Constructing a Conditional Kalirin Null Mouse

描述(由申请人提供)： Kalirin是DBL家族的一种大的、多结构域的Rho GDP/GTP交换因子，几乎只在神经系统中发现。根据对无脊椎动物的遗传研究和对X连锁智力低下相关基因的分析，Kalirin有望在轴突寻路和突触形成中发挥作用。我们对大鼠海马神经元、皮质神经元和交感神经元的研究表明，Kalirin在轴突的起始和生长、树突的生长以及脊椎的形成和维持中发挥着作用。Kalirin有两个针对不同Rho家族GTP酶的全球环境基金结构域和一个潜在的激活域。除了这些催化结构域外，它还具有与各种蛋白质相互作用的结构域。慢性注射可卡因可刺激伏隔核和纹状体中Kalirin的表达，但皮质Kalirin水平没有变化；电惊厥刺激后Kalirin也增加。提出的探索性研究是基于我们的观察，即反义介导的Kalirin在器官型脑片和分离的海马神经元中的表达消除首先导致线性脊椎密度的降低，然后树突树的简化。第一个目标是产生突变小鼠，这将有助于研究多个系统中的神经元发育和可塑性。为了避免胚胎或出生后早期的死亡，并允许产生组织特异性和发育调节的表达消除，将创建两个小鼠品系作为等位基因。这两项都将不受限制地提供给所有合格的调查人员。一个等位基因将产生Kalirin-7空。Kalirin-7是成人脑中最普遍的异构体，只有一个Rho环境基金结构域，以一个独特的PDZ结合基序终止。Kalirin-7出现在发育较晚的阶段，定位于树突棘。第二个等位基因将产生完全的Kalidn基因敲除。两者都将与可用的组织特异性和药物诱导的Cre重组酶小鼠杂交。目标2将研究组织特异性Kalirin条件性基因敲除的生化、组织学和发育后果。

DESCRIPTION (provided by applicant): Kalirin is a large, multidomain Rho GDP/GTP exchange factor (GEF) of the Dbl family found almost exclusively in the nervous system. Based on genetic studies in invertebrates and analysis of genes involved in X-linked mental retardation, Kalirin is expected to play a role in axonal pathfinding and synapse formation. Our studies in rat hippocampal, cortical and sympathetic neurons showed that Kalirin plays a role in axon initiation and outgrowth, dendritic growth, and spine formation and maintenance. Kalirin has two GEF domains with specificity for different Rho family GTPases and a potential kinase domain. In addition to these catalytic domains, it has interactor domains for a variety of proteins. Kalirin expression in the nucleus accumbens and striatum is stimulated by chronic administration of cocaine, with no change in cortical Kalirin levels; Kalirin is also increased following electroconvulsive shock stimulation. The exploratory studies proposed are based on our observations that antisense-mediated elimination of Kalirin expression in organotypic slices and dissociated hippocampal neurons leads first to a reduction in linear spine density followed by simplification of the dendritic tree. Aim 1 is to generate mutant mice that will facilitate studies of neuronal development and plasticity in multiple systems. In order to avoid embryonic or early postnatal lethality, and to allow generation of tissue-specific and developmentally regulated elimination of expression, two mouse lines will be created as floxed alleles. Both will be provided to all qualified investigators without restrictions. One allele will produce the Kalirin-7 null. Kalirin-7 is the most prevalent isoform in the adult brain, with a single Rho GEF domain, terminating with a unique PDZ binding motif. Kalirin-7 appears late in development and is localized to dendritic spines. The second allele will produce the total Kalidn knockout. Both will be bred with available tissue-specific and drug-inducible Cre recombinase mice. Aim 2 will examine the biochemical, histological, and developmental consequences of tissue-specific Kalirin conditional knockouts.