Experimental control of pancreatic development and maintenance

Abstract

To investigate the role of the HOX-like homeoprotein PDX1 in the formation and maintenance of the pancreas, we have genetically engineered mice so that the only source of PDX1 is a transgene that can be controlled by the application of tetracycline or its analogue doxycycline. In these mice the coding region for the tetracyclineregulated transactivator (tTA off) has replaced the coding region of the endogenous Pdx1 gene to ensure correct temporal and spatial expression of the regulatable transactivator. In the absence of doxycycline, tTA off activates the transcription of a bicistronic transgene encoding PDX1 and an enhanced green fluorescent protein reporter, which acts as a visual marker of transgene expression in living cells. Expression of the transgene-encoded PDX1 rescues the Pdx1-null phenotype; the pancreata of these mice develop and function normally. The rescue is conditional; doxycycline-mediated repression of the transgenic Pdx1 throughout gestation recapitulates the Pdx1 null phenotype. Moreover, application of doxycycline at mid-pancreogenesis blocks further development. Adult animals of the rescue genotype that were treated with doxycycline for 3 weeks shut off Pdx1 expression, decreased insulin production, and lost the ability to maintain glucose homeostasis. These results demonstrate the feasibility of controlling the formation of an organ during embryogenesis in utero and the maintenance of the mature organ through the experimental manipulation of a key developmental regulator.

Direct regulation of intestinal fate by Notch

“…To minimize the impact of genetic background effects, all experiments involving conditional transgene activation were conducted by using two tetO-NotchIC stud males. The other mouse strains used have been described: Pdx1-tTA (13), Fabpl Ϫ596 to ϩ21 Cre (14), Z͞AP (15), Pdx1-Cre (16), and RosaNotchIC (17).…”

“…To gain control of Notch activity during embryogenesis, we used an allele of Pdx1 in which the tTA replaces the endogenous Pdx1 coding sequence (13). The Notch pathway was activated by mating Pdx1-tTA mice to tetO-NotchIC responder mice.…”

Pancreas organogenesis: From bud to plexus to gland

“…Using a tetracycline-regulated system, the deletion of Pdx1 function at E11.5 and E12.5 produced a cystic pancreas with very limited or no mature acinar formation, and diminished expression of the critical acinar regulator Ptf1a in such conditional mutants might be the direct cause of the phenotype. Therefore, maintaining the low level of Pdx1 expression in the acinar cells is required for their formation and differentiation (Holland et al, 2002;Hale et al, 2005). In acinar cells, Pdx1 forms a trimeric complex with Pbx1 and Meis2, which controls the nature of the transcriptional activity of PDX1 in acinar versus endocrine cells (Swift et al, 1998).…”

A feat of metabolic proportions: Pdx1 orchestrates islet development and function in the maintenance of glucose homeostasis

“…The role of Pdx1 in the development of the embryonic pancreas beyond the elaboration of the initial buds has been addressed in elegant studies of R. MacDonald and colleagues. Using a mouse model in which the expression of Pdx1 can be selectively repressed by administration of doxycycline, they studied the effect of Pdx1 deletion at various time points after the formation of the initial pancreatic buds [51]. Deletion of Pdx1 at E12.5 led to the formation of a pancreas nearly devoid of acinar cells and islets; the precursor pancreatic epithelium was observed to grow and branch out, producing a truncated ductal tree with immature duct-like cells.…”