Fatty acid oxidation plays a

Fatty Formoterol Hemifumarate β-oxidation plays a crucial role in maintaining body energy homoeostasis mainly during catabolic states. It mainly occurs in mitochondria and involves a cyclical series of reactions that result in the shortening of fatty acids. Fatty acids comprise an efficient source of energy that results in the generation of a large quantity of ATP, and it is involved in the lipid metabolism of cancer cells [30], [31], [32], [33], [34]. In solid tumors, fatty acid β-oxidation activation links to the promotion of breast cancer cells survival triggered by loss of attachment to the matrix during metabolic stress [30]. Also, it is reported that cell survival of human LN18 glioblastoma and HeLa cells depended on fatty acid β-oxidation under conditions of complete nutrient deprivation [31]. Fatty acid β-oxidation connects with the malignant phenotype and regulates by the promyelocytic leukemia protein to active peroxisome proliferator-activated receptor signaling [32]. Recently, a study showed that SIRT6 overexpression induced increased expression of fatty acid β-oxidation–related genes such as carnitine palmitoyltransferase 1 (CPT1), hydroxyacyl-CoA dehydrogenase trifunctional multienzyme complex subunit beta (HADHB), and carnitine O-octanoyltransferase [28], yet the effect of PKCζ on fatty acid β-oxidation is unknown. It is novelty if we can reveal a novel, cooperative model executed by PKCζ and SIRT6 to maintain fatty acid β-oxidation. Here, we report that the phosphorylation of SIRT6 is significantly increased after palmitic acid (PA) treatment in variety of colon cancer cells. PKCζ can interact with SIRT6 directly and phosphorylates SIRT6 at threonine 294 residue in response to PA treatment. SIRT6 Thr294 phosporylation is required for SIRT6 enrichment on chromatin to further bind to the promoters of fatty acid β-oxidation–related genes acyl-CoA synthetase long-chain family member 1 (ACSL1), CPT1, carnitine-acylcarnitine translocase (CACT), and HADHB and then induces the expression of these genes to mediate the fatty acid β-oxidation. Altogether, our data demonstrate that PKCζ plays a major role in regulating gene expression of fatty acid β-oxidation through SIRT6 phosphorylation. These findings further our understanding of the effect of PKCζ on lipid metabolism and may guide the design of new therapeutics to regulate lipid homeostasis.
Materials and Methods
Results
Discussion The present study has identified PKCζ as a new regulator of fatty acid β-oxidation through SIRT6 phosphorylation, thus adding a new layer of knowledge to PKCζ biology. In light of our data, we present a model to hypothesize how PKCζ regulates the expression of fatty acid β-oxidation (Figure 7). In response to PA treatment, PKCζ physically interacts with SIRT6 and phosphorylates SIRT6 at Thr294 residue. The phosphorylated SIRT6 binds to the promoters of fatty acid β-oxidation–related genes to activate the expression of these genes and regulates fatty acid β-oxidation. Our data demonstrate the mechanism of PKCζ on mediating fatty acid β-oxidation through SIRT6 phosphorylation. As we know, this is the first paper to define the role of PKCζ on fatty acid β-oxidation, and it will be benefited for designing new therapeutic target in regulating lipid homeostasis. The phenomenon of SIRT6 phosphorylation was first reported by Thirumurthi et al. They found that SIRT6 was phosphorylated at Ser (338) by the kinase AKT1, which induced the interaction and ubiquitination of SIRT6 by MDM2, targeting SIRT6 for protease-dependent degradation in various breast cancer cell lines [38]. Also, the survival of breast cancer patients was positively correlated with the abundance of SIRT6 and inversely correlated with the phosphorylation of SIRT6 at Ser (338) [38]. Furthermore, two recent studies showed that SIRT6 can phosphorylate by other phosphorylases. C-Jun N-terminal kinase was reported to phosphorylate SIRT6 at serine 10 site to stimulate DNA double-strand break repair in response to oxidative stress by recruiting PARP1 to DNA breaks [39]. Also, SIRT6 phosphorylation by casein kinase 2 alpha 1 (CK2α/CSNK2A1) at Ser338 residue inhibited the proliferation of MCF7 cells, indicating that CSNK2A1-mediated phosphorylation of SIRT6 might be involved in the progression of breast carcinoma and predicted shorter survival of the diagnosed patients [40]. Similar with these studies, the semiexogenous and endogenous phosphorylation of SIRT6 was found to be dramatically enhanced after PA treatment in colon cancer cells in our study (Figure 1). To identify the phosphorylase of SIRT6 after PA treatment, several phosphorylases were detected, and we found that PKCζ can interact with SIRT6 directly in vitro and in vivo and the interaction was significantly enhanced in response to PA treatment (Figure 2). To further confirm the effect of PKCζ on SIRT6, in vitro phosphorylation assays were performed and showed that PKCζ is the phosphorylase of SIRT6 in response to PA treatment (Figure 3F). Also, threonine 294 residue was found to be the phosphorylation site of SIRT6 in vitro and in vivo (Figure 4A-C). These data indicate that PKCζ is a phosphorylase of SIRT6 and phosphorylates SIRT6 on threonine 294 residue after PA treatment.