SUMMARY Berardinelli-Seip congenital lipodystrophy 2 (BSCL2) is caused by loss-of-function mutations in SEIPIN, a protein implicated in both adipogenesis and lipid droplet expansion but whose molecular function remains obscure. Here, we identify physical and functional interactions between SEIPIN and microsomal isoforms of glycerol-3-phosphate acyltransferase (GPAT) in multiple organisms. Compared to controls, GPAT activity was elevated in SEIPIN-deficient cells and tissues and GPAT kinetic values were altered. Increased GPAT activity appears to underpin the block in adipogenesis and abnormal lipid droplet morphology associated with SEIPIN loss. Over-expression of Gpat3 blocked adipogenesis, and Gpat3 knockdown in SEIPIN-deficient preadipocytes partially restored differentiation. GPAT overexpression in yeast, preadipocytes, and fly salivary glands also formed supersized lipid droplets. Finally, pharmacological inhibition of GPAT in Seipin−/− mouse preadipocytes partially restored adipogenesis. These data identify SEIPIN as an evolutionarily conserved regulator of microsomal GPAT and suggest that GPAT inhibitors might be useful for the treatment of human BSCL2 patients.

Diacylglycerol kinase alpha (DGKα) catalyzes the conversion of diacylglycerol (DAG) to phosphatidic acid (PA). Recently, DGKα was identified as a therapeutic target in various cancers, as well as in immunotherapy. Application of small-molecule DGK inhibitors, R59022 and R59949, induces cancer cell death in vitro and in vivo. The pharmacokinetics of these compounds in mice, however, are poor. Thus, there is a need to discover additional DGK inhibitors not only to validate these enzymes as targets in oncology, but also to achieve a better understanding of their biology. In the present study, we investigate the activity of ritanserin, a compound structurally similar to R59022, against DGKα. Ritanserin, originally characterized as a serotonin (5-HT) receptor (5-HTR) antagonist, underwent clinical trials as a potential medicine for the treatment of schizophrenia and substance dependence. We document herein that ritanserin attenuates DGKα kinase activity while increasing the enzyme’s affinity for ATP in vitro. In addition, R59022 and ritanserin function as DGKα inhibitors in cultured cells and activate protein kinase C (PKC). While recognizing that ritanserin attenuates DGK activity, we also find that R59022 and R59949 are 5-HTR antagonists. In conclusion, ritanserin, R59022 and R59949 are combined pharmacological inhibitors of DGKα and 5-HTRs in vitro.

Cancer cells reprogram their metabolism to increase the synthesis of macromolecules for rapid proliferation. Compared to fatty acids, much less is known about the synthesis of phospholipids, which is essential for membrane biogenesis in cancer cells. We found that , which encodes lipin-1, a phosphatidic acid phosphatase (PAP) controlling the rate-limiting step in the phospholipid synthesis pathway, is highly up-regulated in basal-like triple-negative breast cancer (TNBC). Moreover, high expression correlates with the poor prognosis of these patients. Knockdown of increases apoptosis in basal-like TNBC cell lines, whereas it has minimal or less effect on normal human mammary gland epithelial cells (HMECs) and estrogen receptor-positive breast cancer cell lines. Fatty acid incorporation and lipidomics analyses showed that knockdown blocks phospholipid synthesis and changes membrane lipid compositions that ultimately induce the activation of 1 of the 3 branches of unfolded protein responses, the inositol-requiring enzyme-1α pathway. We also show for the first time, to our knowledge, that lipin-1 knockdown significantly inhibits tumor growth using an orthotopic xenograft breast mouse model. Our results suggest that lipin-1 is a potential target for cancer therapy.-He, J., Zhang, F., Tay, L. W. R., Boroda, S., Nian, W., Levental, K. R., Levental, I., Harris, T. E., Chang, J. T., Du, G. Lipin-1 regulation of phospholipid synthesis maintains endoplasmic reticulum homeostasis and is critical for triple-negative breast cancer cell survival.

Background: Lipin 2 is a phosphatidic acid phosphatase (PAP) responsible for DAG formation at the ER membrane during lipogenesis. Results: A combination of biochemical approaches is used to characterize lipin 2 phosphatase activity and regulation. Conclusion: The electrostatic charge of PA regulates activity, but phosphorylation does not.Significance: These findings demonstrate differential regulation of PAP activity within the lipin family.

Our findings demonstrate that a DGKα-GGTase I pathway can be targeted to combat the treatment-resistant mesenchymal cancer phenotype. Combining therapies with greater activity against each GBM subtype may represent a viable therapeutic option against GBM.