Analysis of the phosducin knockout mouse reveals roles for phosducin in regulation of transducin ß subunits level and in the light–dependent transducin translocation

Abstract

Abstract: : Purpose: Phosducin is an abundant photoreceptor–specific protein which interacts with the ßγ subunits of transducin. We generated the phosducin knockout mouse in order to address the physiological function of phosducin in intact rods. Methods:The amounts of phosducin and transducin subunits in the retina were measured by quantitative Western blotting. Light–dependent transducin translocation in rods was quantified by serial tangential cryosectioning with Western blotting. Electroretinographic (ERG) recordings were conducted according to standard protocols. Results:Phosducin knockout mice develop morphologically unaltered rods that do not display any signs of degeneration up to the age of 6 months. The absence of phosducin in their retinas was confirmed by Western blotting. ERG analysis of the dark–adapted animals revealed no statistically significant difference between the a–waves recorded from phosducin knockouts and wild type mice. The amount of transducin α subunit in the rods of phosducin knockouts was normal, whereas the amount of transducin ßγ subunits was ∼30% smaller than that of the wild type mice. The knockout also resulted in the decrease in the light–dependent transducin translocation with the degree of transducin ßγ subunits translocation being affected the most (4.3– and 1.6–fold ßγ translocation in the wild type and phosducin knockout, respectively. The degree of transducin α subunit translocation was also reduced (3.0–fold translocation in wild type vs.1.9–fold in phosducin knockout mice). Conclusions: Phosducin is a multi–function protein. In rods, it is likely to regulate both transducin ßγ subunits level and the light–dependent translocation of transducin subunits.

Keywords: photoreceptors • protein structure/function • electroretinography: non–clinical