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LXR activation causes G1/S arrest through inhibiting SKP2 expression in MIN6 pancreatic beta cells

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Abstract

Liver X receptors (LXRs) are nuclear hormone receptors with central roles in lipid homeostasis. We previously showed that LXR activation induced aberrant lipid metabolism and G1 cell cycle arrest in pancreatic beta cells. In this study, we aimed to identify the molecular target of LXR causing G1 arrest. LXR activation was induced by its agonist, T0901317. A series of luciferase reporters of truncated Skp2 promoter were analyzed in MIN6 cells. mRNA and protein levels of SKP2 and P27 were detected. Flow cytometry assay was used to determine the cell cycle distribution. MTT assay was used to evaluate cell viability. LXR activation increased cell distribution in G1 phase and lipid accumulation. Since dominant-negative Srebp1c could clear the deposited lipid rather than recover the G1 arrest, we identified S-phase kinase-associated protein 2 (Skp2) as a potential target gene of LXR. In deed, LXR activation significantly inhibited Skp2 gene expression and protein amount. We also observed that the luciferase activity of Skp2 promoter was suppressed by T0901317 and the potential LXR regulatory site was narrowed down to a region of nt −289 to −38. Silencing Lxrα and Lxrβ rescued SKP2 protein level and recovered the cellular growth repressed by LXR activation. Moreover, SKP2 overabundance reduced P27 protein level by promoting its degradation, consequently overcame the G1 arrest caused by T0901317. Our findings demonstrate that transrepressing Skp2 expression by LXR activation resulted in defective SKP2-mediated P27 degradation and inhibitory cell growth in beta cells.

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References

  1. L. Bouwens, I. Rooman, Regulation of pancreatic beta-cell mass. Physiol. Rev. 85(4), 1255–1270 (2005). doi:10.1152/physrev.00025.2004

    Article  CAS  PubMed  Google Scholar 

  2. S. Georgia, A. Bhushan, Beta cell replication is the primary mechanism for maintaining postnatal beta cell mass. J. Clin. Investig. 114(7), 963–968 (2004). doi:10.1172/jci22098

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. A.M. Ackermann, M. Gannon, Molecular regulation of pancreatic beta-cell mass development, maintenance, and expansion. J. Mol. Endocrinol. 38(1–2), 193–206 (2007). doi:10.1677/jme-06-0053

    Article  CAS  PubMed  Google Scholar 

  4. C. Jacovetti, A. Abderrahmani, G. Parnaud, J.C. Jonas, M.L. Peyot, M. Cornu, R. Laybutt, E. Meugnier, S. Rome, B. Thorens, M. Prentki, D. Bosco, R. Regazzi, MicroRNAs contribute to compensatory beta cell expansion during pregnancy and obesity. J. Clin. Investig. 122(10), 3541–3551 (2012). doi:10.1172/jci64151

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. M. Prentki, C.J. Nolan, Islet beta cell failure in type 2 diabetes. J. Clin. Investig. 116(7), 1802–1812 (2006). doi:10.1172/JCI29103

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. G.C. Weir, D.R. Laybutt, H. Kaneto, S. Bonner-Weir, A. Sharma, Beta-cell adaptation and decompensation during the progression of diabetes. Diabetes 50(Suppl 1), S154–159 (2001)

    Article  CAS  PubMed  Google Scholar 

  7. G.C. Weir, S. Bonner-Weir, Five stages of evolving beta-cell dysfunction during progression to diabetes. Diabetes 53(Suppl 3), S16–21 (2004)

    Article  CAS  PubMed  Google Scholar 

  8. P. Yesil, E. Lammert, Islet dynamics: a glimpse at beta cell proliferation. Histol. Histopathol. 23(7), 883–895 (2008)

    CAS  PubMed  Google Scholar 

  9. L. Zhang, C. Wang, F-box protein Skp2: a novel transcriptional target of E2F. Oncogene 25(18), 2615–2627 (2006). doi:10.1038/sj.onc.1209286

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. L. Zhong, S. Georgia, S.I. Tschen, K. Nakayama, K. Nakayama, A. Bhushan, Essential role of Skp2-mediated p27 degradation in growth and adaptive expansion of pancreatic beta cells. J. Clin. Investig. 117(10), 2869–2876 (2007). doi:10.1172/JCI32198

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Z. Zhang, J. Li, L. Yang, R. Chen, R. Yang, H. Zhang, D. Cai, H. Chen, The cytotoxic role of intermittent high glucose on apoptosis and cell viability in pancreatic beta cells. J. Diabetes Res. 2014, 712781 (2014). doi:10.1155/2014/712781

    PubMed  PubMed Central  Google Scholar 

  12. S.I. Tschen, S. Georgia, S. Dhawan, A. Bhushan, Skp2 is required for incretin hormone-mediated beta-cell proliferation. Mol. Endocrinol. (Baltimore, Md.) 25(12), 2134–2143 (2011). doi:10.1210/me.2011-1119

    Article  CAS  Google Scholar 

  13. A.C. Carrano, E. Eytan, A. Hershko, M. Pagano, SKP2 is required for ubiquitin-mediated degradation of the CDK inhibitor p27. Nat. Cell Biol. 1(4), 193–199 (1999)

    Article  CAS  PubMed  Google Scholar 

  14. L. Lu, H. Schulz, D.A. Wolf, The F-box protein SKP2 mediates androgen control of p27 stability in LNCaP human prostate cancer cells. BMC Cell Biol. 3, 22 (2002)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. R.H. Unger, Y.T. Zhou, Lipotoxicity of beta-cells in obesity and in other causes of fatty acid spillover. Diabetes 50(Suppl 1), S118–121 (2001)

    Article  CAS  PubMed  Google Scholar 

  16. N.A. van Herpen, V.B. Schrauwen-Hinderling, Lipid accumulation in non-adipose tissue and lipotoxicity. Physiol. Behav. 94(2), 231–241 (2008). doi:10.1016/j.physbeh.2007.11.049

    Article  PubMed  Google Scholar 

  17. K.R. Steffensen, J.A. Gustafsson, Putative metabolic effects of the liver X receptor (LXR). Diabetes 53(Suppl 1), S36–42 (2004)

    Article  CAS  PubMed  Google Scholar 

  18. M. Korach-Andre, A. Archer, R.P. Barros, P. Parini, J.A. Gustafsson, Both liver-X receptor (LXR) isoforms control energy expenditure by regulating brown adipose tissue activity. Proc. Natl. Acad. Sci. USA. 108(1), 403–408 (2011). doi:10.1073/pnas.1017884108

    Article  CAS  PubMed  Google Scholar 

  19. I. Gerin, V.W. Dolinsky, J.G. Shackman, R.T. Kennedy, S.H. Chiang, C.F. Burant, K.R. Steffensen, J.A. Gustafsson, O.A. MacDougald, LXRbeta is required for adipocyte growth, glucose homeostasis, and beta cell function. J. Biol. Chem. 280(24), 23024–23031 (2005). doi:10.1074/jbc.M412564200

    Article  CAS  PubMed  Google Scholar 

  20. Z.X. Meng, J. Nie, J.J. Ling, J.X. Sun, Y.X. Zhu, L. Gao, J.H. Lv, D.Y. Zhu, Y.J. Sun, X. Han, Activation of liver X receptors inhibits pancreatic islet beta cell proliferation through cell cycle arrest. Diabetologia 52(1), 125–135 (2009). doi:10.1007/s00125-008-1174-x

    Article  CAS  PubMed  Google Scholar 

  21. S. Talukdar, F.B. Hillgartner, The mechanism mediating the activation of acetyl-coenzyme A carboxylase-alpha gene transcription by the liver X receptor agonist T0-901317. J. Lipid Res. 47(11), 2451–2461 (2006). doi:10.1194/jlr.M600276-JLR200

    Article  CAS  PubMed  Google Scholar 

  22. A.M. Efanov, S. Sewing, K. Bokvist, J. Gromada, Liver X receptor activation stimulates insulin secretion via modulation of glucose and lipid metabolism in pancreatic beta-cells. Diabetes 53(Suppl 3), S75–78 (2004)

    Article  CAS  PubMed  Google Scholar 

  23. J.T. Rodgers, P. Puigserver, Receptor feasts on sugar and cholesterol. Nat. Med. 13(2), 128–129 (2007). doi:10.1038/nm0207-128

    Article  CAS  PubMed  Google Scholar 

  24. M. Korach-Andre, A. Archer, C. Gabbi, R.P. Barros, M. Pedrelli, K.R. Steffensen, A.T. Pettersson, J. Laurencikiene, P. Parini, J.A. Gustafsson, Liver X receptors regulate de novo lipogenesis in a tissue-specific manner in C57BL/6 female mice. Am. J. Physiol. Endocrinol. Metab. 301(1), E210–222 (2011). doi:10.1152/ajpendo.00541.2010

    Article  CAS  PubMed  Google Scholar 

  25. Z.X. Meng, Y. Yin, J.H. Lv, M. Sha, Y. Lin, L. Gao, Y.X. Zhu, Y.J. Sun, X. Han, Aberrant activation of liver X receptors impairs pancreatic beta cell function through upregulation of sterol regulatory element-binding protein 1c in mouse islets and rodent cell lines. Diabetologia 55(6), 1733–1744 (2012). doi:10.1007/s00125-012-2516-2

    Article  CAS  PubMed  Google Scholar 

  26. A. Takahashi, K. Motomura, T. Kato, T. Yoshikawa, Y. Nakagawa, N. Yahagi, H. Sone, H. Suzuki, H. Toyoshima, N. Yamada, H. Shimano, Transgenic mice overexpressing nuclear SREBP-1c in pancreatic beta-cells. Diabetes 54(2), 492–499 (2005)

    Article  CAS  PubMed  Google Scholar 

  27. T. Zuo, R. Liu, H. Zhang, X. Chang, Y. Liu, L. Wang, P. Zheng, Y. Liu, FOXP3 is a novel transcriptional repressor for the breast cancer oncogene SKP2. J. Clin. Investig. 117(12), 3765–3773 (2007). doi:10.1172/JCI32538

    CAS  PubMed  PubMed Central  Google Scholar 

  28. I.C. Wang, Y.J. Chen, D. Hughes, V. Petrovic, M.L. Major, H.J. Park, Y. Tan, T. Ackerson, R.H. Costa, Forkhead box M1 regulates the transcriptional network of genes essential for mitotic progression and genes encoding the SCF (Skp2-Cks1) ubiquitin ligase. Mol. Cell. Biol. 25(24), 10875–10894 (2005). doi:10.1128/MCB.25.24.10875-10894.2005

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. G. Schneider, D. Saur, J.T. Siveke, R. Fritsch, F.R. Greten, R.M. Schmid, IKKalpha controls p52/RelB at the skp2 gene promoter to regulate G1- to S-phase progression. EMBO J. 25(16), 3801–3812 (2006). doi:10.1038/sj.emboj.7601259

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. L.M. Sarmento, H. Huang, A. Limon, W. Gordon, J. Fernandes, M.J. Tavares, L. Miele, A.A. Cardoso, M. Classon, N. Carlesso, Notch1 modulates timing of G1-S progression by inducing SKP2 transcription and p27 Kip1 degradation. J. Exp. Med. 202(1), 157–168 (2005). doi:10.1084/jem.20050559

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. J. Miyazaki, K. Araki, E. Yamato, H. Ikegami, T. Asano, Y. Shibasaki, Y. Oka, K. Yamamura, Establishment of a pancreatic beta cell line that retains glucose-inducible insulin secretion: special reference to expression of glucose transporter isoforms. Endocrinology 127(1), 126–132 (1990)

    Article  CAS  PubMed  Google Scholar 

  32. R.F. Santerre, R.A. Cook, R.M. Crisel, J.D. Sharp, R.J. Schmidt, D.C. Williams, C.P. Wilson, Insulin synthesis in a clonal cell line of simian virus 40-transformed hamster pancreatic beta cells. Proc. Natl. Acad. Sci. U.S.A. 78(7), 4339–4343 (1981)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Z.X. Meng, J.X. Sun, J.J. Ling, J.H. Lv, D.Y. Zhu, Q. Chen, Y.J. Sun, X. Han, Prostaglandin E2 regulates Foxo activity via the Akt pathway: implications for pancreatic islet beta cell dysfunction. Diabetologia 49(12), 2959–2968 (2006). doi:10.1007/s00125-006-0447-5

    Article  CAS  PubMed  Google Scholar 

  34. M. Nakakuki, H. Shimano, N. Inoue, M. Tamura, T. Matsuzaka, Y. Nakagawa, N. Yahagi, H. Toyoshima, R. Sato, N. Yamada, A transcription factor of lipid synthesis, sterol regulatory element-binding protein (SREBP)-1a causes G(1) cell-cycle arrest after accumulation of cyclin-dependent kinase (cdk) inhibitors. FEBS J. 274(17), 4440–4452 (2007). doi:10.1111/j.1742-4658.2007.05973.x

    Article  CAS  PubMed  Google Scholar 

  35. T. Uchida, T. Nakamura, N. Hashimoto, T. Matsuda, K. Kotani, H. Sakaue, Y. Kido, Y. Hayashi, K.I. Nakayama, M.F. White, M. Kasuga, Deletion of Cdkn1b ameliorates hyperglycemia by maintaining compensatory hyperinsulinemia in diabetic mice. Nat. Med. 11(2), 175–182 (2005). doi:10.1038/nm1187

    Article  CAS  PubMed  Google Scholar 

  36. L. Hengst, S.I. Reed, Translational control of p27Kip1 accumulation during the cell cycle. Science 271(5257), 1861–1864 (1996)

    Article  CAS  PubMed  Google Scholar 

  37. M. Shirane, Y. Harumiya, N. Ishida, A. Hirai, C. Miyamoto, S. Hatakeyama, K. Nakayama, M. Kitagawa, Down-regulation of p27(Kip1) by two mechanisms, ubiquitin-mediated degradation and proteolytic processing. J. Biol. Chem. 274(20), 13886–13893 (1999)

    Article  CAS  PubMed  Google Scholar 

  38. N. Mitro, P.A. Mak, L. Vargas, C. Godio, E. Hampton, V. Molteni, A. Kreusch, E. Saez, The nuclear receptor LXR is a glucose sensor. Nature 445(7124), 219–223 (2007). doi:10.1038/nature05449

    Article  CAS  PubMed  Google Scholar 

  39. N. Mitro, L. Vargas, R. Romeo, A. Koder, E. Saez, T0901317 is a potent PXR ligand: implications for the biology ascribed to LXR. FEBS Lett. 581(9), 1721–1726 (2007). doi:10.1016/j.febslet.2007.03.047

    Article  CAS  PubMed  Google Scholar 

  40. C.P. Chuu, R.Y. Chen, R.A. Hiipakka, J.M. Kokontis, K.V. Warner, J. Xiang, S. Liao, The liver X receptor agonist T0901317 acts as androgen receptor antagonist in human prostate cancer cells. Biochem. Biophys. Res. Commun. 357(2), 341–346 (2007). doi:10.1016/j.bbrc.2007.03.116

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. F. Blaschke, O. Leppanen, Y. Takata, E. Caglayan, J. Liu, M.C. Fishbein, K. Kappert, K.I. Nakayama, A.R. Collins, E. Fleck, W.A. Hsueh, R.E. Law, D. Bruemmer, Liver X receptor agonists suppress vascular smooth muscle cell proliferation and inhibit neointima formation in balloon-injured rat carotid arteries. Circ. Res. 95(12), e110–123 (2004). doi:10.1161/01.RES.0000150368.56660.4f

    Article  CAS  PubMed  Google Scholar 

  42. L.L. Vedin, S.A. Lewandowski, P. Parini, J.A. Gustafsson, K.R. Steffensen, The oxysterol receptor LXR inhibits proliferation of human breast cancer cells. Carcinogenesis 30(4), 575–579 (2009). doi:10.1093/carcin/bgp029

    Article  CAS  PubMed  Google Scholar 

  43. S. Ghisletti, W. Huang, S. Ogawa, G. Pascual, M.E. Lin, T.M. Willson, M.G. Rosenfeld, C.K. Glass, Parallel SUMOylation-dependent pathways mediate gene- and signal-specific transrepression by LXRs and PPARgamma. Mol. Cell 25(1), 57–70 (2007). doi:10.1016/j.molcel.2006.11.022

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. S. Wang, J.F. Raven, A.E. Koromilas, STAT1 represses Skp2 gene transcription to promote p27Kip1 stabilization in Ras-transformed cells. Mol. Cancer Res. 8(5), 798–805 (2010). doi:10.1158/1541-7786.MCR-10-0027

    Article  CAS  PubMed  Google Scholar 

  45. J. Wu, S.W. Lee, X. Zhang, F. Han, S.Y. Kwan, X. Yuan, W.L. Yang, Y.S. Jeong, A.H. Rezaeian, Y. Gao, Y.X. Zeng, H.K. Lin, Foxo3a transcription factor is a negative regulator of Skp2 and Skp2 SCF complex. Oncogene 32(1), 78–85 (2013). doi:10.1038/onc.2012.26

    Article  CAS  PubMed  Google Scholar 

  46. Y. Zhang, Z. Gan, P. Huang, L. Zhou, T. Mao, M. Shao, X. Jiang, Y. Chen, H. Ying, M. Cao, J. Li, J. Li, W.J. Zhang, L. Yang, Y. Liu, A role for protein inhibitor of activated STAT1 (PIAS1) in lipogenic regulation through SUMOylation-independent suppression of liver X receptors. J. Biol. Chem. 287(45), 37973–37985 (2012). doi:10.1074/jbc.M112.403139

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. J.H. Lee, S.M. Park, O.S. Kim, C.S. Lee, J.H. Woo, S.J. Park, E.H. Joe, I. Jou, Differential SUMOylation of LXRalpha and LXRbeta mediates transrepression of STAT1 inflammatory signaling in IFN-gamma-stimulated brain astrocytes. Mol. Cell 35(6), 806–817 (2009). doi:10.1016/j.molcel.2009.07.021

    Article  CAS  PubMed  Google Scholar 

  48. S.B. Joseph, A. Castrillo, B.A. Laffitte, D.J. Mangelsdorf, P. Tontonoz, Reciprocal regulation of inflammation and lipid metabolism by liver X receptors. Nat. Med. 9(2), 213–219 (2003). doi:10.1038/nm820

    Article  CAS  PubMed  Google Scholar 

  49. M. Pascual-Garcia, L. Rue, T. Leon, J. Julve, J.M. Carbo, J. Matalonga, H. Auer, A. Celada, J.C. Escola-Gil, K.R. Steffensen, E. Perez-Navarro, A.F. Valledor, Reciprocal negative cross-talk between liver X receptors (LXRs) and STAT1: effects on IFN-gamma-induced inflammatory responses and LXR-dependent gene expression. J. Immunol. (Baltimore, Md.: 1950) 190(12), 6520–6532 (2013). doi:10.4049/jimmunol.1201393

    Article  CAS  Google Scholar 

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Acknowledgments

This work was supported by grants from the National Key Basic Research Program of China (973 program), (No. 2012CB524901) to X. Han, and the National Natural Science Foundation of China (Nos. 81200559 and 31101011).

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  1. Key Laboratory of Human Functional Genomics of Jiangsu Province, Department of Biochemistry and Molecular Biology, Nanjing Medical University, 140 Hanzhong Road, Nanjing, 210029, People’s Republic of China

    Yating Li, Changwen Jing, Xinyi Tang, Yuanyuan Chen, Xiao Han & Yunxia Zhu

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  1. Yating Li
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  2. Changwen Jing
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Li, Y., Jing, C., Tang, X. et al. LXR activation causes G1/S arrest through inhibiting SKP2 expression in MIN6 pancreatic beta cells. Endocrine 53, 689–700 (2016). https://doi.org/10.1007/s12020-016-0915-8

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