GLUT5 (SLC2A5) enables fructose-mediated proliferation independent of ketohexokinase

Abstract

Background Fructose is an abundant source of carbon and energy for cells to use for metabolism, but only certain cell types use fructose to proliferate. Tumor cells that acquire the ability to metabolize fructose have a fitness advantage over their neighboring cells, but the proteins that mediate fructose metabolism in this context are unknown. Here, we investigated the determinants of fructose-mediated cell proliferation. Methods Live cell imaging and crystal violet assays were used to characterize the ability of several cell lines (RKO, H508, HepG2, Huh7, HEK293T (293T), A172, U118-MG, U87, MCF-7, MDA-MB-468, PC3, DLD1 HCT116, and 22RV1) to proliferate in fructose (i.e., the fructolytic ability). Fructose metabolism gene expression was determined by RT-qPCR and western blot for each cell line. A positive selection approach was used to “train” non-fructolytic PC3 cells to utilize fructose for proliferation. RNA-seq was performed on parental and trained PC3 cells to find key transcripts associated with fructolytic ability. A CRISPR-cas9 plasmid containing KHK-specific sgRNA was transfected in 293T cells to generate KHK-/- cells. Lentiviral transduction was used to overexpress empty vector, KHK, or GLUT5 in cells. Metabolic profiling was done with seahorse metabolic flux analysis as well as LC/MS metabolomics. Cell Titer Glo was used to determine cell sensitivity to 2-deoxyglucose in media containing either fructose or glucose. Results We found that neither the tissue of origin nor expression level of any single gene related to fructose catabolism determine the fructolytic ability. However, cells cultured chronically in fructose can develop fructolytic ability. SLC2A5, encoding the fructose transporter, GLUT5, was specifically upregulated in these cells. Overexpression of GLUT5 in non-fructolytic cells enabled growth in fructose-containing media across cells of different origins. GLUT5 permitted fructose to flux through glycolysis using hexokinase (HK) and not ketohexokinase (KHK). Conclusions We show that GLUT5 is a robust and generalizable driver of fructose-dependent cell proliferation. This indicates that fructose uptake is the limiting factor for fructose-mediated cell proliferation. We further demonstrate that cellular proliferation with fructose is independent of KHK.

Loss of the fructose transporter SLC2A5 inhibits cancer cell migration

“…The gene encoding SLC2A5, a fructose-specific transporter, is highly expressed in cancers whereas it is tightly regulated in healthy tissues ( Douard & Ferraris, 2008 ). Specifically, increased abundance of both SLC2A5 mRNA and protein have been associated with cancer progression and increased frequency of metastasis of many cancers ( Zamora-Leon et al, 1996 ; Chen et al, 2016 ; Bu et al, 2018 ; Hamann et al, 2018 ; Weng et al, 2018 ; Jin et al, 2019 ; Chen et al, 2020 ; Liang et al, 2021 ; Lin et al, 2021 ). In this study, we assessed the importance of the SLC2A5 gene on cancer cell proliferation, migration, extravasation, and colony formation.…”

“…Fructose can be metabolized to fatty acids and triglycerides, providing components essential for the synthesis of membrane lipids to sustain cancer growth and proliferation ( Ter Horst and Serlie, 2017 ). Recent studies have also shown that fructose supplementation stimulates lung cancer cell proliferation in vivo ( Chen et al, 2020 ; Liang et al, 2021 ). Accordingly, we found that supplementation of the cell culture medium with fructose robustly stimulated MIA-PaCa-2 cell proliferation while inactivation of the gene encoding SLC2A5 abolished this response.…”

“…SLC2A5 expression is higher in metastatic liver lesions than in normal liver; and elevated in primary lung tumors; as well as in brain, colon, testis, and uterine cancers, including breast carcinoma cell lines ( Uldry & Thorens, 2004 ). Moreover, recent studies have shown that fructose serves as an efficient energy source for lung cancer cells ( Chen et al, 2020 ) and facilitates tumor cell proliferation ( Liang et al, 2021 ).…”

Fructose Metabolism and Acute Myeloid Leukemia

“…Interestingly, recent evidence from 13 different cell lines from five different originating tissues showed that the ability to metabolize fructose is not tissue site-dependent, since cells that chronically live in an environment containing fructose upregulate GLUT5 and develop the ability to metabolize fructose using hexokinase instead of fructokinase. 77 In contrast with these in vitro studies, genetically deleting fructokinase in mice actually prevented the development of metabolic syndrome. 78…”

Establishing mammalian GLUT kinetics and lipid preferences in a reconstituted-liposome system

“…GLUT5 is highly expressed in breast cancer cells 19,50 39,51 . We investigated D-fructose uptake by GLUT5 in the presence of 200 µM of MSNBA, but find no competition for transport (Fig.…”