Achaete-scute-like-3 (ASCL3) Regulates Vascular Muscle Cell Proliferation

描述（由申请人提供）：在发育过程中，许多涉及细胞命运决定和控制细胞增殖、分化、凋亡和迁移的级联反应是由碱性螺旋-环-螺旋类（bHLH）的转录因子（TF）调节的，包括从无脊椎动物到人类高度保守的achaeet - scsce bHLH家族。尽管哺乳动物ASCL1已成为神经生物学广泛研究的对象，但关于该家族其他成员ASCL2至5，特别是普遍表达的ASCL3的作用的信息却很少。我们的初步数据显示ASCL3、ASCL2和ASCL1在血管平滑肌细胞（VSMC）中表达。关注ASCL3，我们发现其表达在血管紧张素II刺激增殖时增加，PDGF-BB在较小程度上增加，而在Notch1激活或BMP2治疗抑制生长的情况下减少。此外，ASCL3的稳定过表达导致体外VSMC增殖增加，并且ASCL3在体内血管病变后的新生内膜中明显上调。这些数据表明，ASCL3积极促进VSMC的增殖，并在病理性血管重构的发展中发挥潜在作用，将其定义为“血管修复”因子。这些观察结果导致了我们的中心假设，即ASCL3是一种调节VSMC增殖的促生长bHLH转录因子。

DESCRIPTION (provided by applicant): Numerous cascades involved in cell fate determination during development and governing cell proliferation, differentiation, apoptosis and migration are regulated by transcription factors (TF) in the basic-helix-loop-helix class (bHLH), including those in the achaete-scute family of bHLH, which has been highly conserved from invertebrates to humans. Although the mammalian ASCL1 has been the object of extensive studies in neurobiology, yet very little information is available on the role of the other members of the family, ASCL2 to 5, particularly the ubiquitously expressed ASCL3. Our preliminary data show that ASCL3, ASCL2 and ASCL1 are expressed in vascular smooth muscle cells (VSMC). Focusing on ASCL3, we found that its expression is increased upon stimulation of proliferation with angiotensin II, and to a lesser extent PDGF-BB, while being reduced in the context of growth inhibition by Notch1 activation or BMP2 treatment. Furthermore, stable over-expression of ASCL3 leads to increased proliferation of VSMC in vitro and ASCL3 is clearly up-regulated in neointima upon vascular lesion in vivo. These data suggest that ASCL3 contributes actively to proliferation in VSMC with a potential role in development of pathological vascular remodeling, defining it as a "vasculopatic" factor. These observations led to our central hypothesis that ASCL3 is a growth-promoting bHLH transcription factor regulating VSMC proliferation. To address the implications and predictions of this hypothesis we propose to undertake: Specific aim 1: Characterization of ASCL3 expression profiles in relation to VSMC proliferation through (1.1) analysis of ASCL3 expression in response to different stimuli in vitro, focusing on factors of relevance in vascular pathologies to correlate it to the (1.2) analysis of ASCLS expression in vascular remodeling in vivo, and Specific aim 2: Analysis of the effects of ASCL3 on proliferation of VSMC in vitro through characterization of the (2.1) effects of gain- and loss-of-function of ASCLS on VSMC proliferation and, naturally derived, the (2.2) identification of ASCL3 target genes regulating VSMC proliferation. These specific aims will be addressed using immunochemistry and quantitative analysis of expression in vivo and in vitro in combination with growth factors of relevance in vascular remodeling as well as in the context of gain- and loss-of function through genetic and biochemical approaches. Development of this proposal will define ASCL3 as a novel bHLH TF with an active role in vascular proliferation and remodeling. This work is bound to transcend the field of vascular biology by shedding light on the general roles of this ubiquitous TF in cellular proliferation at large, with a potential to impact other fields where deregulated cell growth is at the roots of the involved biological processes as it is the case for cancer or developmental biology.