Genetic control of skeletal development by Pbx1

描述（申请人提供）：细胞增殖与终末分化的程度和时间的完美协调是一个遗传控制的过程，是所有组织和器官系统发育的基础。对这一严格调控过程的放松可能导致人类出生缺陷和肿瘤转化。Pbx1是一种同源结构域蛋白，可与Hox蛋白协同结合DNA以调节其DNA结合特异性，并且是外展果蝇（EXD）的同源物，其在果蝇身体计划模式中的功能已被遗传学证明。我们正在进行的努力和长期目标是利用转基因小鼠模型来评估Pbx家族的Hox辅助因子对哺乳动物模式和形态发生的贡献。在胚胎中，Pbx1 （Pbx1- 1-）的缺乏导致妊娠晚期死亡，轴骨和尾骨广泛的模式缺陷，第二鳃弓神经嵴细胞衍生的骨骼结构的同质转化，软骨细胞增殖明显减少，伴有软骨细胞早性肥大和骨过早骨化。与pbx1 - 1-不同，Pbx2 - 1-和pbx3 - 1-小鼠在模式或骨骼发育/成熟方面均未表现出明显异常。尽管如此,Pbx1-l -;pbx2 - 1突变体在子宫内死亡较早，并表现出骨骼缺陷的急剧恶化。本研究的目标是通过以下具体目标，精细地解剖同源盒基因Pbx1对图图化和骨骼发育的遗传控制：1)利用现有的Cre小鼠，通过产生基因敲除小鼠，使Pbxl在神经嵴和软骨细胞中以组织特异性的方式失活，从基因上分离Pbxl在图图化中的早期作用与其在软骨增殖、分化和软骨内成骨中的后期作用；2)利用具有明显生长缺陷的转基因间充质细胞（Pbx1- 1-），如小鼠胚胎成纤维细胞（MEFs）和Micromass间充质细胞培养，表征Pbx1在软骨细胞增殖中的作用；3)确定Pbx1与相关家族成员Pbx2在骨骼发育中的独特和重叠功能，并仅在软骨形成的遗传控制中。这些研究的完成将促进我们对Pbx1模式和骨骼发育的遗传调控的理解。从更广泛的角度来看，这项工作将揭示骨骼发育扰动的戏剧性影响，并有望影响我们对影响颅面、轴骨和尾骨发育的人类出生缺陷发病机制的理解。

DESCRIPTION (provided by applicant): Perfect coordination of the extent and timing of cellular proliferation with terminal differentiation is a genetically controlled process, fundamental for the development of all tissues and organ systems. Deregulations of this tightly regulated process can cause human birth defects and neoplastic transformation. Pbx1 is a homeodomain protein that collaboratively binds DNA with Hox proteins to modulate their DNA binding specificities and is a homolog of Drosophila extradenticle (EXD), whose function in patterning the fly body plan has been demonstrated genetically. Our ongoing efforts and long-term goals utilize genetically modified mouse models to assess the contributions of the Pbx family of Hox cofactors to mammalian patterning and morphogenesis. In embryos, the lack of Pbx1 (Pbx1-l-) results in late gestational lethality, widespread patterning defects of the axial and appendicular skeleton, homeotic transformation of second branchial arch neural crest cell-derived skeletal structures, markedly diminished chondrocyte proliferation, accompanied by precocious chondrocyte hypertrophy, and premature ossification of bone. Unlike Pbx1-l-, both Pbx2 -l-and Pbx3-l- mice do not display gross abnormalities either in patterning or in skeletal development/maturation. Nonetheless, Pbx1-l-; Pbx2-l- mutants die earlier in utero and show drastic exacerbation of the skeletal defects. The goal of this proposal is to finely dissect the genetic control of patterning and skeletal development by the homeobox gene Pbx1, through the following specific aims: 1) genetically uncouple the early roles of Pbxl in patterning from its later roles in cartilage proliferation, differentiation and endochondral ossification, through the generation of knockout mice where Pbxl is inactivated in a tissue-specific manner in neural crest and chondrocytes, by utilizing available Cre mice; 2) characterize the role of Pbx1 in chondrocyte proliferation by using genetically modified (Pbx1-l-) mesenchymal cells in culture, such as Mouse Embryonic Fibroblasts (MEFs), which show a striking growth defect, and Micromass Mesenchyme Cultures; 3) identify unique and overlapping functions of Pbx1 with the related family member Pbx2 in skeletal development and also exclusively in the genetic control of chondrogenesis. Completion of these studies will advance our understanding of the genetic regulation of patterning and skeletal development by Pbx1. Under a broader perspective, this work will shed light on the dramatic effects of the perturbations of skeletal development and hopefully impact on our understanding of the pathogenesis of human birth defects that affect the development of the craniofacial, axial and appendicular skeleton.