Inhibition of glucose metabolism prevents glycosylation of the glutamine transporter ASCT2 and promotes compensatory LAT1 upregulation in leukemia cells.

Florence Polet,Adan Pinto,Ruben Martherus,Cyril Corbet,Olivier Feron

doi:10.18632/oncotarget.10131

Abstract

Leukemia cells are highly dependent on glucose and glutamine as bioenergetic and biosynthetic fuels. Inhibition of the metabolism of glucose but also of glutamine is thus proposed as a therapeutic modality to block leukemia cell growth. Since glucose also supports protein glycosylation, we wondered whether part of the growth inhibitory effects resulting from glycolysis inhibition could indirectly result from a defect in glycosylation of glutamine transporters. We found that ASCT2/SLC1A5, a major glutamine transporter, was indeed deglycosylated upon glucose deprivation and 2-deoxyglucose exposure in HL-60 and K-562 leukemia cells. Inhibition of glycosylation by these modalities as well as by the bona fide glycosylation inhibitor tunicamycin however marginally influenced glutamine transport and did not impact on ASCT2 subcellular location. This work eventually unraveled the dispensability of ASCT2 to support HL-60 and K-562 leukemia cell growth and identified the upregulation of the neutral amino acid antiporter LAT1/SLC7A5 as a mechanism counteracting the inhibition of glycosylation. Pharmacological inhibition of LAT1 increased the growth inhibitory effects and the inactivation of the mTOR pathway resulting from glycosylation defects, an effect further emphasized during the regrowth period post-treatment with tunicamycin. In conclusion, this study points towards the underestimated impact of glycosylation inhibition in the interpretation of metabolic alterations resulting from glycolysis inhibition, and identifies LAT1 as a therapeutic target to prevent compensatory mechanisms induced by alterations in the glycosylating process.

Highlights

Many transporters and receptors are N-glycosylated, and their cell-surface expression depends on proper folding and the degree of N-glycan branching [1]
We found that in the absence of Glc, HL-60 cells exhibited over time a shift of ASCT2 immunoblot signal towards lower molecular weight (MW) (Figure 2A)
The lower ASCT2 MW band observed upon glucose deprivation was observed when HL60 cells were exposed to either inhibitor (Figure 2B)

Summary

Introduction

Many transporters and receptors are N-glycosylated, and their cell-surface expression depends on proper folding and the degree of N-glycan branching [1]. While inhibitors of glycolysis are proposed to inhibit tumor cell growth through the blockade of a preferential source of energy fuels in cancer cells and tumor stroma cells [3,4,5,6], little is known about the contribution of the inhibition of glycosylation on tumor metabolism. Other bioenergetic and biosynthetic fuels than glucose are taken up into cells by a variety of transporters that are potentially glycosylated. SLC1A5 ( known as ASCT2) is the most described glutamine transporter in cancer cells. Part of the explanation for the prominent role of ASCT2 in cancer comes from the coupling of this transporter with a neutral amino acids antiporter [19]. Most glutamine transporters including ASCT2 have been reported to be glycosylated but whether inhibition of glucose uptake and glycolysis influences the capacity of leukemia cells to utilize glutamine through alterations of the glycosylating process, is unknown

Methods

Results

Conclusion