Phospholipase C rescues visual defect in norpA mutant of Drosophila melanogaster.
Mutations in the norpA gene of Drosophila melanogaster severely affect the light-evoked photoreceptor potential with strong mutations rendering the fly blind. The norpA gene has been proposed to encode phosphatidylinositol-specific phospholipase C (PLC), which enzymes play a pivotal role in one of the largest classes of signaling pathways known. A chimeric norpA minigene was constructed by placing the norpA cDNA behind an R1-6 photoreceptor cell-specific rhodopsin promoter. This minigene was transferred into norpAP24 mutant by P-element-mediated germline transformation to determine whether it could rescue the phototransduction defect concomitant with restoring PLC activity. Western blots of head homogenates stained with norpA antiserum show that norpA protein is restored in heads of transformed mutants. Moreover, transformants exhibit a large amount of measurable PLC activity in heads, whereas heads of norpAP24 mutant exhibit very little to none. Immunohistochemical staining of tissue sections using norpA antiserum confirm that expression of norpA protein in transformants localizes in the retina, more specifically in rhabdomeres of R1-6 photoreceptor cells, but not R7 or R8 photoreceptor cells. Furthermore, electrophysiological analyses reveal that transformants exhibit a restoration of light-evoked photoreceptor responses in R1-6 photoreceptor cells, but not in R7 or R8 photoreceptor cells. This is the strongest evidence thus far supporting the hypothesis that the norpA gene encodes phospholipase C that is utilized in phototransduction.
McKay
R R
RR
Department of Biological Sciences, State University of New York, Buffalo 14260, USA.
Chen
D M
DM
Miller
K
K
Kim
S
S
Stark
W S
WS
Shortridge
R D
RD
D000818:Animals; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000235:genetics / Q000502:physiology*; D004596:Electroretinography; D005260:Female; D006257:Head; D008297:Male; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D054799:Phospholipase C beta; D010727:Phosphoric Diester Hydrolases / Q000235:genetics* / Q000502:physiology*; D017956:Photoreceptor Cells, Invertebrate / Q000502:physiology; D014170:Transformation, Genetic; D010738:Type C Phospholipases; D014786:Vision Disorders / Q000235:genetics*
D029721:Drosophila Proteins; D010727:Phosphoric Diester Hydrolases; D010738:Type C Phospholipases; C468371:NorpA protein, Drosophila; D054801:Phosphoinositide Phospholipase C; D054799:Phospholipase C beta; D043265:Phosphatidylinositol Diacylglycerol-Lyase