The cooperative interaction between two motifs of an enhancer element of the chicken alpha A-crystallin gene, alpha CE1 and alpha CE2, confers lens-specific expression.

An 84 bp element located between nucleotides -162 and -79 of the chicken alpha A-crystallin gene exhibits lens-specific enhancer activity. Transient transfection experiments using 5' deletion and linker scanner mutants has indicated that the 84 bp enhancer element is composed of three motifs, alpha CE1 (-162 to -134), alpha CE3 (-135 to -121) and alpha CE2 (-119 to -99). Neither alpha CE1 or alpha CE3 motif alone can exhibit enhancer activity even when trimerized, whereas together they can direct some degree of lens-specific expression. alpha CE2 alone shows low transcriptional activity when trimerized. A combination of alpha CE1 with alpha CE2 exerts full lens-specific enhancer activity comparable with that of the 84 bp enhancer element, indicating that alpha CE1 and alpha CE2 motifs are sufficient to confer lens-specific expression. Transcriptional activation by these two motifs from a distance required the additional presence of either or both motifs adjacent to the beta-actin basal promoter. Gel shift experiments indicated that the alpha CE1, alpha CE2 and alpha CE3 motifs specifically bind nuclear proteins. alpha CE1 binds a protein predominantly present in lens cells, whereas alpha CE2- and alpha CE3-binding proteins differ between lens and lung cells. Mutations within the alpha CE1 and alpha CE2 motifs that failed to bind nuclear factors in vitro resulted in loss of transcriptional activation, indicating that these nuclear factors play a key role in controlling lens-specific expression.