Functional role of glycosphingolipids in contact inhibition of growth in a human mammary epithelial cell line

Huang, Xiaohua; Schurman, Nathan; Handa, Kazuko; Hakomori, Sen‐itiroh

doi:10.1002/1873-3468.12709

Cited by 11 publications

(9 citation statements)

References 50 publications

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“…We further characterized the impacts of Gb3‐cSrc association on drug resistance in WiDr cells carrying the R273H TP53 mutation. Previous reports from other laboratories showed that addition of exogenous GM3, Gb3 and GD3 altered cell viability and other behaviors 50,51 . Shiga toxin 1B subunit (STxB or verotoxin) specifically bound to Gb3 and induced cancer cell death 52–54 .…”

Section: Resultsmentioning

confidence: 96%

Gb3‐cSrc complex in glycosphingolipid‐enriched microdomains contributes to the expression of p53 mutant protein and cancer drug resistance via β‐catenin–activated RNA methylation

et al. 2020

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Glucosylceramide synthase (GCS) is a key enzyme catalyzing ceramide glycosylation to generate glucosylceramide (GlcCer), which in turn serves as the precursor for cells to produce glycosphingolipids (GSLs). In cell membranes, GSLs serve as essential components of GSL‐enriched microdomains (GEMs) and mediate membrane functions and cell behaviors. Previous studies showed that ceramide glycosylation correlates with upregulated expression of p53 hotspot mutant R273H and cancer drug resistance. Yet, the underlying mechanisms remain elusive. We report herewith that globotriaosylceramide (Gb3) is associated with cSrc kinase in GEMs and plays a crucial role in modulating expression of p53 R273H mutant and drug resistance. Colon cancer cell lines, either WiDr homozygous for missense‐mutated TP53 (R273H+/+) or SW48/TP53‐Dox bearing heterozygous TP53 mutant (R273H/+), display drug resistance with increased ceramide glycosylation. Inhibition of GCS with Genz‐161 (GENZ 667161) resensitized cells to apoptosis in these p53 mutant‐carrying cancer cells. Genz‐161 effectively inhibited GCS activity, and substantially suppressed the elevated Gb3 levels seen in GEMs of p53‐mutant cells exposed to doxorubicin. Complex formation between Gb3 and cSrc in GEMs to activate β‐catenin was detected in both cultured cells and xenograft tumors. Suppression of ceramide glycosylation significantly decreased Gb3‐cSrc in GEMs, β‐catenin, and methyltransferase‐like 3 for m6A RNA methylation, thus altering pre‐mRNA splicing, resulting in upregulated expression of wild‐type p53 protein, but not mutants, in cells carrying p53 R273H. Altogether, increased Gb3‐cSrc complex in GEMs of membranes in response to anticancer drug induced cell stress promotes expression of p53 mutant proteins and accordant cancer drug resistance.

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Section: Resultsmentioning

confidence: 96%

Gb3‐cSrc complex in glycosphingolipid‐enriched microdomains contributes to the expression of p53 mutant protein and cancer drug resistance via β‐catenin–activated RNA methylation

et al. 2020

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“…The enzyme encoded by the A4GALT gene catalyzes the synthesis of P1 and PK antigens and is also widely used in P1PK blood group systematic typing due to its gene polymorphism [34] . In addition, studies have reported that knocking down A4GALT can significantly inhibit contact inhibition, and exogenous adding A4GALT can inhibit the proliferation of low-density cells, suggesting that A4GALT is involved in the 'contact inhibition' of cell growth [35] . Other studies have found that A4GALT-mediated GSLs can enhance the metastasis ability of mesenchymal and epithelial cancer cells, and the loss of A4GALT mediated by CRISPR-Cas9 can weaken galactose induced epithelial-mesenchymal transformation (EMT) [36] .…”

Section: Discussionmentioning

confidence: 99%

A4GALT mediated the glycosylation of B7-H3 in human colorectal cancer cell lines

Han

Chen³

et al. 2022

Preprint

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B7-H3 is one of the most important members of the B7/CD28 family, and its expression level is abnormally high in a variety of tumors. B7-H3 inhibits T cell activation via binding to the corresponding receptors on T cells, thereby mediating tumor immune escape. Glycosylation is a common post-translational modification of proteins, which plays an essential role in protein expression patterns and biological functions. Current evidence has shown that the abnormal glycosylation of B7-H3 in tumors is of great significance for protein expression and ligand-receptor binding. Therefore, in-depth exploration of the underlying mechanism of glycosylation modification of B7-H3 is expected to provide new insights for tumor immunotherapy. To investigate the underlying mechanism of glycosyltransferase-mediated glycosylation of B7-H3. Firstly, the CHIPBase database was used to screen glycosyltransferases with a high correlation with the protein expression of B7-H3. Then their siRNAs were designed and synthesized to transfect into cells, and the western blotting assay was performed for further screening. Secondly, the siRNA and overexpression plasmid of the screened glycosyltransferase were respectively transfected into cells to verify the effect on the expression level of B7-H3. Thirdly, the effect of glycosyltransferase on the expression level of B7-H3 was explored by changing its substrate level. Finally, the co-immunoprecipitation experiment was conducted to verify whether protein binding existed between B7-H3 and the glycosyltransferase. The glycosyltransferase called A4GALT had the highest correlation with the protein expression of B7-H3. After knocking down or overexpressing A4GALT, the protein expression level of B7-H3 both changed significantly. When knocked down GALT to reduce the galactosyl donors, B7-H3 was significantly down-regulated. And B7-H3 was up-regulated while increasing the galactose in the medium. In addition, the Co-immunoprecipitation experiment proved that there was protein binding between B7-H3 and A4GALT. A4GALT can positively regulate the expression of B7-H3, and changing the level of galactosyl donors can also positively regulate the expression of B7-H3. There is a protein interaction between A4GALT and B7-H3.

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“…Cholera toxin is produced by Vibrio cholerae and is a multifunctional protein that influences various cells, including the immune system, and which also acts as an anti-inflammatory agent (Beharati K and Ganguly NK, 2011; Baldauf K et al, 2015). Cholera toxin was shown to suppress carcinogenesis in colon cancer (Doulberis M et al, 2015) and may exert contact inhibition by binding to glycosphingolipids in MCF10A cells (Huang X et al, 2017). One of the functions of cholera toxin is to activate adenylate cyclase, which increases the intracellular cAMP level.…”

Section: Discussionmentioning

confidence: 99%

Hypo-osmotic Stress Induces ATP Release via Volume-regulated Anion Channels in Undifferentiated Mammary Cells

Furuya

Takahashi

Hirata

et al. 2021

Preprint

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The high interstitial ATP concentration in the cancer microenvironment is a major source of adenosine, which acts as a strong immune suppressor. However, the source of ATP release has not been elucidated. We measured the ATP release during hypotonic stress using a real-time ATP luminescence imaging system in primary cultured mammary cells and in breast cell lines. In primary cultured cells, ATP was intermittently released with transient-sharp peaks, while in breast cell lines ATP was released with a slowly rising diffuse pattern. The diffuse ATP release pattern was changed to a transient-sharp pattern by cholera toxin treatment and the reverse change was induced by transforming growth factor (TGF) β treatment. DCPIB, an inhibitor of volume-regulated anion channels (VRACs), only suppressed the diffuse pattern. The inflammatory mediator sphingosine-1-phosphate (S1P) induced a diffuse ATP release pattern isovolumetrically. The knockdown of A isoform of leucine-rich repeat-containing protein 8 (LRRC8A), the essential molecular entity of VRACs, using shRNA suppressed the diffuse pattern. These results suggest that abundantly expressed VRACs are a conduit of ATP release in undifferentiated cells, including cancer cells.

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Functional role of glycosphingolipids in contact inhibition of growth in a human mammary epithelial cell line

Cited by 11 publications

References 50 publications

Gb3‐cSrc complex in glycosphingolipid‐enriched microdomains contributes to the expression of p53 mutant protein and cancer drug resistance via β‐catenin–activated RNA methylation

Gb3‐cSrc complex in glycosphingolipid‐enriched microdomains contributes to the expression of p53 mutant protein and cancer drug resistance via β‐catenin–activated RNA methylation

A4GALT mediated the glycosylation of B7-H3 in human colorectal cancer cell lines

Hypo-osmotic Stress Induces ATP Release via Volume-regulated Anion Channels in Undifferentiated Mammary Cells

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