Miller Camille, Cai Yafei, Patton Tadd, Graves Sarai Hacker, Li Honglin, Sabbatini Maria Eugenia
Department of Biological Sciences, Augusta University, Augusta, Georgia.
Department of Biochemistry and Molecular Biology, Medical College of Georgia, Cancer Center, Augusta University, Augusta, Georgia; and.
Am J Physiol Gastrointest Liver Physiol. 2017 Mar 1;312(3):G314-G326. doi: 10.1152/ajpgi.00176.2016. Epub 2017 Jan 19.
Alcoholism causes an imbalance of endoplasmic reticulum (ER) homeostasis in pancreatic acini. In those cells, the ER is involved in the synthesis and folding of pancreatic enzymes. Ubiquitin-fold modifier 1 (Ufm1) is part of a novel ubiquitin-like modification system involved in maintaining ER homeostasis. Among the components of the Ufm1 system, Regulator of C53 and DDRGK1 (RCAD) has recently been identified as a Ufm1-specific E3 ligase that promotes ufmylation of DDRGK1, an RCAD-interacting protein. We determined the importance of RCAD in the proper synthesis and secretion of pancreatic enzymes using mice with genetically deleted RCAD. The pancreas of RCAD-deficient mice was of normal size and histology. Using quantitative PCR and Western blotting, we found that amylase was upregulated in pancreas organs from RCAD-knockout (KO) mice. Constitutive amylase secretion was much higher in isolated pancreatic acini from RCAD KO mice, whereas CCK-stimulated amylase secretion was disturbed. RCAD deficiency caused a downregulation in expression of ER chaperone BiP, which affected ER homeostasis and activated both apoptosis and trypsin. We also found that both RCAD and DDRGK1 transcript levels were upregulated in pancreatic acini from alcohol-preferring rats. Elevated expression of RCAD and DDRGK1 was associated with increased ER stress and UPR activation. Because of the lack of BiP expression, caspase 3 and trypsin activation we enhanced in RCAD-deficient pancreatic acini upon treatment with ethanol and CCK. In conclusion, the RCAD/BiP pathway is required for proper synthesis and secretion of pancreatic enzymes. In alcoholism, increased levels of components of the Ufm1 system could prevent the deleterious effects of alcohol in the pancreas by regulating BiP levels. RCAD/BiP pathway is required for the proper synthesis and secretion of amylase from pancreatic acini, as well as for the maintenance of the ER homeostasis. In alcoholism, the exocrine pancreas could increase the levels of components of the Ufm1 system to protect itself from alcohol's deleterious effects by regulating the expression of ER chaperone BiP.
酒精中毒会导致胰腺腺泡内质网（ER）稳态失衡。在这些细胞中，内质网参与胰腺酶的合成与折叠。泛素折叠修饰因子1（Ufm1）是一种新型泛素样修饰系统的一部分，该系统参与维持内质网稳态。在Ufm1系统的组成成分中，C53和DDRGK1调节因子（RCAD）最近被鉴定为一种Ufm1特异性E3连接酶，可促进与RCAD相互作用的蛋白DDRGK1的泛素化修饰。我们利用基因敲除RCAD的小鼠，确定了RCAD在胰腺酶正常合成与分泌中的重要性。RCAD缺陷小鼠的胰腺大小和组织学正常。通过定量PCR和蛋白质印迹法，我们发现RCAD基因敲除（KO）小鼠胰腺器官中的淀粉酶上调。来自RCAD基因敲除小鼠的分离胰腺腺泡中组成型淀粉酶分泌要高得多，而胆囊收缩素（CCK）刺激的淀粉酶分泌受到干扰。RCAD缺陷导致内质网伴侣蛋白BiP的表达下调，这影响了内质网稳态，并激活了细胞凋亡和胰蛋白酶。我们还发现，在偏爱酒精的大鼠的胰腺腺泡中，RCAD和DDRGK1的转录水平均上调。RCAD和DDRGK1表达的升高与内质网应激增加和未折叠蛋白反应（UPR）激活有关。由于缺乏BiP表达，在用乙醇和CCK处理后，RCAD缺陷的胰腺腺泡中半胱天冬酶3和胰蛋白酶的激活增强。总之，RCAD/BiP途径是胰腺酶正常合成与分泌所必需的。在酒精中毒中，Ufm1系统成分水平的升高可通过调节BiP水平来预防酒精对胰腺的有害影响。RCAD/BiP途径对于胰腺腺泡中淀粉酶的正常合成与分泌以及内质网稳态的维持是必需的。在酒精中毒中，外分泌胰腺可通过调节内质网伴侣蛋白BiP的表达来提高Ufm1系统成分的水平，以保护自身免受酒精的有害影响。
