A mouse model of the zebrafish meltdown mutant

描述（由申请人提供）：在这项资助提案中，我们概述了一项计划，根据我们最近在斑马鱼中的发现，在小鼠中模拟人类癌细胞入侵。我们已经证明，平滑肌肌球蛋白重链（myh 11）基因突变导致斑马鱼融化（mlt）突变幼虫后肠囊状扩张。Mlt（Myh 11）蛋白，这是限制平滑肌细胞，非自主激活上皮细胞表达的人类proinvasion基因直系同源物在肠道中的斑马鱼mlt幼虫。因此，mlt突变体可用于模拟人癌细胞侵袭。在这里，我们建议获得转基因小鼠，其中mlt点突变被工程改造成小鼠Myh 11基因座。在哺乳动物模型系统中的mlt肠缺陷的表型学将使我们能够解决关于上皮结构和癌细胞侵袭的问题，这些问题在使用斑马鱼时是不可行的。首先，mlt小鼠可用于获得用于微阵列实验和器官培养测定的基质细胞系。这些实验可能导致鉴定编码指导人类癌细胞侵袭的保守基质信号的基因。第二，mlt突变体可以与其他模拟人类息肉综合征的小鼠突变体交配。这些双突变体将有助于确定Myh 11的改变是否会促进良性肠息肉的侵袭性转化。第三，mlt基因敲入的小鼠可以经受已知引起肠癌形成的化学诱变方案。这些实验将使我们能够测试myh 11突变是否会增强肿瘤的生长和/或转移。因此，mlt突变小鼠的衍生将使实验能够帮助确定人MYH 11在癌细胞侵袭中的作用。这项资助提案中描述的工作将有助于确定癌细胞侵袭和转移的分子机制，并可能导致预防癌症进展的新治疗方法。

DESCRIPTION (provided by applicant): In this grant proposal, we outline a plan to model human cancer cell invasion in the mouse based on our recent discoveries in zebrafish. We have shown that mutation of the smooth muscle myosin heavy chain (myh11) gene leads to cystic expansion of the posterior intestine in zebrafish meltdown (mlt) mutant larvae. The Mlt (Myh11) protein, which is restricted to smooth muscle cells, non-autonomously activates epithelial expression of human proinvasion gene orthologs in the intestine of zebrafish mlt larvae. Thus, mlt mutants may be used to model human cancer cell invasion. Here, we propose to derive transgenic mice in which the mlt point mutation is engineered into the mouse Myh11 locus. Phenocopy of mlt intestinal defects in a mammalian model system will allow us to address questions concerning epithelial architecture and cancer cell invasion that are not feasible using zebrafish. First, the mlt mice may be used to obtain stromal cell lines for use in microarray experiments and organ culture assays. These experiments may lead to the identification of genes encoding conserved stromal signals that direct human cancer cell invasion. Second, the mlt mutants may be mated to other mouse mutants that model human polyposis syndromes. These double mutants will help determine if alteration of Myh11 promotes invasive transformation of benign intestinal polyps. Third, the mlt knock-in mice may be subjected to chemical mutagenesis protocols known to cause formation of intestinal cancers. These experiments will allow us to test whether the myh11 mutation enhances tumor growth and or metastasis. Thus, derivation of mlt mutant mice will enable experiments that may help define the role of human MYH11 in cancer cell invasion. The work described in this grant proposal will help define the molecular mechanisms of cancer cell invasion and metastasis and may lead to new treatments to prevent cancer progression.