Society for Endocrinology Annual Meeting 2001
London, UK
03 December 2001 - 04 December 2001
Society for Endocrinology
Differential expression of the two mouse insulin genes in MIN6 cells
Endocrinology and Reproduction Research, Kings College London, UK.
Mice and rats are unusual amongst mammals for having 2 structurally similar but non-allelic insulin genes. Both genes are functional but their expression patterns relative to each other have not been conclusively determined. The insulin 1 gene (ins 1) has been shown in several studies to be selectively glucose responsive in mice and rats. We have used the MIN6 cell line to examine passage-related changes in ins 1 and 2 expression and to test glucose responsiveness of the 2 genes. Quantitative PCR techniques were used to measure mRNA levels for ins 1, ins 2 and beta-actin in MIN6 pseudoislets (cell clusters) at early (P32), intermediate (P40) and late (P64) passage and in primary mouse islets. When expressed per fg beta-actin mRNA, mouse islets displayed similar ratios of ins 1:ins 2 expression (57:43% total insulin mRNA). Passage 32 MIN6 cells also showed similar expression patterns to islets (60 : 40% total insulin mRNA). However by passage 40 ins 1 expression fell to less than 10% of total insulin mRNA and at passage 64 this value was reduced to 2% of total insulin mRNA. Glucose responsiveness of the 2 genes was assessed by incubating pseudoislets at either 0 or 20mM glucose for 24 hours. Significant increases in ins 1 expression were detected in both passage 40 but not P64 MIN6 pseudoislets (P40 at 0mM glucose 80.8 plus/minus 4.4, at 20mM glucose 125.3 plus/minus 7.8 fg ins 1/fg actin, P<0.01) but no effect of glucose was observed on expression of the ins 2 gene. Our results suggest that ins 1 and 2 are differentially regulated in MIN6 cells. Expression patterns of the 2 genes change with passage, with expression of the glucose responsive gene (ins 1) diminishing to low levels at relatively early passages.
Endocrine Abstracts 2 P26