Mammalian Developmental Genetics And Animal Models Of Hu

哺乳动物分子遗传学部分（SMMG）专注于胚胎发育的分子遗传学。转录调控是发育调控的一个关键因素，涉及一个精心制作的顺式调控靶基因序列库，以及多种调控基因表达的因子的合作和物理相互作用。存在于许多细胞环境中的核心元件被认为与细胞或组织特异性因子形成复合物，以建立基因表达的阳性和阴性控制，并在发育中的胚胎中实现细胞特化和组织身份。我们研究这个过程的分子遗传学。我们的大部分工作涉及在小鼠发育过程中的同源异型盒基因（称为Lhx基因）的LIM类成员的功能评价。在这些实验的过程中，我们确定了两种新的蛋白质类，分别由Ldb和Ssdp基因家族编码。这些辅因子与Lhx编码的LIM同源结构域因子和其他转录调节因子的相互作用对胚胎发育至关重要。我们研究的其他主题是Wnt抑制剂Dkk家族的成员，该家族在发育中的中枢神经系统中形成Lhx基因的作用和模式形成之间的联系。 密切相关的Lhx 1和Lhx 5基因是我们正在进行的工作的主题。这两个基因在发育中的中枢神经系统的许多区域中显著表达，包括脊髓和小脑。早期的研究表明，Lhx 1无效胚胎缺乏前头部结构，Lhx 5无效突变体在海马发育中受损。lhx 1基因缺失的胚胎在发育的早期阶段就死亡了，这就排除了对可能的大脑缺陷进行彻底分析的可能性。我们还担心Lhx 1和Lhx 5之间可能存在功能冗余。因此，我们产生了有条件的Lhx 1突变体以及Lhx 1/5双突变体，以便能够更密切地研究这两个基因在大脑发育中的单独或联合作用。分析的重点是这些突变体中发育中的小脑。缺乏Lhx 1或Lhx 5的胚胎的小脑表型并不显著。然而，Lhx 1/5双突变体的检查显示，原始小脑的浦肯野细胞基本上是缺失的，如钙结合蛋白标记物的缺乏所定义的。我们的实验表明，Lhx 1和Lhx 5是必不可少的钙结合蛋白阳性浦肯野细胞的产生，并在这个功能方面有冗余。 我们目前工作的另一个方面涉及Lhx 2在脑垂体发育中的功能。我们以前的研究已经确定了这个基因在大脑和眼睛发育以及造血中的作用。由于在视杯形成之前，眼原基的发育停滞，因此胚胎是无眼的。此外，前脑细胞增殖不足导致新皮质发育不全和海马原基发育不全。Lhx 2缺陷突变体在子宫内死亡，可能是由于细胞非自主性缺陷导致的严重贫血。这里我们描述了一个额外的表型。Lhx 2的缺失损害脑垂体后叶的形成。我们观察到过度增殖和缺乏适当的分化的前体细胞在这个组织。Lhx 2功能的丧失还导致垂体前叶和中叶的紊乱，可能继发于垂体后叶形成的缺陷。我们早期的工作已经确定Lhx 3和Lhx 4是垂体发育的重要调节因子。目前的工作表明，Lhx 2在这一过程中发挥了重要作用。 Wnt抑制剂Dkk家族的成员在发育中的胚胎中广泛表达。我们以前报道过Dkk 1，Lhx和头部诱导过程中的其他转录活性的介质。严重的喙部缺损是Dkk 1功能性消融的结果。我们最近的工作涉及Dkk 2基因敲除小鼠的表型。这些突变体的特征在于角膜细胞更新的严重缺陷，表明Wnt途径调节对于维持眼表完整性是必不可少的。角膜是一种非常有序的透明结构，其上皮细胞每隔三或四周被干细胞的衍生物取代，干细胞的衍生物位于角膜缘利姆布斯，角膜缘是角膜和邻接的结膜上皮细胞之间的过渡区。年轻成年Dkk 2无效小鼠的角膜是不透明的，并且含有通常在皮肤和结膜中发现的细胞，包括毛囊、皮脂腺和杯状细胞。Wnt信号在突变的利姆布斯区域上调。因此，Dkk 2已被鉴定为调节角膜上皮更新的Wnt抑制剂。

The Section on Mammalian Molecular Genetics (SMMG) is focused on the molecular genetics of embryonic development. Transcriptional control is a key element of developmental regulation, involving an elaborate repertoire of cis-regulatory target gene sequences as well as the cooperation and physical interaction of multiple factors that regulate gene expression. Core elements, present in many cell contexts, are thought to form complexes with cell- or tissue-specific factors to establish positive and negative control of gene expression and to bring about cell specification and tissue identity in the developing embryo. We study the molecular genetics of this process. Much of our work deals with the functional evaluation of members of the LIM class of homeobox genes (termed Lhx genes) during mouse development. During the course of these experiments, we identified two novel classes of proteins, encoded by the Ldb and Ssdp gene families, respectively. These are co-factors whose interaction with the Lhx-encoded LIM-homeodomain factors and with other transcriptional regulators is essential for embryonic development. Additional topics of our studies are members of the Dkk family of Wnt inhibitors that form a link between the action of Lhx genes and pattern formation in the developing central nervous system. The closely related Lhx1 and Lhx5 genes are topics of our ongoing work. The two genes are prominently expressed in many regions of the developing central nervous system, including the spinal cord and the cerebellum. Earlier work had shown that Lhx1 null embryos lack anterior head structures and that Lhx5 null mutants are impaired in hippocampal development. Lhx1 null embryos die at an early stage of development, precluding a thorough analysis of possible brain defects. We were also concerned that functional redundancies may exist between Lhx1 and Lhx5. We therefore generated conditional Lhx1 mutants as well as Lhx1/5 double mutants to be able to examine more closely the individual or combined roles of both genes in brain development. The analysis is focused on the developing cerebellum in these mutants. The cerebellar phenotype of embryos that lack either Lhx1 or Lhx5 is not remarkable. However, the examination of Lhx1/5 double mutants revealed that the Purkinje cells of the primordial cerebellum are largely missing, as defined by the absence of the Calbindin marker. Our experiment shows that both Lhx1 and Lhx5 are essential for the generation of Calbindin-positive Purkinje cells and that there is redundancy in this functional aspect. Another aspect of our current work concerns functions of Lhx2 in the context of the development of the pituitary gland. Our previous studies had established a role for this gene in brain and eye development and in hematopoiesis. Null embryos were anophthalmic because of a developmental arrest of the eye anlagen prior to the formation of the optic cup. In addition, deficient cell proliferation in the forebrain resulted in hypoplasia of the neocortex and aplasia of the hippocampal anlagen. The Lhx2-deficient mutants died in utero, possibly because of a cell non-autonomous defect of definitive erythropoiesis that caused severe anemia.Here we describe an additional phenotype. Deletion of Lhx2 impairs formation of the posterior lobe of the pituitary gland. We observe over-proliferation and a lack of proper differentiation of precursor cells in this tissue. Loss of Lhx2 function also leads to a disorganization of the anterior and intermediate lobes of the pituitary gland, possibly secondary to defects in the formation of the posterior pituitary. Our earlier work had identified Lhx3 and Lhx4 as essential regulators of pituitary development. The present work shows that Lhx2 plays an important role in the process as well. Members of the Dkk family of Wnt inhibitors are widely expressed in the developing embryo. We previously reported on Dkk1, a mediator of Lhx and other transcriptional activity during head induction. Severe rostral defects are the result of Dkk1 functional ablation. Our recent work has dealt with the phenotype of Dkk2 knockout mice. These mutants are characterized by a profound defect in cornea cell turnover, indicating that Wnt pathway regulation is essential for the maintenance of ocular surface integrity. The cornea is a very ordered, transparent structure whose epithelium is replaced every three or four weeks by derivatives of stem cells that reside in the limbus, a transitional zone between the corneal and the abutting conjunctival epithelium. The cornea of young adult Dkk2 null mice is opaque and contains cells normally found in skin and conjunctiva, including hair follicles, sebaceous glands and goblet cells. Wnt signals are upregulated in the mutant limbus region. Dkk2 has therefore been identified as a Wnt inhibitor that regulates the turnover of corneal epithelia.