Regulation of amylase gene expression in diabetic mice is mediated by a cis-acting upstream element close to the pancreas-specific enhancer.

Abstract

We localized the c/s-acting sequences that mediate the regulation of a pancreatic amylase gene, Amy-2.2, in diabetic mice. We constructed three hybrid genes containing sequences from the 5'-flanking region of the amylase gene upstream of the heterologous elastase promoter linked to the CAT structural gene. These constructs were transferred to the germ line of transgenic mice by microinjection of fertilized eggs. The amylase sequences had two effects on expression of the elastase promoter: Basal expression was increased, and expression in diabetic animals was reduced to -~2% of basal levels. A 30-bp amylase fragment was sufficient to transfer both of these regulatory functions to the elastase promoter. Sequences within this 30-bp fragment are included in the binding site for the pancreatic nuclear protein PTFI. The close association of the PTFl-binding site and the regulatory functions is consistent with a mechanism based on interference with activation by PTFI in diabetic pancreas. PTFl-binding activity is not reduced in diabetic pancreas. The data presented here demonstrate that the 5'-flanking region of the pancreatic amylase gene contains a novel insulin-dependent element (IDE) that mediates the loss of expression in diabetic animals.

Experimental control of pancreatic development and maintenance

“…Because efficient amylase gene transcription is dependent on insulin (35,36), the decrease of amylase mRNA accumulation may be an indirect effect mediated through decreased insulin production. Although the level of elastase enzyme is affected by long-term diabetes (37), elastase I gene transcription is not affected in the short term by insulin (38). A direct effect on the elastase I gene would be consistent with the demonstrated ability of PDX1 to bind and activate the transcriptional enhancer of the elastase I gene in acinar cells (19).…”

Elements responsible for hormonal control and tissue specificity of L-type pyruvate kinase gene expression in transgenic mice.

“…This L-PK glucose-insulin response element is significantly different from those described so far in the literature (1, 16, 24, 26, 29). These elements have usually been identified through transient- (1,24,29) or stable-transfection experiments in cultured cells (26), except for the insulin-dependent element of the a-amylase gene, initially discovered in transgenic mice (16).…”

Exocrine pancreas transcription factor 1 binds to a bipartite enhancer element and activates transcription of acinar genes.

“…Sequences required for XP enhancer function were defined by deletion and substitution analysis (1,4,13,17,23,26,32,42,56). These studies demonstrated the importance of a 20-nucleotide conserved sequence (the pancreatic consensus) present in each of the identified XP-specific enhancers and in the 5' flanking region of other XP genes (1,5,10,52,54).…”

“…cis-acting DNA sequences which confer XP-specific transcription initially were identified in 5' flanking sequences of the chymotrypsin B gene by using transient transfection of an XP cell line (59) and of the elastase I gene by using transgenic mice (39,40,55). In subsequent gene transfer experiments, it was shown that transcriptional control of these and the amylase 2A and trypsin I genes is mediated by XP-specific enhancers (1,13,42).Sequences required for XP enhancer function were defined by deletion and substitution analysis (1,4,13,17,23,26,32,42,56). These studies demonstrated the importance of a 20-nucleotide conserved sequence (the pancreatic consensus) present in each of the identified XP-specific enhancers and in the 5' flanking region of other XP genes (1,5,10,52,54).…”