Ruchon A F, Marcinkiewicz M, Siegfried G, Tenenhouse H S, DesGroseillers L, Crine P, Boileau G
Département de Biochimie, Institut de Recherches Cliniques de Montréal et Département de Médecine, Université de Montréal, Montréal, Canada.
J Histochem Cytochem. 1998 Apr;46(4):459-68. doi: 10.1177/002215549804600405.
Mutations in PEX, a phosphate-regulating gene with homology to endopeptidase on the X chromosome, were recently identified in patients with X-linked hypophosphatemia (XLH), an inherited disorder of phosphate homeostasis characterized by growth retardation and rachitic and osteomalacic bone disease. To understand the mechanism by which loss of PEX function elicits the mutant phenotype, a study of its mRNA localization and ontogenesis was undertaken. Using the reverse transcriptase-nested polymerase chain reaction (RT-nested PCR) with polyA+ RNA purified from mouse testis, a 337-bp Pex cDNA fragment was generated and cloned in the pCRII plasmid. The cDNA was used to generate sense and anti-sense Pex riboprobes for in situ hybridization (ISH) and Northern analysis. To survey a large number of different tissues, sagittal sections of embryos and newborn mice were examined. ISH showed the presence of Pex mRNA in osteoblasts and odontoblasts. Pex gene expression was detectable on Day 15 of embryonic development, which coincides with the beginning of intercellular matrix deposition in bones. Finally, Northern analysis of total RNA from calvariae and teeth of 3-day-old and adult mice showed that the abundance of the 7-kb Pex transcript is decreased in adult bones and in nongrowing teeth. The present study demonstrates that Pex mRNA is expressed in bones and teeth and suggests that this putative endopeptidase plays an important role in the development of these tissues.
与X染色体上的内肽酶具有同源性的磷酸盐调节基因PEX发生的突变，最近在X连锁低磷血症（XLH）患者中被发现。XLH是一种遗传性磷酸盐稳态紊乱疾病，其特征为生长发育迟缓以及佝偻病和骨软化症。为了了解PEX功能丧失引发突变表型的机制，我们对其mRNA定位和个体发生进行了研究。使用从小鼠睾丸中纯化的polyA + RNA进行逆转录巢式聚合酶链反应（RT - 巢式PCR），生成了一个337bp的Pex cDNA片段，并将其克隆到pCRII质粒中。该cDNA用于生成正义和反义Pex核糖探针，用于原位杂交（ISH）和Northern分析。为了检测大量不同组织，我们检查了胚胎和新生小鼠的矢状切片。ISH显示成骨细胞和成牙本质细胞中存在Pex mRNA。在胚胎发育的第15天可检测到Pex基因表达，这与骨骼中细胞间基质沉积的开始时间一致。最后，对3日龄和成年小鼠颅骨和牙齿的总RNA进行Northern分析表明，7kb Pex转录本的丰度在成年骨骼和非生长牙齿中降低。本研究表明Pex mRNA在骨骼和牙齿中表达，并提示这种假定的内肽酶在这些组织的发育中起重要作用。
