Metabolic regulation of the CCN family genes by glycolysis in chondrocytes

Abstract

The CCN family consists of 6 genes in the mammalian genome and produces multifunctional proteins involved in a variety of biological processes. Recent reports indicate the profound roles of CCN2 in energy metabolism in chondrocytes, and Ccn2 deficiency is known to alter the expression of 2 other family members including Ccn3. However, almost nothing is known concerning the regulation of the CCN family genes by energy metabolism. In order to gain insight into this critical issue, we initially and comprehensively evaluated the effect of inhibition of glycolysis on the expression of all of the CCN family genes in chondrocytic cells. Upon the inhibition of a glycolytic enzyme, repression of CCN2 expression was observed, whereas CCN3 expression was conversely induced. Similar repression of CCN2 was conferred by the inhibition of aerobic ATP production, which, however, did not induce CCN3 expression. In contrast, glucose starvation significantly enhanced the expression of CCN3 in those cells. The results of a reporter gene assay using a molecular construct containing a CCN3 proximal promoter revealed a dose-dependent induction of the CCN3 promoter activity by the glycolytic inhibitor in chondrocytic cells. These results unveiled a critical role of glycolytic activity in the regulation of CCN2 and CCN3, which activity mediated the mutual regulation of these 2 major CCN family members in chondrocytes.

CCN3 (NOV) Drives Degradative Changes in Aging Articular Cartilage

“…CCN3 has antiproliferative activities in several tumor cell lines [ 34 , 35 , 36 ]. In addition, CCN3 was previously shown to be induced by glucose starvation [ 37 ]. Another report suggests that recombinant CCN3 ameliorates 1L-1β induced metalloproteinases in cultured chondrocytes by the activation of PI3K/AKT/mTOR pathway [ 38 , 39 ].…”

RFX1‐mediated CCN3 induction that may support chondrocyte survival under starved conditions

“…The inhibition of glycolysis by glucose starvation was performed essentially as previously described (Akashi et al, 2018). Briefly, after having been seeded onto six‐well culture plates, HCS‐2/8 cells were grown until they reached confluence.…”

“…The original reporter gene construct employed to evaluate the promoter activity of human CCN3 (from −1403 to −45, counted from the translation initiation site) was constructed for our previous work, using pGL3‐control (Promega) as a backbone (Akashi et al, 2018). To make a deletion mutant of the human CCN3 promoter, this reporter construct was used as a template, and three deletion mutants of the human CCN3 proximal promoter were prepared by PCR using specific primers.…”

“…As a result, we found that the expression of CCN2 was significantly reduced by a glycolysis inhibitor, while the expression of CCN3 and CCN5 was increased. No significant difference was observed in the gene expression levels of other CCN family members (Akashi et al, 2018). Moreover, CCN3 expression was also induced by glucose starvation, and it was revealed that CCN3 is under the control of glycolysis activity.…”

“…Subsequent reporter gene analysis confirmed that the transcriptional activity of a CCN3 promoter was increased by the glycolysis inhibitor in a dose‐dependent manner. Therefore, it was suggested that the induction of CCN3 expression by glycolysis inhibition is regulated at a transcriptional level via the proximal promoter (Akashi et al, 2018).…”