Transcription profiling by array of wild type and Pcsk5 Vcc mutant mouse embryos

We have identified an ENU-induced recessive mouse mutation (Vcc) with a pleiotropic phenotype overlapping including cardiac, tracheo-esophageal, anorectal, axial skeletal antero-posterior patterning defects, limb malformations, presacral mass, renal and palatal agenesis, and pulmonary hypoplasia. It results from a C470R mutation in the proprotein convertase PCSK5 (PC5/6, SPC6) that fails to complement a null allele, which also has a similar phenotype. The mutation ablates a predicted disulfide bond in the P domain, and results in reduced secretion, abnormal cellular localisation, and loss of proprotein convertase activity. We analysed whole genome gene expression in a mouse line carrying ENU-generated point mutation in Pcsk5 gene on a mixed background : 25% C57BL6/J on C3H/HeH (mutation originally introduced into C57BL6/J). We show that GDF11, a regulator of Hox expression, anteroposterior patterning and nephrogenesis, is a PCSK5 target, and that Pcsk5 mutation results in abnormal expression of several paralogous caudal Hox genes (Hoxa, Hoxc, Hoxd), and Mnx1 (Hlxb9) that are necessary for caudal embryo development. Our data establishes novel and pleiotropic functions for Pcsk5 in mammalian development and the coordinated expression of caudal Hox genes; and identifies GDF11 as a novel cleavage target for PCSK5. We propose that Pcsk5, at least in part via GDF11, coordinately regulates the expression of caudal Hox paralogs, to control antero-posterior patterning, nephrogenesis, and skeletal and anorectal development.我们发现了一种由N-乙基-N-亚硝基脲（ENU）诱导的隐性小鼠突变体（Vcc），其具有多效性表型，涵盖心脏、气管食管、肛门直肠、轴骨骼前后轴模式建成缺陷、肢体畸形、骶前肿块、肾与腭发育不全以及肺发育不全。该表型源于前蛋白转化酶PCSK5（PC5/6、SPC6）中发生的C470R突变，该突变无法互补空等位基因，而空等位基因本身亦呈现相似表型。该突变消除了P结构域中一处预测的二硫键，导致蛋白分泌水平降低、细胞定位异常，并使前蛋白转化酶活性丧失。我们对携带Pcsk5基因ENU诱导点突变的小鼠品系开展了全基因组基因表达分析，该品系具有混合遗传背景：以C3H/HeH为遗传背景，导入25%的C57BL/6J基因组（该突变最初被引入C57BL/6J小鼠中）。研究证实，调控同源异型框（Hox）基因表达、前后轴模式建成以及肾发生的生长分化因子11（GDF11）是PCSK5的靶标；Pcsk5突变会导致多个尾部Hox基因旁系同源基因（Hoxa、Hoxc、Hoxd）以及胚胎尾部发育必需的Mnx1（Hlxb9）的表达异常。本研究明确了Pcsk5在哺乳动物发育及尾部Hox基因协同表达中的全新多效功能，并证实GDF11是PCSK5的新型切割靶标。我们提出，Pcsk5至少可部分通过GDF11协同调控尾部Hox基因旁系同源物的表达，进而控制前后轴模式建成、肾发生以及骨骼与肛门直肠的发育。