{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,27]],"date-time":"2026-09-27T05:03:36Z","timestamp":1790485416130,"version":"4.1.0"},"reference-count":36,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2021,2,18]],"date-time":"2021-02-18T00:00:00Z","timestamp":1613606400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100006034","name":"University of Southern California","doi-asserted-by":"publisher","award":["N\/A"],"award-info":[{"award-number":["N\/A"]}],"id":[{"id":"10.13039\/100006034","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Leydig cells contain significant amounts of constitutively produced steroidogenic acute regulatory protein (STAR; STARD1). Hormone-induced STAR plays an essential role in inducing the transfer of cholesterol into the mitochondria for hormone-dependent steroidogenesis. STAR acts at the outer mitochondrial membrane, where it interacts with a protein complex, which includes the translocator protein (TSPO). Mutations in STAR cause lipoid congenital adrenal hyperplasia (lipoid CAH), a disorder characterized by severe defects in adrenal and gonadal steroid production; in Leydig cells, the defects are seen mainly after the onset of hormone-dependent androgen formation. The function of constitutive STAR in Leydig cells is unknown. We generated STAR knockout (KO) MA-10 mouse tumor Leydig cells and showed that STAR KO cells failed to form progesterone in response to dibutyryl-cAMP and to TSPO drug ligands, but not to 22(R)-hydroxycholesterol, which is a membrane-permeable intermediate of the CYP11A1 reaction. Electron microscopy of STAR KO cells revealed that the number and size of lipid droplets were similar to those in wild-type (WT) MA-10 cells. However, the density of lipid droplets in STAR KO cells was drastically different than that seen in WT cells. We isolated the lipid droplets and analyzed their content by liquid chromatography\u2013mass spectrometry. There was a significant increase in cholesteryl ester and phosphatidylcholine content in STAR KO cell lipid droplets, but the most abundant increase was in the amount of diacylglycerol (DAG); DAG 38:1 was the predominantly affected species. Lastly, we identified genes involved in DAG signaling and lipid metabolism which were differentially expressed between WT MA-10 and STAR KO cells. These results suggest that constitutive STAR in Leydig cells is involved in DAG accumulation in lipid droplets, in addition to cholesterol transport. The former event may affect cell functions mediated by DAG signaling.<\/jats:p>","DOI":"10.3390\/ijms22042021","type":"journal-article","created":{"date-parts":[[2021,2,18]],"date-time":"2021-02-18T20:56:21Z","timestamp":1613681781000},"page":"2021","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":80,"title":["Role of Constitutive STAR in Leydig Cells"],"prefix":"10.3390","volume":"22","author":[{"given":"Melanie","family":"Galano","sequence":"first","affiliation":[{"name":"Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, CA 90089, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuchang","family":"Li","sequence":"additional","affiliation":[{"name":"Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, CA 90089, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8053-8374","authenticated-orcid":false,"given":"Lu","family":"Li","sequence":"additional","affiliation":[{"name":"Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, CA 90089, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chantal","family":"Sottas","sequence":"additional","affiliation":[{"name":"Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, CA 90089, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1183-8568","authenticated-orcid":false,"given":"Vassilios","family":"Papadopoulos","sequence":"additional","affiliation":[{"name":"Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, CA 90089, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2021,2,18]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"113","DOI":"10.1016\/0303-7207(91)90210-J","article-title":"Acute action of luteinizing hormone on mouse Leydig cells: Accumulation of mitochondrial phosphoproteins and stimulation of testosterone synthesis","volume":"81","author":"Epstein","year":"1991","journal-title":"Mol. Cell Endocrinol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1095\/biolreprod59.2.263","article-title":"Leydig cell protein synthesis and steroidogenesis in response to acute stimulation by luteinizing hormone in rats","volume":"59","author":"Luo","year":"1998","journal-title":"Biol. Reprod."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1868","DOI":"10.1210\/me.2012-1159","article-title":"Identification of a dynamic mitochondrial protein complex driving cholesterol import, trafficking, and metabolism to steroid hormones","volume":"26","author":"Rone","year":"2012","journal-title":"Mol. Endocrinol."},{"key":"ref_4","first-page":"S233","article-title":"Steroidogenic Acute Regulatory Protein: Structure, Functioning, and Regulation","volume":"84","author":"Tugaeva","year":"2019","journal-title":"Biochemistry"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"663","DOI":"10.1016\/j.bbalip.2007.02.012","article-title":"Steroidogenic acute regulatory protein (StAR), a novel mitochondrial cholesterol transporter","volume":"1771","author":"Miller","year":"2007","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"11540","DOI":"10.1073\/pnas.94.21.11540","article-title":"Targeted disruption of the mouse gene encoding steroidogenic acute regulatory protein provides insights into congenital lipoid adrenal hyperplasia","volume":"94","author":"Caron","year":"1997","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1870","DOI":"10.1056\/NEJM199612193352503","article-title":"The pathophysiology and genetics of congenital lipoid adrenal hyperplasia","volume":"335","author":"Bose","year":"1996","journal-title":"N. Engl. J. Med."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"4139","DOI":"10.1016\/j.bmcl.2012.04.058","article-title":"Identification of small-molecule inhibitors of the steroidogenic acute regulatory protein (STARD1) by structure-based design","volume":"22","author":"Akula","year":"2012","journal-title":"Bioorg. Med. Chem. Lett."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"10223","DOI":"10.1074\/jbc.M611221200","article-title":"Cholesterol binding does not predict activity of the steroidogenic acute regulatory protein, StAR","volume":"282","author":"Baker","year":"2007","journal-title":"J. Biol. Chem."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"90","DOI":"10.1016\/j.mce.2015.03.014","article-title":"Translocator protein-mediated pharmacology of cholesterol transport and steroidogenesis","volume":"408","author":"Papadopoulos","year":"2015","journal-title":"Mol. Cell Endocrinol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"719","DOI":"10.1111\/andr.12733","article-title":"Cholesterol accumulation, lipid droplet formation, and steroid production in Leydig cells: Role of translocator protein (18-kDa)","volume":"8","author":"Chung","year":"2020","journal-title":"Andrology"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1546","DOI":"10.1016\/j.celrep.2016.10.037","article-title":"SEIPIN Regulates Lipid Droplet Expansion and Adipocyte Development by Modulating the Activity of Glycerol-3-phosphate Acyltransferase","volume":"17","author":"Pagac","year":"2016","journal-title":"Cell Rep."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1186\/s12944-017-0473-y","article-title":"Perilipins: A diversity of intracellular lipid droplet proteins","volume":"16","author":"Itabe","year":"2017","journal-title":"Lipids Health Dis."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"843","DOI":"10.1016\/j.biocel.2007.04.008","article-title":"Deleted in liver cancer-1 (DLC-1): A tumor suppressor not just for liver","volume":"40","author":"Liao","year":"2008","journal-title":"Int. J. Biochem. Cell Biol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"399","DOI":"10.1007\/s00210-007-0131-4","article-title":"Phospholipase D signaling: Orchestration by PIP2 and small GTPases","volume":"374","author":"Schmidt","year":"2007","journal-title":"Naunyn Schmiedebergs Arch. Pharmacol."},{"key":"ref_16","first-page":"2641","article-title":"Lipid phosphate phosphatase 3 participates in transport carrier formation and protein trafficking in the early secretory pathway","volume":"126","author":"Johannes","year":"2013","journal-title":"J. Cell Sci."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"28314","DOI":"10.1016\/S0021-9258(18)46930-X","article-title":"The purification, cloning, and expression of a novel luteinizing hormone-induced mitochondrial protein in MA-10 mouse Leydig tumor cells. Characterization of the steroidogenic acute regulatory protein (StAR)","volume":"269","author":"Clark","year":"1994","journal-title":"J. Biol. Chem."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1828","DOI":"10.1126\/science.7892608","article-title":"Role of steroidogenic acute regulatory protein in adrenal and gonadal steroidogenesis","volume":"267","author":"Lin","year":"1995","journal-title":"Science"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"4778","DOI":"10.1073\/pnas.92.11.4778","article-title":"Human steroidogenic acute regulatory protein: Functional activity in COS-1 cells, tissue-specific expression, and mapping of the structural gene to 8p11.2 and a pseudogene to chromosome 13","volume":"92","author":"Sugawara","year":"1995","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"2299","DOI":"10.1093\/hmg\/4.12.2299","article-title":"T-->A transversion 11 bp from a splice acceptor site in the human gene for steroidogenic acute regulatory protein causes congenital lipoid adrenal hyperplasia","volume":"4","author":"Tee","year":"1995","journal-title":"Hum. Mol. Genet."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"289","DOI":"10.3389\/fendo.2017.00289","article-title":"Monitoring of Dual CRISPR\/Cas9-Mediated Steroidogenic Acute Regulatory Protein Gene Deletion and Cholesterol Accumulation Using High-Resolution Fluorescence","volume":"8","author":"Lee","year":"2017","journal-title":"Front. Endocrinol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"6785","DOI":"10.1073\/pnas.83.18.6785","article-title":"Rapid formation of diacylglycerol from phosphatidylcholine: A pathway for generation of a second messenger","volume":"83","author":"Besterman","year":"1986","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"978","DOI":"10.1016\/j.bbadis.2017.01.026","article-title":"Steroidogenic acute regulatory protein (StAR) overexpression attenuates HFD-induced hepatic steatosis and insulin resistance","volume":"1863","author":"Qiu","year":"2017","journal-title":"Biochim Biophys Acta Mol. Basis Dis."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"244","DOI":"10.1095\/biolreprod.104.037721","article-title":"Involvement of protein kinase C and cyclic adenosine 3\u2019,5\u2019-monophosphate-dependent kinase in steroidogenic acute regulatory protein expression and steroid biosynthesis in Leydig cells","volume":"73","author":"Jo","year":"2005","journal-title":"Biol. Reprod."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"33","DOI":"10.1016\/0006-291X(84)90376-0","article-title":"Steroidogenic effect of exogenous phospholipase C on bovine adrenal fasciculata cells","volume":"123","author":"Hadjian","year":"1984","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"312","DOI":"10.1016\/0014-5793(85)80393-8","article-title":"Effect of phorbol ester and phospholipase C on LH-stimulated steroidogenesis in purified rat Leydig cells","volume":"188","author":"Papadopoulos","year":"1985","journal-title":"FEBS Lett."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1353","DOI":"10.1016\/0300-9084(88)90006-5","article-title":"Stimulation of progesterone production by phorbol-12-myristate-13-acetate in MA-10 Leydig tumor cells","volume":"70","author":"Chaudhary","year":"1988","journal-title":"Biochimie"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"10819","DOI":"10.1016\/S0021-9258(19)85156-6","article-title":"The nature of protein kinase C activation by physically defined phospholipid vesicles and diacylglycerols","volume":"260","author":"Boni","year":"1985","journal-title":"J. Biol. Chem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"279","DOI":"10.1016\/j.cmet.2011.12.018","article-title":"FAT SIGNALS--lipases and lipolysis in lipid metabolism and signaling","volume":"15","author":"Zechner","year":"2012","journal-title":"Cell Metab."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"14253","DOI":"10.1016\/S0021-9258(17)43852-X","article-title":"Positional specificity of hormone-sensitive lipase from rat adipose tissue","volume":"258","author":"Fredrikson","year":"1983","journal-title":"J. Biol. Chem."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"484","DOI":"10.1007\/s12020-016-1043-1","article-title":"SF-1 deficiency causes lipid accumulation in Leydig cells via suppression of STAR and CYP11A1","volume":"54","author":"Hatano","year":"2016","journal-title":"Endocrine"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"15837","DOI":"10.1074\/jbc.M601097200","article-title":"Perilipin promotes hormone-sensitive lipase-mediated adipocyte lipolysis via phosphorylation-dependent and -independent mechanisms","volume":"281","author":"Miyoshi","year":"2006","journal-title":"J. Biol. Chem."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Federico, L., Yang, L., Brandon, J., Panchatcharam, M., Ren, H., Mueller, P., and Smyth, S.S. (2018). Lipid phosphate phosphatase 3 regulates adipocyte sphingolipid synthesis, but not developmental adipogenesis or diet-induced obesity in mice. PLoS ONE, 13.","DOI":"10.1371\/journal.pone.0198063"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"286","DOI":"10.1002\/j.1460-2075.1995.tb07002.x","article-title":"A dual functional signal mediator showing RhoGAP and phospholipase C-delta stimulating activities","volume":"14","author":"Homma","year":"1995","journal-title":"Embo J."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1016\/j.febslet.2005.11.073","article-title":"Mitochondrial targeting of growth suppressor protein DLC2 through the START domain","volume":"580","author":"Ng","year":"2006","journal-title":"FEBS Lett."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"88","DOI":"10.1210\/endo-108-1-88","article-title":"Characterization of Several Clonal Lines of Cultured Leydig Tumor Cells: Gonadotropin Receptors and Steroidogenic Responses","volume":"108","author":"Ascoli","year":"1981","journal-title":"Endocrinology"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/22\/4\/2021\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T05:25:48Z","timestamp":1760160348000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/22\/4\/2021"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,2,18]]},"references-count":36,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2021,2]]}},"alternative-id":["ijms22042021"],"URL":"https:\/\/doi.org\/10.3390\/ijms22042021","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,2,18]]}}}