Cho Yoon Mi, Jang Yoon Seong, Jang Young Min, Chung Sang Mi, Kim Ho Shik, Lee Jeong Hwa, Jeong Seong Whan, Kim In Kyung, Kim Jung Jin, Kim Kwang Soo, Kwon Oh Joo
Department of Biochemistry, 2MRC for Cell Death Disease Research Center, Korea.
Exp Mol Med. 2009 Jun 30;41(6):440-52. doi: 10.3858/emm.2009.41.6.049.
When we treated rat bone marrow stromal cells (rBMSCs) with neuronal differentiation induction media, typical unfolded protein response (UPR) was observed. BIP/GRP78 protein expression was time-dependently increased, and three branches of UPR were all activated. ATF6 increased the transcription of XBP1 which was successfully spliced by IRE1. PERK was phosphorylated and it was followed by eIF2alpha phosphorylation. Transcription of two downstream targets of eIF2alpha, ATF4 and CHOP/GADD153, were transiently up-regulated with the peak level at 24 h. Immunocytochemical study showed clear coexpression of BIP and ATF4 with NeuN and Map2, respectively. UPR was also observed during the neuronal differentiation of mouse embryonic stem (mES) cells. Finally, chemical endoplasmic reticulum (ER) stress inducers, thapsigargin, tunicamycin, and brefeldin A, dose-dependently increased both mRNA and protein expressions of NF-L, and, its expression was specific to BIP-positive rBMSCs. Our results showing the induction of UPR during neuronal differentiations of rBMSCs and mES cells as well as NF-L expression by ER stress inducers strongly suggest the potential role of UPR in neuronal differentiation.
当我们用神经元分化诱导培养基处理大鼠骨髓基质细胞（rBMSCs）时，观察到典型的未折叠蛋白反应（UPR）。BIP/GRP78蛋白表达呈时间依赖性增加，并且UPR的三个分支均被激活。ATF6增加了XBP1的转录，而XBP1被IRE1成功剪接。PERK发生磷酸化，随后eIF2α发生磷酸化。eIF2α的两个下游靶点ATF4和CHOP/GADD153的转录短暂上调，在24小时达到峰值水平。免疫细胞化学研究表明，BIP和ATF4分别与NeuN和Map2明显共表达。在小鼠胚胎干细胞（mES）的神经元分化过程中也观察到了UPR。最后，化学内质网（ER）应激诱导剂毒胡萝卜素、衣霉素和布雷菲德菌素A剂量依赖性地增加了NF-L的mRNA和蛋白表达，并且其表达对BIP阳性的rBMSCs具有特异性。我们的结果表明，在rBMSCs和mES细胞的神经元分化过程中诱导了UPR，以及ER应激诱导剂诱导了NF-L表达，这强烈提示了UPR在神经元分化中的潜在作用。
