Knock-in of α3 connexin prevents severe cataracts caused by an α8 point mutation

A G22R point mutation in α8 connexin (Cx50) has been previously shown to cause a severe cataract by interacting with endogenous wild-type α3 connexin (Cx46) in mouse lenses. Here, we tested whether a knocked-in α3 connexin expressed on the locus of the endogenous α8 connexin could modulate the severe cataract caused by the α8-G22R mutation. We found that the α3(-/-) α8(G22R/-) mice developed severe cataracts with disrupted inner fibers and posterior rupture while the α3(-/-) α8(G22R/ KIα3) lens contained relatively normal inner fibers without lens posterior rupture. The α8-G22R mutant proteins produced typical punctate staining of gap junctions between fiber cells of α3(-/-) α8(G22R/KIα3) lenses, but not in those of α3(-/-) α8(G22R/-) lenses. Thus, we hypothesize that the knocked-in α3 connexin subunits interact with the α8-G22R connexin subunits to form functional gap junction channels and rescue the lens phenotype. Using an electrical coupling assay consisting of paired Xenopus oocytes, we demonstrated that only co-expression of mutant α8-G22R and wild-type α3 connexin subunits forms functional gap junction channels with reduced conductance and altered voltage sensitivity compared with the channels formed by α3 connexin subunits alone. Thus, knocked-in α3 connexin and mutant α8-G22R connexin probably form heteromeric gap junction channels that influence lens homeostasis and lens transparency.