Identification of expression cells and elucidation of a function for a novel gene encoding a PrP-like protein newly separated and identified

我们在PrP基因敲除小鼠(PRNP-/-长崎)的脑中发现了异常的mRNA物种，其中mRNAs是编码PrP样蛋白的新序列，该蛋白是一种在一级氨基酸结构上与PrPC有23%同源性的膜糖蛋白。对该基因的表达组织和细胞进行了鉴定，并获得了以下认识：(1)在成年野生型小鼠中，睾丸和心脏生理性地高水平表达多普利/PrPLP基因，BAT在脑中几乎检测不到。Northern印迹法检测到新生野生型小鼠脑内有瞬时表达的多谱勒/PrPLP基因，在出生后1周左右达到高峰。原位杂交和免疫组织化学检测表明，脑血管内皮细胞表达转录本。此外，在新生野生型小鼠脾毛细血管和肠粘膜固有层均可见D/PrPLP的表达。这些结果提示，在血管生成，尤其是血脑屏障的成熟过程中，DUPLE/PrPLP起着重要的作用。(2)在新生小鼠的睾丸中，可以在未来的淋巴管专化的叶细胞中发现mRNA的表达。生后3周开始，生精周期第IV-VId的精原细胞开始强表达Depple/PrPLP基因。因此，多普利/PrPLP可能在精子发生和血-睾丸屏障中起重要作用。(3)Northern印迹显示PrP基因敲除小鼠脑内GFAP和LM基因表达异常。一种用于正常小鼠的转基因PrP^C成功地将PrP基因敲除的小鼠从胶质细胞激活中拯救出来。此外，早在浦肯野细胞变性开始之前，胶质细胞的激活就很明显。这些发现有力地表明，在缺乏PRP^C的情况下，异位的多谱勒/PrPLP积极参与了脑内神经胶质细胞的激活。

We identified aberrant mRNA species in the brain of PrP knockout mice (Prnp-/- Nagasaki), which mRNAs were the novel sequence encoding PrP-like protein (Dopple/PrPLP), a putative membrane glycoprotein with 23 % identity to PrpC in the primary amino acid structure. The expression tissues and cells for this gene were identified, and the following knowledge was acquired.(1) In adult wild-type mice, Dopple/PrPLP mRNA was physiologically expressed at a high level by testis and heart, bat was barely detectable in brain. However, transient expression of Dopple/PrPLP mRNA was detectable by Northern blotting in the brain of neonatal wild-type mice, showing maximal expression around 1 week after birth. In situ hybridization paired with immunohistochemistry Identified brain endothelial cells as expressing the transcripts. Moreover, in the neonatal wild-type mice, the Dopple/PrPLP mRNA localized in capillaries of spleen and lamina propria mucosa of gut. These findings suggested a role of Dopple/PrPLP in angiogenesis, in particular blood-brain barrier maturation.(2) In a testis of neonatal mice, the expression of mRNA was found in the leaf cells specializing in the lymph vessel in the future. On the other hand, from the 3rd week after birth, the spermatogonia at the stage IV-VI day of the spermatogenic cycle became to strongly express Dopple/PrPLP mRNA. Hence, Dopple/PrPLP may play an important role in the spermatogenesis and blood-testis barrier.(3) Northern blotting demonstrated unregulated expression of the genes for GFAP and LM in the brains of PrP knockout mice. A transgene for normal mouse PrP^C- successfully rescued PrP knockout mice from the glial activation. Moreover, the glial cell activation was notable well before the onset of the Purkinje cell degeneration. These findings strongly suggest that ectopic Dopple/PrPLP in the absence of Prp^C is actively involved in the glial-cell activation in the brain.