Two Distinct Aquaporin 0s are Required for Development and Transparency of the Zebrafish Lens

Purpose: AQP0, formerly known as MIP26, likely has multiple separate functions in the mammalian lens including water transport, formation of thin junctions and interactions with other lens components. Though mammalian genomes contain only one AQP0 gene the zebrafish genome contains two, Aqp0a and Aqp0b, and the putative multiple functions of the single mammalian protein may be divided between these two genes. Our purpose is to exploit this gene duplication and divergence to illuminate the multiple functions of AQP0 in the lens. Methods: Whole mount in situ hybridization and western blots were used to determine the expression pattern of Aqp0a and Aqp0b. The role of both proteins was studied in vivo by microinjection of antisense morpholino oligos (MO) in zebrafish. The water permeability of both proteins was tested using the Xenopus oocyte swelling assay and a yeast shrinkage assay. Results: We showed that both genes, like their mammalian counterparts, are expressed in the lens. Morpholino knock-down of either gene alone led to cataract formation, indicating that both genes are necessary for normal lens development and transparency. We demonstrated that full-length Aqp0a is a functional water channel when expressed in Xenopus oocytes and in yeast, whereas Aqp0b was not. However, addition of an HA-tag at its N-terminus converted Aqp0b to a water channel in Xenopus oocytes. Conclusions: We propose that Aqp0a is the primary water channel of the lens and that Aqp0b, though possibly a secondary water channel, has an unidentified function in the lens.