FUNCTIONAL REGIONS OF THE HOMEODOMAIN PROTEIN IDX-1 REQUIRED FOR TRANSACTIVATION OF THE RAT SOMATOSTATIN GENES

The insulin-, glucagon- and somatostatin-producing cells (beta, alpha and delta, respectively) in the pancreatic islets derive from a common precursor stem cell and differentiate sequentially during embryonic d evelopment. The homeodomain protein islet duodenum HOX (IDX)-1 [insuli n promoter factor (IPF)-1/somatostatin transactivating factor (STF)-1) ] is a transcription factor critically required for both the developme nt of the pancreas and the transcriptional expression of the insulin g ene. IDX-1 may also act to determine the differentiation of the common pancreatic precursor to beta, alpha and delta cells. Although IDX-1 i s detected in most adult mouse islet beta-cells and regulates insulin gene transcription, it is also found in 158 of the delta-cells and tra nsactivates the rat somatostatin gene. The roles of different domains of IDX-1 involved in the transactivation of the somatostatin gene are unclear. In this study, we have created a series of amino- and carboxy -terminal deletions, as well as point substitution mutations to deline ate functional domains within the IDX-1 protein. We find that deletion s amino-proximal to the homeodomain enhance DNA-binding to the TAAT-1 transcriptional control element within the somatostatin gene promoter. However, these amino-terminal deletions result in substantial decreas es in transactivation of a transcriptional reporter containing the TAA T-1 element. Paradoxically, coexpression of the transcriptionally inac tive, amino-terminally deleted IDX-1 mutant proteins, either with the wild-type IDX-1 or with themselves, results in a marked enhancement of transactivation of the transcriptional TAAT-1 element reporter. We pr ovide evidence that this synergistic enhancement of transactivation is mediated by protein-protein interactions among the regions of IDX-1 l ocated carboxyl-proximal to the homeodomain. Although successive delet ions into the carboxyterminal region do not alter DNA-binding, these d eletions result in a biphasic enhancement and diminution of transactiv ation. The IDX-1 homeodomain mediates sequence-specific DNA-binding be cause substitution mutations within this region abolish DNA-binding. A ll of the amino- and carboxy-terminal deletion proteins were present i n nuclear extracts of transfected cells, suggesting that nuclear local ization signals reside within the IDX-1 homeodomain. The mapping of th e functional domains of IDX-1 may facilitate understanding of IDX-1-me diated gene regulation and islet cell development.