MIST1 regulates the pancreatic acinar cell expression of Atp2c2, the gene encoding secretory pathway calcium ATPase 2.

MIST1 is a transcription factor expressed in pancreatic acinar cells and other serous exocrine cells. Mice harboring a targeted deletion of the Mist1 gene (Mist1(-/-)) exhibit alterations in acinar regulated exocytosis and aberrant Ca(2+) signaling that are normally controlled by acinar cell Ca(2+)-ATPases. Previous studies indicated that total sarcoendoplasmic reticulum Ca(2+)-ATPases (SERCA) and plasma membrane Ca(2+)-ATPases (PMCA) remained unaffected in Mist1(-/-) acinar cultures. Therefore, we have assessed the expression of Atp2c2, the gene that encodes the secretory pathway Ca(2+)-ATPase 2 (SPCA2). We revealed a dramatic decrease in pancreatic expression of Atp2a2 mRNA and SPCA2 protein in Mist1(-/-) mice. Surprisingly, this analysis indicated that the acinar-specific Atp2c2 mRNA is a novel transcript, consisting of only the 3' end of the gene and the protein and localizes to the endoplasmic reticulum. Expression of SPCA2 was also lost in Mist1(-/-) secretory cells of the salivary glands and seminal vesicles, suggesting that Atp2c2 transcription is regulated by MIST1. Indeed, inducible MIST1 expression in Mist1(-/-) pancreatic acinar cells restored normal Atp2c2 expression, supporting a role for MIST1 in regulating the Atp2c2 gene. Based on these results, we have identified a new Atp2c2 transcript, the loss of which may be linked to the Mist1(-/-) phenotype.

Identification of a transcription factor, BHLHB8, involved in mouse seminal vesicle epithelium differentiation and function.

The seminal vesicle is a male accessory sex organ that develops from segments of the Wolffian duct adjacent to the urogenital sinus. It produces most of the seminal plasma in both humans and rodents. To date, very few transcription factors have been linked to the development and differentiation of seminal vesicles. In this study, we have examined the role of basic helix-loop-helix (BHLH) B8 transcription factor expressed at high levels in the adult seminal vesicle and during seminal gland differentiation. Immunofluorescent studies indicate that BHLHB8 is expressed within the epithelial layer of the seminal layer of the seminal vesicle following branching morphogenesis but prior to full maturation of cell morphology and function. Analysis of mice that do not express BHLHB8 (Bhlhb8(-/-)) indicates no deficiency in the initial development of the seminal vesicle. However, morphological and ultrastructural analysis indicates disruption of the epithelial cellular architecture. The seminal vesicle epithelial layer of 2-mo-old Bhlhb8(-/-) mice shows extensive cellular degeneration based on the appearance of reduced microvilli, altered granule size, and dilated endoplasmic reticulum and Golgi apparatus. The seminal vesicle epithelial cells also degenerate prematurely, as evidenced by disruption of nuclear architecture and significant accumulations of autophagic bodies. These results identify BHLHB8 as a regulator in establishing and stabilizing the secreting epithelial cells of the seminal vesicle.

Mice lacking the transcription factor Mist1 exhibit an altered stress response and increased sensitivity to caerulein-induced pancreatitis.

Several animal models have been developed to investigate the pathobiology of pancreatitis, but few studies have examined the effects that altered pancreatic gene expression have in these models. In this study, the sensitivity to secretagogue-induced pancreatitis was examined in a mouse line that has an altered acinar cell environment due to the targeted deletion of Mist1. Mist1 is an exocrine specific transcription factor important for the complete differentiation and function of pancreatic acinar cells. Mice lacking the Mist1 gene [Mist1 knockout (KO) mice] exhibit cellular disorganization and functional defects in the exocrine pancreas but no gross morphological defects. Following the induction of pancreatitis with caerulein, a CCK analog, we observed elevated serum amylase levels, necrosis, and tissue damage in Mist1 KO mice, indicating increased pancreatic damage. There was also a delay in the regeneration of acinar tissue in Mist1 KO animals. Molecular profiling revealed an altered activation of stress response genes in Mist1 KO pancreatic tissue compared with wild-type (WT) tissue following the induction of pancreatitis. In particular, Western blot analysis for activating transcription factor 3 and phosphorylated eukaryotic initiation factor 2alpha (eIF2alpha), mediators of endoplasmic reticulum (ER) stress, indicated limited activation of this pathway in Mist1 KO animals compared with WT controls. Conversely, Mist1 KO pancreatic tissue exhibits increased expression of growth arrest and DNA damage inducible 34 protein, an inhibitor of eIF2alpha phosphorylation, before and after the induction of pancreatitis. These finding suggest that activation of the ER stress pathway is a protective event in the progression of pancreatitis and highlight the Mist1 KO mouse line as an important new model for studying the molecular events that contribute to the sensitivity to pancreatic injury.

Mist1 is necessary for the establishment of granule organization in serous exocrine cells of the gastrointestinal tract.

Establishing a pool of granules at the luminal border is a key step during exocrine cell development in the pancreas and is necessary for efficient release of digestive enzymes through regulated exocytosis. Several proteins have been linked to maintaining granule organization, but it is unclear which regulatory mechanisms are necessary to establish organization. Based on temporal and spatial expression, the transcription factor Mist1 is an excellent candidate, and analysis of mice that do not express Mist1 (Mist1KO) reveal disrupted cell morphology in adult pancreatic acini. To address Mist1's role in establishing granule location, we have characterized the organization of pancreatic acini throughout development in Mist1KO mice. Using various histological approaches, we have determined that correct granule organization is never established in pancreatic acini of Mist1KO mice. Further examination indicates that this disruption in granule targeting may be the primary defect in Mist1KO mice as granule organization is affected in other serous exocrine cells that normally express Mist1. To identify a mechanistic link between granule targeting and the loss of Mist1 function, intercellular junctions and the expression of Rab3D were assessed. While both of these factors are affected in Mist1KO mice, these changes alone do not account for the disorganization observed in Mist1KO tissues. Therefore, we conclude that Mist1 is necessary for complete differentiation and maturation of serous exocrine cells through the combined regulation of several exocrine specific genes.