Abstract
LEFTY2 (endometrial bleeding associated factor; EBAF or LEFTYA), a cytokine released shortly before menstrual bleeding, is a negative regulator of cell proliferation and tumour growth. LEFTY2 down-regulates Na+/H+ exchanger activity with subsequent inhibition of glycolytic flux and lactate production in endometrial cancer cells. Glucose can be utilized not only for glycolysis but also for glycogen formation. Both glycolysis and glycogen formation require cellular glucose uptake which could be accomplished by the Na+ coupled glucose transporter-1 (SGLT1; SLC5A1). The present study therefore explored whether LEFTY2 modifies endometrial SGLT1 expression and activity as well as glycogen formation. Ishikawa and HEC1a cells were exposed to LEFTY2, SGLT1 and glycogen synthase (GYS1) transcript levels determined by qRT-PCR. SGLT1, GYS1 and phospho-GYS1 protein abundance was quantified by western blotting, cellular glucose uptake from 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino)-2-Deoxyglucose (2-NBDG) uptake, and cellular glycogen content utilizing an enzymatic assay and subsequent colorimetry. As a result, a 48-hour treatment with LEFTY2 significantly increased SGLT1 and GYS1 transcript levels as well as SGLT1 and GYS1 protein abundance in both Ishikawa and HEC1a cells. 2-NBDG uptake and cellular glycogen content were upregulated significantly in Ishikawa (type 1) but not in type 2 endometrial HEC1a cells, although there was a tendency of increased 2-NBDG uptake. Further, none of the effects were seen in human benign endometrial cells (HESCs). Interestingly, in both Ishikawa and HEC1a cells, a co-treatment with TGF-β reduced SGLT1, GYS and phospho-GYS protein levels, and thus reduced glycogen levels and again HEC1a cells had no significant change. In conclusion, LEFTY2 up-regulates expression and activity of the Na+ coupled glucose transporter SGLT1 and glycogen synthase GYS1 in a cell line specific manner. We further show the treatment with LEFTY2 fosters cellular glucose uptake and glycogen formation and TGF-β can negate this effect in endometrial cancer cells.
Highlights
LEFTY2 is a member of the transforming growth factor beta (TGF-β) superfamily
We further show the treatment with LEFTY2 fosters cellular glucose uptake and glycogen formation and TGF-β can negate this effect in endometrial cancer cells
The present study addressed the effect of LEFTY2 on the Na+ coupled glucose transporter SGLT1 and glucose utilization in Ishikawa and HEC1a cells
Summary
LEFTY2 (endometrial bleeding associated factor; EBAF or LEFTYA) is a member of the transforming growth factor beta (TGF-β) superfamily. Unlike other TGF-β family members, LEFTY2 does not function via receptor-mediated SMADdependent signaling, but rather by antagonizing the signaling of TGF- β and Nodal [2]. Since the normal function of the TGF-β signaling pathway is suppression of cellular proliferation and transformation, it could be proposed that the action of LEFTY2 could be a potential oncoprotein by counteracting TGF-β-mediated signaling. LEFTY2 is highly enriched in embryonic stem cells and participates in the regulation of ‘stemness’ and embryonic differentiation [6,7,8,9]. This expression has been shown to re-appear in cancers, such as breast and melanoma [10]
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