Molecular characterization and functional study of the bHLH transcription factor MIST1
Abstract
MIST1 is a basic helix-loop-helix (bHLH) transcription factor that is expressed in serous secretory cells of multiple organs. In the pancreas, MIST1 expression is restricted to pancreatic acinar cells. Mouse Mist1 null (Mist1 KO) pancreata consist of acinar cells exhibiting dramatic differentiation defects including hyperproliferation, lack of cell polarity, failure to establish cell-cell junctions and compromised secretory function. Despite extensive characterization of Mist1KO mice, the molecular properties of the protein and the mechanisms by which it regulates pancreatic acinar cell differentiation are largely unknown. In this thesis, I dissected the MIST1 protein with the aim of identifying the position of its functional motifs that are responsible for transcription activation/repression. MIST1 and MIST1 bHLH, a MIST1 truncation that only contains the central bHLH domains of the protein, were expressed in pancreatic acinar cells of Mist1 KO mice, and the pancreata of the transgenic mice were studied to compare the activities of MIST1 and MIST1bHLH in rescuing the Mist1KO phenotype. Both MIST1 and MIST1bHLH completely restored Mist1KO pancreatic acinar cells in all aspects of the Mist1KO phenotype, including proliferation control, cellular organization and MIST1 target gene expression. These results support the contention that all functional motifs necessary for the transcriptional activity of MIST1 reside within the central bHLH region. To reveal the molecular mechanisms behind the function of MIST1 in acinar cells, Mist1KO pancreatic acinar cells were also examined for structural defects involving organization of their actin cytoskeleton. Expression profiling of the RHO family GTPases, a group of key regulators of cellular actin organization, revealed that MIST1 regulates expression of Rnd2, a RHO family GTPase 2 gene. Indeed, MIST1 binds to the Rnd2 gene promoter and represses Rnd2 gene expression in pancreatic acinar cells. Interestingly, RND2 overexpression correlates with acinar cell de-differentiation caused by various genetic or chemical reagents. Additionally, diminished Rnd2 expression in pancreatic AR42J cells results in suppressed cell proliferation and altered cell morphology. Taken together, I propose that MIST1 controls pancreatic acinar cell differentiation through its regulation of the Rnd2 gene. In normal acinar cells, MIST1 maintains low levels of RND2. Elevation of Rnd2 expression upon Mist1 deletion leads to acinar cell de-differentiation, a condition that primes the exocrine pancreas for acinar-to-ductal metaplasia (ADM) and pancreatic cancer.
Degree
Ph.D.
Advisors
Konieczny, Purdue University.
Subject Area
Molecular biology
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