Silencing of solute carrier family 13 member 5 disrupts energy homeostasis and inhibits proliferation of human hepatocarcinoma cells

Abstract

The solute carrier family 13 member 5 (SLC13A5), a sodiumcoupled citrate transporter, plays a key role in importing citrate from the circulation into liver cells. Recent evidence has revealed that SLC13A5 deletion protects mice from high-fat diet-induced hepatic steatosis and that mutation of the SLC13A5 orthologues in Drosophila melanogaster and Caenorhabditis elegans promotes longevity. However, despite the emerging importance of SLC13A5 in energy homeostasis, whether perturbation of SLC13A5 affects the metabolism and malignancy of hepatocellular carcinoma is unknown. Here, we sought to determine whether SLC13A5 regulates hepatic energy homeostasis and proliferation of hepatoma cells. RNAi-mediated silencing of SLC13A5 expression in two human hepatoma cell lines, HepG2 and Huh7, profoundly suppressed cell proliferation and colony formation, and induced cell cycle arrest accompanied by increased expression of cyclin-dependent kinase inhibitor p21 and decreased expression of cyclin B1. Furthermore, such suppressive effects were also observed on the growth of HepG2 cell-derived xenografts expressing SLC13A5-shRNA in nude mice. Metabolically, knockdown of SLC13A5 in HepG2 and Huh7 cells was associated with a decrease in intracellular levels of citrate, the ratio of ATP/ADP, phospholipid content, and ATP citrate lyase expression. Moreover, both in vitro and in vivo assays demonstrated that SLC13A5 depletion promotes activation of the AMP-activated protein kinase, which was accompanied by deactivation of oncogenic mechanistic target of rapamycin signaling. Together, our findings expand the role of SLC13A5 from facilitating hepatic energy homeostasis to influencing hepatoma cell proliferation and suggest a potential role of SLC13A5 in the progression of human hepatocellular carcinoma.Metabolic reprogramming has long been regarded as a hallmark of cancer, through which cancer cells switch from oxidative phosphorylation to glycolysis for ATP production even in microenvironments with sufficient oxygen (1, 2). To accommodate the challenge of rapid proliferation, cancer cell metabolism is remodeled to increase the biosynthesis of macromolecules including nucleotides, proteins, and lipids as building blocks of new cells (3,4). Although the precise mechanisms underlying this metabolic reprogramming remain elusive, malignant cells often have perturbed metabolism that allows for the accumulation of many metabolic intermediates facilitating the production of cellular building materials (4,5). Numerous studies have demonstrated that the use of glycolysis and the tricarboxylic acid (TCA) 2 cycle intermediates for biosynthesis is a key feature of altered metabolism in cancer cells (6, 7). Among others, citrate, an intermediate metabolite located at the crossroads of glucose metabolism and energy production, is an important sensor of energy homeostasis (8).Accumulating evidence suggests that citrate is involved in both physiological and pathophysiological processes including histone acetylation, insulin secretion, inflammation, ca...

Flipping a citrate switch on liver cancer cells

“…1). Knocking down SLC13A5 in two human hepatoma cell lines caused a decrease in proliferation and cell cycle arrest at the G 1 phase, which were associated with decreased expression of two proteins involved in cell cycle control, p21 and cyclin B1 (5). Interestingly, similar effects were also observed when SLC13A5 was knocked down in an in vitro model of tumorigenicity (anchorage-dependent clonogenicity) and in an in vivo model, ectopic xenografts in mice, derived from human liver cancer cell lines.…”

“…To examine the role of the citrate transporter SLC13A5 in liver cancer, Li et al (5) used knockdown of SLC13A5 in liver cancer cell lines to monitor the impact on modulating cell growth and oncogenic signaling (Fig. 1).…”

Hepatitis B reactivation in chronic hepatitis C patients treated with interferon or pan‐oral direct‐acting antivirals

“…(2,3) In contrast, Chen et al found that baseline levels of HBV DNA were not associated with HBV reactivation during the DAA-based therapies; however, this conclusion was based on only eight cases whose baseline level of HBV DNA varied from undetectable to high (1.1 3 10 6 IU/mL). Further analysis on the incidence of HBV reactivation among patients with different baseline levels of HBV DNA may yield alternative conclusions.…”

“…(1) We were well aware of the difference in the definition of hepatitis B virus (HBV) reactivation (HBVr) across the studies. However, due to the limited data reported in the included studies, we were unable to use a standardized definition, HBV replication 2 log increase from baseline levels, or a new appearance of HBV DNA to a level of 100 IU/mL (2) to adjust the pooled results. However, we have shown in the discussion that the pooled rate of HBVr (12.2%) could be higher in patients treated with DAAs if we standardize the definition.…”

Is mIndy a mediator of energy metabolism reprogramming in hepatocellular carcinoma induced by interleukin‐6/signal transducer and activator of transcription 3 signaling?

“…The hypothesis was supported by a recent study that knocking down of mIndy inhibited human HCC cell proliferation by disrupting energy homeostasis in vitro. (5) Taken together, the oncogenic role of IL-6/STAT3 signing was not only mediated by its the well-known downstream antiapoptotic proteins and proliferationrelated cyclins, but factors that contributed to the remodeling of energy metabolism, such as mIndy, might be also needed to adapt the rapid cell proliferation in the development of HCC. In addition, although the contribution of mIndy in HCC cell growth was confirmed in vitro, whether Indy is required for HCC growth in vivo will be an interesting topic for further studies.…”