Room light decreases rhodopsin in Drosophila rhabdomeres.

 

Selimovic, Asmir, George Denny and William S. Stark. Department of Biology, Saint Louis University, St. Louis, MO 63103. e-mail starkws@slu.edu

 

This laboratoryıs long-lived interest in turnover of rhodopsin in Drosophila visual receptors (Stark et al., 1988) has been rejuvenated in recent years with modern techniques and new insights into relevant gene involvements. The accompanying figure (top) shows visualization of photoreceptor organelles (rhabdomeres) using optical neutralization of the cornea, comparing live white-eyed flies maintained in the dark vs. light; a transgene with the normal rhodopsin promoter (ninaE) drove green fluorescent protein (GFP) labeled Rh1 into R1-6 rhabdomeres. Lower rhabdomere fluorescence in flies kept in the light suggests that light forces rhodopsin turnover, and a haze of fluorescence surrounding these rhabdomeres suggests that we are also visualizing rhodopsin-containing endosomes in the cytoplasm.

            We quickly realized that this finding was not new to our lab. Decades ago, we had quantified a 2-fold dark-light difference in rhabdomeric Rh1 using microscope photometry of rhodopsin-metarhodopsin conversions in the deep pseudopupil (Zinkl et al., 1990). At that time, we presented this finding but did not emphasize it since our purpose was to extend the findings of Ostroy and coworkers that norpA mutants had a light-induced retinal degeneration (Ostroy, 1978; Meyertholen et al., 1987).

            Additionally, a recent study coincidentally offered an explanation of the mechanism of the norpA mutantıs retinal degeneration as well as lightıs involvement in the give-and-take between rhabdomeres and endosomes (Chinchore et al., 2009). Using immunocytochemistry, they found that light exposure moves rhodopsin from rhabdomeres to Rab7-positive endosomes; an overload in endosomes caused by tenacious arrestin binding was offered as the explanation for degeneration in the norpA mutant. They also found that 13 hours of darkness allowed rhodopsin to be cleared from endosomes while newly synthesized rhodopsin transport into the rhabdomere continued.

            We quantified rhodopsin using photometry of the deep pseudopupil in live white-eyed flies to replicate our earlier finding (Zinkl et al., 1990) and to confirm Chinchore et al.ıs (2009) finding that a return to dark re-establishes rhodopsin in the rhabdomere. The accompanying figure (bottom) shows a substantial rhodopsin decrease for light-reared flies when compared with dark-reared flies. We further show a higher rhodopsin level in light-reared flies that had been returned to the dark for 1 day and for 2 days.

            In summary, we used confocal microscopy and microscope photometry, both based on photoreceptor imaging in living flies, to confirm that room light levels of illumination cause rhodopsin to move from rhabdomeres into endosomes and that a return to darkness re-establishes the full amount of rhodopsin in rhabdomeres.

            Acknowledgements: This study was a spin-off of a collaboration with Prof. J. C. Eissenberg at Saint Louis University, and he passed the stock with GFP labeled rhodopsin on to us from Prof. J. E. OıTousa at the University of Notre Dame.

            References: Chinchore, Y., et al., (2009) PLoS Genet 5(2): e1000377; Meyertholen, E. P., et al. (1987) J Comp Phys 161: 793-798; Ostroy, S. E. (1978) J Gen Physiol 72: 717-732; Stark, W.S., et al. (1988) J Neurocytol 17: 499-509; Zinkl G., et al. (1990) Vis Neurosc 5: 429-439.