentially expressed in the Pasteuria endospore encumbered J2 s, of which 229 were up-regulated and 353 were down-regulated. Pasteuria infection caused a suppression of the protein synthesis machinery of the nematode. Several of the differentially expressed transcripts were putatively involved in nematode innate immunity, signaling, stress responses, endospore attachment process and post-attachment behavioral modification of the juveniles. The expression profiles of fifteen selected transcripts were validated to be true by the qRT PCR. RNAi based silencing of transcripts coding for fructose bisphosphate aldolase and glucosyl transferase caused a reduction in endospore attachment as compared to the controls, whereas, silencing of aspartic protease and ubiquitin coding transcripts resulted in higher incidence of endospore attachment on the nematode cuticle.
Conclusions
Here we provide evidence of an early transcriptional response by the nematode upon infection by Pasteuria prior to root invasion. We found that adhesion of Pasteuria endospores to the cuticle induced a down-regulated protein response in the nematode. In addition, we show that fructose bisphosphate aldolase, glucosyl transferase, aspartic protease and ubiquitin coding transcripts are involved in modulating the endospore attachment on the nematode cuticle. Our results add new and significant information to the existing knowledge on early molecular interaction between M. incognita and P. penetrans.
Electronic supplementary material
The online version of this article (10.1186/s12864-018-5230-8) contains supplementary material, which is available to authorized users.
... Cers have been found to be toxic to oocytes and embryos both in vivo and in vitro (20), and the inhibition of acidsphingomyelinases, the enzyme that hydrolyzes SMs to Cers, blocks apoptosis in oocytes (21). GlcCer synthase, the enzyme that catalyzes the glycosylation of Cers to form GlcCers, is essential for oocyte and embryo membrane formation (22,23). These studies lay the foundation of demonstrating the importance of sphingolipids in fertility and potentially EAI, but they do not clarify which sphingolipid species are associated with EAI. ...
Objective:
To characterize the peritoneal fluid (PF) sphingolipid profile in endometriosis-associated infertility (EAI), and to assess the plausible functional role(s) of ceramides in oocyte maturation potential.
Design:
Retrospective case-control study and in vitro mouse oocyte study.
Setting:
University-affiliated hospital and university laboratory.
Subjects:
Twenty-seven infertile patients diagnosed with endometriosis and 20 infertile patients who did not have endometriosis; BALB/c female mice.
Intervention(s):
None.
Main outcome measure(s):
PF sphingolipid concentrations. Number of metaphase II (MII) mouse oocytes.
Result(s):
Liquid chromatography-tandem mass spectrometry revealed 11 significantly elevated PF sphingolipids in infertile women with severe endometriosis compared with infertile women without endometriosis (change >50%, false discovery rate ≤10%). Logistic regression analysis identified three very-long-chain ceramides potentially associated with EAI. Functional studies revealed that very-long-chain ceramides may compromise or induce murine MII oocyte maturation. The oocyte maturation effects induced by the very long-chain ceramides were triggered by alterations in mitochondrial superoxide production in a concentration-dependent manner. Scavenging of mitochondrial superoxide reversed the maturation effects of C24:0 ceramide.
Conclusion(s):
EAI is associated with accumulation of PF very-long-chain ceramides. Mouse studies demonstrated how ceramides affect MII oocyte maturation, mediating through mitochondrial superoxide. These results provide an opportunity for direct functional readout of pathophysiology in EAI, and future therapies targeted at this sphingolipid metabolism may be harnessed for improved oocyte maturation.
... It may thus be surmised that mechanical properties of the cell membrane changed, leading to delayed cytokinesis and increased binucleation of hepatocytes. Our results corroborate data from Nomura et al. [64] showing that Ugcg-deficient C. elegans embryos developed multinucleated cells causing decreased brood size of the animals. Also AtillaGokcumen et al. demonstrated that alterations of the lipid content in HeLa S3 cells, achieved by siRNA treatments of synthesizing enzymes, changed their physical properties and caused cytokinesis failure [65]. ...
Hepatocellular carcinoma (HCC) is one of the most frequent cancers. In vitro studies suggest that growth and response to therapy of human carcinomas may depend on glycosphingolipid (GSL) expression. Glucosylceramide synthase (GCS), encoded by the gene Ugcg, is the basic enzyme required for the synthesis of GSLs. Gene array analysis implied that Ugcg is significantly overexpressed in human HCC as compared to non-tumorous liver tissue. Therefore we have investigated whether tumor - genesis and - growth is altered in the absence of GSLs. An endogenous liver cancer model has been initiated by application of diethylnitrosamine in mice lacking Ugcg specifically in hepatocytes. We have now shown that hepatocellular tumor initiation and growth in mice is significantly inhibited by hepatic GSL deficiency in vivo. Neither the expression of cell cycle proteins, such as cyclins and pathways such as the MAP-kinase/Erk pathway nor the mTOR/Akt pathway as well as the number of liver infiltrating macrophages and T cells were essentially changed in tumors lacking GSLs. Significantly elevated bi-nucleation of atypical hepatocytes, a feature for impaired cytokinesis, was detected in tumors of mice lacking liver-specific GSLs. A reduction of proliferation and restricted growth of tumor microspheres due to delayed, GSL-dependent cytokinesis, analogous to the histopathologic phenotype in vivo could be demonstrated in vitro. GSL synthesis inhibition may thus constitute a potential therapeutic target for hepatocellular carcinoma.
