Upregulation of human glycolipid transfer protein (GLTP) induces necroptosis in colon carcinoma cells

Shrawan Kumar Mishra; Daniel J. Stephenson; Charles E. Chalfant; Rhoderick E. Brown

doi:10.1016/j.bbalip.2018.11.002

Upregulation of human glycolipid transfer protein (GLTP) induces necroptosis in colon carcinoma cells

Shrawan Kumar Mishra, Daniel J. Stephenson, Charles E. Chalfant, Rhoderick E. Brown

The Hormel Institute

Research output: Contribution to journal › Article › peer-review

21 Scopus citations

Abstract

Human GLTP on chromosome 12 (locus 12q24.11) encodes a 24 kD amphitropic lipid transfer protein (GLTP) that mediates glycosphingolipid (GSL) intermembrane trafficking and regulates GSL homeostatic levels within cells. Herein, we provide evidence that GLTP overexpression inhibits the growth of human colon carcinoma cells (HT-29; HCT-116), but spares normal colonic cells (CCD-18Co). Mechanistic studies reveal that GLTP overexpression arrested the cell cycle at the G1/S checkpoint via upregulation of cyclin-dependent kinase inhibitor-1B (Kip1/p27) and cyclin-dependent kinase inhibitor 1A (Cip1/p21) at the protein and mRNA levels, and downregulation of cyclin-dependent kinase-2 (CDK2), cyclin-dependent kinase-4 (CDK4), cyclin E and cyclin D1 protein levels. Assessment of the biological fate of HCT-116 cells overexpressing GLTP indicated no increase in cell death suggesting induction of quiescence. However, HT-29 cells overexpressing GLTP underwent cell death by necroptosis as revealed by phosphorylation of human mixed lineage kinase domain-like protein (pMLKL) via receptor-interacting protein kinase-3 (RIPK-3), elevated cytosolic calcium, and plasma membrane permeabilization by pMLKL oligomerization. Overexpression of W96A-GLTP, an ablated GSL binding site mutant, failed to arrest the cell cycle or induce necroptosis. Sphingolipid assessment (ceramide, monohexosylceramide, sphingomyelin, ceramide-1-phosphate, sphingosine, and sphingosine-1-phosphate) of HT-29 cells overexpressing GLTP revealed large decreases (>5-fold) in sphingosine-1-phosphate with minimal change in 16:0-ceramide, tipping the ‘sphingolipid rheostat’ (S1P/16:0-Cer ratio) towards cell death. Depletion of RIPK-3 or MLKL abrogated necroptosis induced by GLTP overexpression. Our findings establish GLTP upregulation as a previously unknown suppressor of human colon carcinoma HT-29 cells via interference with cell cycle progression and induction of necroptosis.

Original language	English (US)
Pages (from-to)	158-167
Number of pages	10
Journal	Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
Volume	1864
Issue number	2
DOIs	https://doi.org/10.1016/j.bbalip.2018.11.002
State	Published - Feb 2019

Bibliographical note

Funding Information:
Research support provided by NIH HL-125353 (CEC & REB), GM-45928 (REB), HD087198 (CEC), RR031535 (CEC), the Veteran's Administration (VA Merit Review, I BX001792 (CEC)), Research Career Scientist Award 13F-RCS-002 (CEC), Southern Minnesota Paint-the-Town Pink Cancer Research Funds (REB) and the Hormel Foundation (REB) is gratefully acknowledged.

Keywords

Colon cancer
Glycolipid transfer protein
Necroptosis
Programmed cell death
Sphingolipid rheostat
Sphingolipidomics

PubMed: MeSH publication types

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1016/j.bbalip.2018.11.002

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6448591

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title = "Upregulation of human glycolipid transfer protein (GLTP) induces necroptosis in colon carcinoma cells",

abstract = "Human GLTP on chromosome 12 (locus 12q24.11) encodes a 24 kD amphitropic lipid transfer protein (GLTP) that mediates glycosphingolipid (GSL) intermembrane trafficking and regulates GSL homeostatic levels within cells. Herein, we provide evidence that GLTP overexpression inhibits the growth of human colon carcinoma cells (HT-29; HCT-116), but spares normal colonic cells (CCD-18Co). Mechanistic studies reveal that GLTP overexpression arrested the cell cycle at the G1/S checkpoint via upregulation of cyclin-dependent kinase inhibitor-1B (Kip1/p27) and cyclin-dependent kinase inhibitor 1A (Cip1/p21) at the protein and mRNA levels, and downregulation of cyclin-dependent kinase-2 (CDK2), cyclin-dependent kinase-4 (CDK4), cyclin E and cyclin D1 protein levels. Assessment of the biological fate of HCT-116 cells overexpressing GLTP indicated no increase in cell death suggesting induction of quiescence. However, HT-29 cells overexpressing GLTP underwent cell death by necroptosis as revealed by phosphorylation of human mixed lineage kinase domain-like protein (pMLKL) via receptor-interacting protein kinase-3 (RIPK-3), elevated cytosolic calcium, and plasma membrane permeabilization by pMLKL oligomerization. Overexpression of W96A-GLTP, an ablated GSL binding site mutant, failed to arrest the cell cycle or induce necroptosis. Sphingolipid assessment (ceramide, monohexosylceramide, sphingomyelin, ceramide-1-phosphate, sphingosine, and sphingosine-1-phosphate) of HT-29 cells overexpressing GLTP revealed large decreases (>5-fold) in sphingosine-1-phosphate with minimal change in 16:0-ceramide, tipping the {\textquoteleft}sphingolipid rheostat{\textquoteright} (S1P/16:0-Cer ratio) towards cell death. Depletion of RIPK-3 or MLKL abrogated necroptosis induced by GLTP overexpression. Our findings establish GLTP upregulation as a previously unknown suppressor of human colon carcinoma HT-29 cells via interference with cell cycle progression and induction of necroptosis.",

keywords = "Colon cancer, Glycolipid transfer protein, Necroptosis, Programmed cell death, Sphingolipid rheostat, Sphingolipidomics",

author = "Mishra, {Shrawan Kumar} and Stephenson, {Daniel J.} and Chalfant, {Charles E.} and Brown, {Rhoderick E.}",

note = "Funding Information: Research support provided by NIH HL-125353 (CEC & REB), GM-45928 (REB), HD087198 (CEC), RR031535 (CEC), the Veteran's Administration (VA Merit Review, I BX001792 (CEC)), Research Career Scientist Award 13F-RCS-002 (CEC), Southern Minnesota Paint-the-Town Pink Cancer Research Funds (REB) and the Hormel Foundation (REB) is gratefully acknowledged. ",

year = "2019",

month = feb,

doi = "10.1016/j.bbalip.2018.11.002",

language = "English (US)",

volume = "1864",

pages = "158--167",

journal = "Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids",

issn = "1388-1981",

publisher = "Elsevier B.V.",

number = "2",

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TY - JOUR

T1 - Upregulation of human glycolipid transfer protein (GLTP) induces necroptosis in colon carcinoma cells

AU - Mishra, Shrawan Kumar

AU - Stephenson, Daniel J.

AU - Chalfant, Charles E.

AU - Brown, Rhoderick E.

N1 - Funding Information: Research support provided by NIH HL-125353 (CEC & REB), GM-45928 (REB), HD087198 (CEC), RR031535 (CEC), the Veteran's Administration (VA Merit Review, I BX001792 (CEC)), Research Career Scientist Award 13F-RCS-002 (CEC), Southern Minnesota Paint-the-Town Pink Cancer Research Funds (REB) and the Hormel Foundation (REB) is gratefully acknowledged.

PY - 2019/2

Y1 - 2019/2

N2 - Human GLTP on chromosome 12 (locus 12q24.11) encodes a 24 kD amphitropic lipid transfer protein (GLTP) that mediates glycosphingolipid (GSL) intermembrane trafficking and regulates GSL homeostatic levels within cells. Herein, we provide evidence that GLTP overexpression inhibits the growth of human colon carcinoma cells (HT-29; HCT-116), but spares normal colonic cells (CCD-18Co). Mechanistic studies reveal that GLTP overexpression arrested the cell cycle at the G1/S checkpoint via upregulation of cyclin-dependent kinase inhibitor-1B (Kip1/p27) and cyclin-dependent kinase inhibitor 1A (Cip1/p21) at the protein and mRNA levels, and downregulation of cyclin-dependent kinase-2 (CDK2), cyclin-dependent kinase-4 (CDK4), cyclin E and cyclin D1 protein levels. Assessment of the biological fate of HCT-116 cells overexpressing GLTP indicated no increase in cell death suggesting induction of quiescence. However, HT-29 cells overexpressing GLTP underwent cell death by necroptosis as revealed by phosphorylation of human mixed lineage kinase domain-like protein (pMLKL) via receptor-interacting protein kinase-3 (RIPK-3), elevated cytosolic calcium, and plasma membrane permeabilization by pMLKL oligomerization. Overexpression of W96A-GLTP, an ablated GSL binding site mutant, failed to arrest the cell cycle or induce necroptosis. Sphingolipid assessment (ceramide, monohexosylceramide, sphingomyelin, ceramide-1-phosphate, sphingosine, and sphingosine-1-phosphate) of HT-29 cells overexpressing GLTP revealed large decreases (>5-fold) in sphingosine-1-phosphate with minimal change in 16:0-ceramide, tipping the ‘sphingolipid rheostat’ (S1P/16:0-Cer ratio) towards cell death. Depletion of RIPK-3 or MLKL abrogated necroptosis induced by GLTP overexpression. Our findings establish GLTP upregulation as a previously unknown suppressor of human colon carcinoma HT-29 cells via interference with cell cycle progression and induction of necroptosis.

AB - Human GLTP on chromosome 12 (locus 12q24.11) encodes a 24 kD amphitropic lipid transfer protein (GLTP) that mediates glycosphingolipid (GSL) intermembrane trafficking and regulates GSL homeostatic levels within cells. Herein, we provide evidence that GLTP overexpression inhibits the growth of human colon carcinoma cells (HT-29; HCT-116), but spares normal colonic cells (CCD-18Co). Mechanistic studies reveal that GLTP overexpression arrested the cell cycle at the G1/S checkpoint via upregulation of cyclin-dependent kinase inhibitor-1B (Kip1/p27) and cyclin-dependent kinase inhibitor 1A (Cip1/p21) at the protein and mRNA levels, and downregulation of cyclin-dependent kinase-2 (CDK2), cyclin-dependent kinase-4 (CDK4), cyclin E and cyclin D1 protein levels. Assessment of the biological fate of HCT-116 cells overexpressing GLTP indicated no increase in cell death suggesting induction of quiescence. However, HT-29 cells overexpressing GLTP underwent cell death by necroptosis as revealed by phosphorylation of human mixed lineage kinase domain-like protein (pMLKL) via receptor-interacting protein kinase-3 (RIPK-3), elevated cytosolic calcium, and plasma membrane permeabilization by pMLKL oligomerization. Overexpression of W96A-GLTP, an ablated GSL binding site mutant, failed to arrest the cell cycle or induce necroptosis. Sphingolipid assessment (ceramide, monohexosylceramide, sphingomyelin, ceramide-1-phosphate, sphingosine, and sphingosine-1-phosphate) of HT-29 cells overexpressing GLTP revealed large decreases (>5-fold) in sphingosine-1-phosphate with minimal change in 16:0-ceramide, tipping the ‘sphingolipid rheostat’ (S1P/16:0-Cer ratio) towards cell death. Depletion of RIPK-3 or MLKL abrogated necroptosis induced by GLTP overexpression. Our findings establish GLTP upregulation as a previously unknown suppressor of human colon carcinoma HT-29 cells via interference with cell cycle progression and induction of necroptosis.

KW - Colon cancer

KW - Glycolipid transfer protein

KW - Necroptosis

KW - Programmed cell death

KW - Sphingolipid rheostat

KW - Sphingolipidomics

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SN - 1388-1981

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EP - 167

JO - Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids

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ER -

Upregulation of human glycolipid transfer protein (GLTP) induces necroptosis in colon carcinoma cells

Abstract

Bibliographical note

Keywords

PubMed: MeSH publication types

UN SDGs

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