Orientation-dependent Influence of an Intergenic Enhancer on the Promoter Activity of the Divergently Transcribed Mouse Shsp/[alpha]B-crystallin and Mkbp/HspB2 Genes

The mouse Shsp/[alpha]B-crystallin and Mkbp/HspB2 genes are closely linked and divergently transcribed. In this study, we have analyzed the contribution of the intergenic enhancer to Shsp/[alpha]B-crystallin and Mkbp/HspB2 promoter activity using dual-reporter vectors in transient transfection and transgenic mouse experiments. Deletion of the enhancer reduced Shsp/[alpha]B-crystallin promoter activity by 30- and 93-fold and Mkbp/HspB2 promoter activity by 6- and 10-fold in transiently transfected mouse lens [alpha]-TN4 and myoblast C2C12 cells, respectively. Surprisingly, inversion of the enhancer reduced Shsp/[alpha]B-crystallin promoter activity by 17-fold, but did not affect Mkbp/HspB2 promoter activity in the transfected cells. In contrast, enhancer activity was orientation-independent in combination with a heterologous promoter in transfected cells. Transgenic mouse experiments established the orientation dependence and Shsp/[alpha]B-crystallin promoter preference of the intergenic enhancer in its native context. The orientation dependence and preferential effect of the Shsp/[alpha]B-crystallin enhancer on the Shsp/[alpha]B-crystallin promoter provide an example of adaptive changes in gene regulation accompanying the functional diversification of duplicated genes during evolution.