g Glutamine consumption measurement in response to CtBP knockdown or knockdown under HG and LG conditions

g Glutamine consumption measurement in response to CtBP knockdown or knockdown under HG and LG conditions. some other branching pathways such as one carbon metabolism and pentose phosphate pathway (PPP) were also found to be important channels to convert glucose to other essential downstream molecules Toll-like receptor modulator for cancer cell growth5,6. Glutamine is mainly utilized through the glutaminolysis pathway and the research about this pathway has attracted great attention in recent years, because cancer cells were found to rely on this pathway for durable supply of carbon and nitrogen7. The crosstalk between glycolysis and glutaminolysis has been noticed a long time ago; however, how Toll-like receptor modulator these two processes influence each other ARMD5 is controversial. Previous studies indicated that the interactive activity of these two pathways is mediated by some intermediate metabolites such as pyruvate. Pyruvate is the end product of glycolysis and glutamine can also be used to produce pyruvate; however, the latter process is more complicated and needs to go through several enzymatic reactions belonging to the TCA cycle8. The other way of interaction is through serine synthesis pathway6. Glutamate provides an amine group to 3-phosphopyruvate, a product converted from glycolysis intermediate 3-phosphoglycerate, to form 3-phosphoserine, the precursor of serine. The third interactive mechanism between glycolysis and glutaminolysis correlates with synthesis of nucleotide hexosamine, a substrate for protein glycosylation, which requires the input from Toll-like receptor modulator both glucose and glutamine. Wellen et al.9 found that glycolysis is required for glutamine uptake in multiple types of mammalian cells and the mediating factor, IL3Ra, was found to be glycosylated by hexosamine that is synthesized dependent on glucose. Consistently, the conclusion of glycolysis promoting glutamine uptake has also been demonstrated in another independent study using B cell as a model. In this study, withdrawal of glucose led to almost 10 times decrease in glutamine metabolism, but the mechanism has not been elaborated10. Recent studies indicate that fast proliferating cells, in particular, the cancer cells, require the durable supply of both energy and metabolites used as building blocks. Both glucose and glutamine are consumed to fulfill this requirement11,12. Glucose, for instance, goes through the aerobic glycolysis process to accelerate the output of ATP. However, glutamine is directly transported into the TCA cycle to exaggerate the output of intermediate metabolites, especially the citrate, which can be further converted to acetyl-CoA as a building block for the synthesis of fatty acid etc. Glutaminolysis mainly occurs in mitochondria where glutamine is converted to -ketoglutarate and enters the TCA cycle. Upon DNA damage, glutaminolysis is halted temporarily to contribute to cell cycle inhibition13. One of the known mediators of DNA damage response in controlling glutaminolysis is SIRT4. SIRT4 is a mitochondrion-localized Sirtuin family protein with both deacetylation and ADP-ribosylation enzymatic activities14. SIRT4 catalyzes the ADP-ribosylation of gutamate dehydrogenase (GDH), an enzyme converting glutamate to -ketoglutarate, leading to repressed glutaminolysis14. Our previous data indicated that expression is repressed by a transcriptional co-repressor CtBP to contribute to the maintenance of pH homeostasis of breast cancer cells, which benefits cancer cells for their growth15. Here we further report that CtBP repression of expression is regulated by glycolysis activity in cancer cells and highlight a novel pathway that mediates the crosstalk between glycolysis and glutaminolysis. Results Correlated glucose and glutamine consumption in cancer cells Glucose and glutamine are two major carbon sources for cancer cells and both of them can enter the TCA Toll-like receptor modulator cycle to produce energy (Fig. ?(Fig.1a).1a). In order to investigate whether glycolysis impacts glutaminolysis, we cultured MCF-7 cells in high glucose (HG, 4.5?g/L glucose) medium and low glucose (LG, 1?g/L glucose) medium but supplied with the same initial amount of glutamine (2?mM). We did not observe obvious increased apoptosis associated with 1?g/L glucose culture condition for MCF-7 cells and MDA-MB-231 cells (data not shown). As expected, the Toll-like receptor modulator cells cultured in HG medium showed a much faster proliferation than the cells in LG medium (Fig. ?(Fig.1b).1b). To monitor the ability of glutamine consumption by each individual cell in HG and LG culture conditions, the glutamine consumption was normalized to.

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