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）脑中鉴定了异常mRNA种类，这些mRNA是编码PrP样蛋白（Dopple/PrPLP）的新序列，这是一种推定的膜糖蛋白，在一级氨基酸结构上与PrpC具有23%的同一性。鉴定了该基因的表达组织和细胞，并获得了以下知识。(1)在成年野生型小鼠中，Dopple/PrPLP mRNA在睾丸和心脏中以高水平生理表达，在脑中几乎检测不到。然而，通过北方印迹法在新生野生型小鼠脑中检测到Dopple/PrPLP mRNA的瞬时表达，在出生后1周左右显示出最大表达。原位杂交结合免疫组织化学鉴定脑内皮细胞表达转录本。新生野生型小鼠Dopple/PrPLP mRNA定位于脾脏毛细血管和肠道粘膜固有层。这些发现表明Dopple/PrPLP在血管生成，特别是血脑屏障成熟中的作用。(2)在新生小鼠的睾丸中，mRNA的表达被发现在未来专门用于淋巴管的叶细胞中。另一方面，从出生后第3周开始，在生精周期的第IV-VI阶段的精原细胞开始强烈表达Dopple/PrPLP mRNA。Dopple/PrPLP可能在精子发生和血睾屏障中起重要作用。(3)北方印迹显示PrP基因敲除小鼠脑中GFAP和LM基因表达不受调节。正常小鼠PrP^C-的转基因成功地将PrP敲除小鼠从神经胶质激活中拯救出来。此外，在浦肯野细胞变性发生之前，胶质细胞活化是显著的。这些发现有力地表明，在Prp^C缺失的情况下，异位Dopple/PrPLP积极参与脑中的胶质细胞活化。

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.