Functional characterization of the bHLH protein MIST1 during pancreatic development

John Michael Rukstalis, Purdue University

Abstract

Mist1 is a tissue specific member of the basic Helix-Loop-Helix (bHLH) class of transcription factors that is expressed in multiple exocrine tissues throughout the body including the pancreas, prostate, and salivary glands. In an attempt to elucidate the role of Mist1 in the development of these tissue lineages, the Mist1 gene was inactivated in the mouse through homologous recombination. Mice lacking Mist1 (Mist1 KO) are viable and phenotypically normal upon gross examination. Fine histological examination of the pancreas reveals a disruption in apical-basal polarity, alterations in the expression of proteins involved in cell-cell contact, and improper exocytosis of secretory granules. Examination of gene expression patterns in the Mist1 KO reveals changes consistent with pancreatic damage and a human clinical condition known as chronic pancreatitis. As these mice age, large tubular lesions develop within the pancreas that are seen in multiple models of pancreatic injury and disease. Though many genes with altered expression have been identified in the Mist1KO mice, the specific gene targets of this transcription factor remain unknown. In an attempt to identify the primary genetic defects that occur during the development of the Mist1KO phenotype, an Affymetrix gene array analysis was performed on pancreatic RNA samples from post natal day 1 wild-type and Mist1KO mice. The expression of one hundred and eighty-two transcripts were identified as being significantly different, with forty of the genes exhibiting a two-fold or greater change in expression levels. One identified transcript that was absent in the Mist1KO sample was the gap junction gene Connexin 32 (Cx32). Detailed analysis of Cx32 gene expression reveals that it is co-expressed with Mist1 in all exocrine organs, and is simultaneously lost in these organs upon deletion of the Mist1 gene. In vitro analysis of the Cx32 promoter demonstrates that Mist1 can positively regulate Cx32 expression in pancreatic acinar cell lines. From this data, we conclude that Cx32 is one of the primary gene targets of Mist1, and in combination with other primary target genes contributes to the Mist1KO phenotype.

Degree

Ph.D.

Advisors

Konieczny, Purdue University.

Subject Area

Molecular biology

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