Diacylglycerol kinase epsilon in bovine and rat photoreceptor cells. Light-dependent distribution in photoreceptor cells

NATALINI, P.M.; ZULIAN, S.E.; ILINCHETA DE BOSCHERO, M.G.; GIUSTO, N.M.

EXPERIMENTAL EYE RESEARCH

ACADEMIC PRESS LTD-ELSEVIER SCIENCE LTD

Lugar: Amsterdam; Año: 2013 p. 139 - 150

0014-4835

The present study shows the selective light-dependent distribution of 1,2-diacylglycerol kinase epsilon (DAGKe) in photoreceptor cells from bovine and albino rat retina. Immunofluorescence microscopy in isolated rod outer segments from bleached bovine retinas (BBROS) revealed a higher DAGKe signal than that found in rod outer segments from dark-adapted bovine retinas (BDROS). The light-dependent outer segment localization of DAGKe was also observed by immunohistochemistry in retinas from albino rats. DAGK activity, measured in terms of phosphatidic acid formation from a) [3H]DAG and ATP in the presence of EGTA and R59022, a type I DAGK inhibitor, or b) [g-32P]ATP and 1-stearoyl, 2-arachidonoylglycerol (SAG), was found to be significantly higher in BBROS than in BDROS. Higher light-dependent DAGK activity (condition b) was also found when ROS were isolated from dark-adapted rat retinas exposed to light. Western blot analysis of isolated ROS proteins from bovine and rat retinas confirmed that illumination increases DAGKe content in the outer segments of these two species. Light-dependent DAGKe localization in the outer segment was not observed when U73122, a phospholipase C inhibitor, was present prior to the exposure of rat eye cups (in situ model) to light. Conversely, no increased PA synthesis from [3H]DAG and ATP was observed in the presence of neomycin prior to the exposure of bovine eyecups to light. Interestingly, when BBROS were pre-phosphorylated with ATP in the presence of 1,2-dioctanoyl sn-glycerol (diC8) or phorbol dibutirate (PDBu) as PKC activation conditions, higher DAGK activity was observed than in dephosphorylated controls. Taken together, our findings suggest that the selective distribution of DAGKe in photoreceptor cells is a light-dependent mechanism that promotes increased SAG removal and synthesis of 1-stearoyl, 2-arachidonoyl phosphatidic acid in the sensorial portion of this cell, thus demonstrating a novel mechanism of light-regulated DAGK activity in the photoreceptors of two vertebrate species.