Abstract
Purpose: :
The subterranean blind mole rat (Spalax galili) has regressed, subcutaneous eyes. Compared to its mouse counterpart, the αB–crystallin promoter of the mole rat has little lens but enhanced muscle activity in transgenic mice. The purpose of this study is to identify the molecular basis for the difference in αB–crystallin promoter activity between mice and mole rats.
Methods: :
Site–specific mutagenesis, transient transfections (using mouse muscle C2C12 and lens αTN4–1 cells) and transgenic mice were performed by conventional methods. Electrophoretic mobility shift assays were used to study DNA:protein interactions.
Results: :
A 31–nt sequence (–285/–255) was identified in the blind mole rat promoter that differs from that in the mouse. –273CA in the mouse promoter was mutated to G, as occurs in the blind mole rat promoter. While the activity of the wild type and mutant mouse αB–crystallin promoter was similar in transfected cells, the mutant lost lens activity and gained skeletal muscle activity in transgenic mice; thus, the activity of the mutated mouse promoter resembled that of the mole rat promoter. Gel mobility assays showed that the homologous 31nt promoter sequences of blind mole rat and mouse bind distinct nuclear factor(s). In addition, the gel mobility shift pattern of the mutated mouse sequence also resembled that generated with the mole rat sequence. Competition analysis of gel mobility shift assays suggested that AP2 may be engaged in the interactions with the mole rat sequence.
Conclusions: :
We have identified a specific position within the blind mole rat αB–crystallin promoter that may play a critical role in the evolutionary changes of the promoter function.
Keywords: crystalline lens • gene/expression