A novel mechanism of transcriptional repression of p^27kip1 through Notch/HRT2 signaling in vascular smooth muscle cells

Summary

Vascular smooth muscle cell (VSMC) proliferation occurs in vascular obstructive events such as atherosclerosis and restenosis. We previously showed that Notch receptors are induced in smooth muscle cells during vascular remodeling. Our goal was to determine the mechanisms employed by Notch signaling to regulate proliferation. Activation of Notch1 and Notch4 induced the VSMC-selective target genes HRT1 and HRT2, promoted cell cycle transit in smooth muscle cells, and led to loss of density-dependent growth inhibition. This was associated with a reduction in levels of the cyclin-dependent kinase inhibitor (cdk) p27kip1.Over-expression of p27 kip1 resulted in a dose-dependent rescue of the Notch-induced phenotype and exit from the cell cycle. In addition, HRT2 expression was sufficient to promote S-phase entry, and we demonstrate that HRT2 interacts directly with the p27 kip1 promoter to repress transcription. Transcriptional repression occurred within the ∼774bp minimal p27kip1 promoter region and mutational analysis demonstrated that repression is largely dependent on a conserved class-C domain. Our data show that Notch signaling acts to promote a proliferative phenotype in VSMC by modulation of the G1/S-phase checkpoint. In addition, we define a novel mechanism by which the Notch effector, HRT2, interacts directly with the class-C domain of the p27kip1 promoter, repressing its expression. These studies identify a novel transcriptional target of HRT2, and show that Notch effectors directly control cell cycle regulatory components. We suggest that this mechanism is relevant to hyperproliferative states in VSMC seen during vascular remodeling and repair.

Keywords

Notch signaling - smooth muscle - HRT2 - p27kip1 - proliferation

• References

• 1 Karsan A. The role of notch in modeling and maintaining the vasculature. Can J Physiol Pharmacol 2005; 83: 14-23.
• 2 Joutel A, Tournier-Lasserve E. Notch signalling pathway and human diseases. Semin Cell Dev Biol 1998; 09: 619-25.
• 3 Garg V, Muth AN, Ransom JF. et al. Mutations in NOTCH1 cause aortic valve disease. Nature 2005; 437: 270-4.
• 4 Iso T, Kedes L, Hamamori Y. HES and HERP families: multiple effectors of the Notch signaling pathway. J Cell Physiol 2003; 194: 237-55.
• 5 Lindner V, Booth C, Prudovsky I. et al. Members of the Jagged/Notch gene families are expressed in injured arteries and regulate cell phenotype via alterations in cell matrix and cell-cell interaction. Am J Pathol 2001; 159: 875-83.
• 6 Domenga V, Fardoux P, Lacombe P. et al. Notch3 is required for arterial identity and maturation of vascular smooth muscle cells. Genes Dev 2004; 18: 2730-5.
• 7 Wang W, Prince CZ, Mou Y. et al. Notch3 signaling in vascular smooth muscle cells induces c-FLIP expression via ERK/MAPK activation. Resistance to Fas ligand-induced apoptosis. J Biol Chem 2002; 277: 21723-9.
• 8 Wang W, Campos AH, Prince CZ. et al. Coordinate Notch3-hairy-related transcription factor pathway regulation in response to arterial injury. Mediator role of platelet-derived growth factor and ERK. J Biol Chem 2002; 277: 23165-71.
• 9 Morrow D, Scheller A, Birney YA. et al. Notchmediated CBF-1/RBP-J{kappa-dependent regulation of human vascular smooth muscle cell phenotype in vitro. Am J Physiol Cell Physiol 2005; 289: C1188-96.
• 10 Morrow D, Sweeney C, Birney YA. et al. Cyclic strain inhibits Notch receptor signaling in vascular smooth muscle cells in vitro . Circ Res 2005; 96: 567-75.
• 11 Sweeney C, Morrow D, Birney YA. et al. Notch 1 and 3 receptor signaling modulates vascular smooth muscle cell growth, apoptosis, and migration via a CBF-1/RBP-Jk dependent pathway. Faseb J 2004; 18: 1421-3.
• 12 Proweller A, Pear WS, Parmacek MS. Notch signaling represses myocardin-induced smooth muscle cell differentiation. J Biol Chem 2005; 280: 8994-9004.
• 13 Murata K, Hattori M, Hirai N. et al. Hes1 Directly Controls Cell Proliferation through the Transcriptional Repression of p27Kip1. Mol Cell Biol 2005; 25: 4262-71.
• 14 Abderrahmani A, Niederhauser G, Lenain V. et al. The hairy and enhancer of split1 is a negative regulator of the repressor element silencer transcription factor. FEBS Lett 2005; 579: 6199-204.
• 15 Kim HK, Siu G. The notch pathway intermediate HES-1 silences CD4 gene expression. Mol Cell Biol 1998; 18: 7166-75.
• 16 Fujimori K, Fujitani Y, Kadoyama K. et al. Regulation of lipocalin-type prostaglandinD synthase gene expression by Hes-1 through E-box and interleukin-1 beta via two NF-kappa B elements in rat leptomeningeal cells. J Biol Chem 2003; 278: 6018-26.
• 17 Yan B, Raben N, Plotz P. The human acid alpha-glucosidase gene is a novel target of the Notch-1/Hes-1 signaling pathway. J Biol Chem 2002; 277: 29760-4.
• 18 Fischer A, Klattig J, Kneitz B. et al. Hey basic helixloop-helix transcription factors are repressors of GATA4 and GATA6 and restrict expression of the GATA target gene ANF in fetal hearts. Mol Cell Biol 2005; 25: 8960-70.
• 19 Firulli AB, Han D, Kelly-Roloff L. et al. A comparative molecular analysis of four rat smooth muscle cell lines. In Vitro Cell Dev Biol Anim 1998; 34: 217-26.
• 20 Rothman A, Kulik TJ, Taubman MB. et al. Development and characterization of a cloned rat pulmonary arterial smooth muscle cell line that maintains differentiated properties through multiple subcultures. Circulation 1992; 86: 1977-86.
• 21 Aoki K, Barker C, Danthinne X. et al. Efficient generation of recombinant adenoviral vectors by Crelox recombination in vitro . Mol Med 1999; 05: 224-31.
• 22 Small D, Kovalenko D, Kacer D. et al. Soluble Jagged 1 represses the function of its transmembrane form to induce the formation of the Src-dependent chord-like phenotype. J Biol Chem 2001; 276: 32022-30.
• 23 Small D, Kovalenko D, Soldi R. et al. Notch activation suppresses fibroblast growth factor-dependent cellular transformation. J Biol Chem 2003; 278: 16405-13.
• 24 Krah DL. A simplified multiwell plate assay for the measurement of hepatitis A virus infectivity. Biologicals 1991; 19: 223-7.
• 25 Minami S, Ohtani-Fujita N, Igata E. et al. Molecular cloning and characterization of the human p27Kip1 gene promoter. FEBS Lett 1997; 411: 1-6.
• 26 Campos AH, Wang W, Pollman MJ. et al. Determinants of Notch-3 receptor expression and signaling in vascular smooth muscle cells: implications in cellcycle regulation. Circ Res 2002; 91: 999-1006.
• 27 Wang W, Prince CZ, Hu X. et al. HRT1 modulates vascular smooth muscle cell proliferation and apoptosis. Biochem Biophys Res Commun 2003; 308: 596-601.
• 28 Ronchini C, Capobianco AJ. Induction of cyclin D1 transcription and CDK2 activity by Notch(ic): implication for cell cycle disruption in transformation by Notch(ic). Mol Cell Biol 2001; 21: 5925-34.
• 29 Carlson TR, Yan Y, Wu X. et al. Endothelial expression of constitutively active Notch4 elicits reversible arteriovenous malformations in adult mice. Proc Natl Acad Sci USA 2005; 102: 9884-9.
• 30 Krebs LT, Shutter JR, Tanigaki K. et al. Haploinsufficient lethality and formation of arteriovenous malformations in Notch pathway mutants. Genes Dev 2004; 18: 2469-73.
• 31 Duarte A, Hirashima M, Benedito R. et al. Dosagesensitive requirement for mouse Dll4 in artery development. Genes Dev 2004; 18: 2474-8.
• 32 Xue Y, Gao X, Lindsell CE. et al. Embryonic lethality and vascular defects in mice lacking the Notch ligand Jagged1. Hum Mol Genet 1999; 08: 723-30.
• 33 Lawson ND, Scheer N, Pham VN. et al. Notch signaling is required for arterial-venous differentiation during embryonic vascular development. Development 2001; 128: 3675-83.
• 34 Joutel A, Tournier-Lasserve E. Notch signalling pathway and human diseases. Semin Cell Dev Biol 1998; 09: 619-25.
• 35 Ruchoux MM, Domenga V, Brulin P. et al. Transgenic mice expressing mutant Notch3 develop vascular alterations characteristic of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy. Am J Pathol 2003; 162: 329-42.
• 36 Tanner FC, Boehm M, Akyurek LM. et al. Differential effects of the cyclin-dependent kinase inhibitors p27(Kip1), p21(Cip1), and p16(Ink4) on vascular smooth muscle cell proliferation. Circulation 2000; 101: 2022-5.
• 37 Pagano M, Tam SW, Theodoras AM. et al. Role of the ubiquitin-proteasome pathway in regulating abundance of the cyclin-dependent kinase inhibitor p27. Science 1995; 269: 682-5.
• 38 Carrano AC, Eytan E, Hershko A. et al. SKP2 is required for ubiquitin-mediated degradation of the CDK inhibitor p27. Nat Cell Biol 1999; 01: 193-9.
• 39 Ito E, Iwahashi Y, Yanagisawa Y. et al. Two short sequences have positive effects on the human p27Kip1 gene transcription. Gene 1999; 228: 93-100.
• 40 Inoue T, Kamiyama J, Sakai T. Sp1 and NF-Y synergistically mediate the effect of vitamin D(3) in the p27(Kip1) gene promoter that lacks vitamin D response elements. J Biol Chem 1999; 274: 32309-17.
• 41 Sarmento LM, Huang H, Limon A. et al. Notch1 modulates timing of G1-S progression by inducing SKP2 transcription and p27 Kip1 degradation. J Exp Med 2005; 202: 157-68.
• 42 Iso T, Chung G, Hamamori Y. et al. HERP1 is a cell type-specific primary target of Notch. J Biol Chem 2002; 277: 6598-607.
• 43 Shimizu K, Chiba S, Saito T. et al. Functional diversity among Notch1, Notch2, and Notch3 receptors. Biochem Biophys Res Commun 2002; 291: 775-9.
• 44 Rangarajan A, Talora C, Okuyama R. et al. Notch signaling is a direct determinant of keratinocyte growth arrest and entry into differentiation. Embo J 2001; 20: 3427-36.
• 45 Doi H, Iso T, Yamazaki M. et al. HERP1 inhibits myocardin-induced vascular smooth muscle cell differentiation by interfering with SRF binding to CArG box. Arterioscler Thromb Vasc Biol 2005; 25: 2328-34.
• 46 Noseda M, Fu Y, Niessen K. et al. Smooth Muscle alpha-actin is a direct target of Notch/CSL. Circ Res 2006; 98: 1468-70.
• 47 Noseda M, McLean G, Niessen K. et al. Notch activation results in phenotypic and functional changes consistent with endothelial-to-mesenchymal transformation. Circ Res 2004; 94: 910-7.