Retinoid deprivation reduces phospholipase C (PLC-β), product of the Drosophila norpA gene

Purpose. Northern and Western (Stark, et al., ARVO-95, IOVS 36, s771, 1995) and promoter-reporter analyses (Sun, et al., Exp. Eye Res. 57, 177-187, 1993) showed that retinoid deprivation reduces Drosophila opsin, likely at the transcriptional level. We examined phospholipase C (PLC) to determine if deprivation affected downstream phototransduction molecules. Methods. Flies were retinoid deprived on Sang's medium (Sun et al.). An antiserum against the major gene product of the norpA gene (Zhu, et al., J. Biol. Chem. 268, 15994-16001, 1993) was used in Western blots and in immunocytochemistry with 1 μm LR White sections. SDS-PAGE gels were blotted and visualized by autoradiography with ¹²⁵I labeled protein G. We used FITC conjugated secondaries to localize PLC using fluorescence and confocal microscopes; negative controls included norpA mutants and secondary only. Results. Western blots revealed the expected 130-kDa band (McKay, et al., J. Biol. Chem. 270, 13271-13276, 1995) and showed that retinoid deprivation reduced PLC by about half. Our fluorescence immunocytochemistry confirmed rhabdomeric localization of PLC (see McKay, et al.). Rhabdomeres showed equal fluorescence, eliminated by norpA, in retinoid deprived and replete flies, but rhabdomeres were smaller in deprived flies, in agreement with Sapp, et al. (Exp. Eye Res. 53, 73-79, 1991). Conclusions. Retinoid deprivation reduced PLC but not as severely as opsin's near elimination (Stark, et al.; deCouet & Tanimura, Eur. J. Cell. Biol. 44, 50-56, 1987). Thus, the PLC recuction may be based on the reduction in the size of the organelle, suggesting that it results as a consequence of opsin reduction. In other words, it is unlikely that retinoids regulate the PLC gene promoter directly. An interesting contrast involves Drosophila retinoid binding protein (DRBP) which is nearly eliminated by retinoid deprivation (R. D. Lee, MS Thesis, Saint Louis University, Biology, 1994) and is expressed in cone (Semper) cells rather than in receptors (Shim, et al., Neurosci. Abs. 21, 1174, 1995), suggesting possible direct control by retinoids.