Pancreatic and Duodenal Homeobox Protein 1 (Pdx-1) Maintains Endoplasmic Reticulum Calcium Levels Through Transcriptional Regulation of Sarco-endoplasmic Reticulum Calcium ATPase 2b (SERCA2b) in the Islet {beta} Cell [Molecular Bases of Disease]

September 30th, 2014 by Johnson, J. S., Kono, T., Tong, X., Yamamoto, W. R., Zarain-Herzberg, A., Merrins, M. J., Satin, L. S., Gilon, P., Evans-Molina, C.

While the Pancreatic duodenal homeobox 1 (Pdx-1) transcription factor is known to play an indispensable role in β cell development and secretory function, recent data also implicate Pdx-1 in the maintenance of endoplasmic reticulum (ER) health. The sarco endoplasmic reticulum Ca2+ ATPase 2b (SERCA2b) pump maintains a steep Ca2+ gradient between the cytosol and ER lumen. In models of diabetes, our data demonstrated loss of β cell Pdx-1 that occurs in parallel with altered SERCA2b expression, while in silico analysis of the SERCA2b promoter revealed multiple putative Pdx-1 binding sites. We hypothesized that Pdx-1 loss under inflammatory and diabetic conditions leads to decreased SERCA2b levels and activity with concomitant alterations in ER health. To test this, siRNA-mediated knockdown of Pdx-1 was performed in INS-1 cells. Results revealed reduced SERCA2b expression and decreased ER Ca2+, which was measured using fluorescence lifetime imaging microscopy. Co-transfection of human Pdx-1 with a reporter fused to the human SERCA2 promoter increased luciferase activity 3-4 fold relative to an empty vector control, and direct binding of Pdx-1 to the proximal SERCA2 promoter was confirmed by chromatin immunoprecipitation. To determine whether restoration of SERCA2b could rescue ER stress induced by Pdx-1 loss, Pdx1+/- mice were fed a high fat diet. Isolated islets demonstrated an increased spliced-to-total Xbp1 ratio, while SERCA2b overexpression reduced the Xbp1 ratio to that of wild-type controls. Together, these results identify SERCA2b as a novel transcriptional target of Pdx-1 and define a role for altered ER Ca2+ regulation in Pdx-1 deficient states.
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