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Screening of a Drosophila genomic DNA library at reduced stringency hybridization conditions using a rat PLC alpha cDNA probe yielded a gene which encodes a member of the protein disulfide isomerase/PLC alpha family. The gene has been localized to band 74C on the left arm of the third chromosome and has been designated dpdi. Northern analysis shows that the dpdi gene encodes a transcript that is 2.3 kb in length and is present throughout development as well as in both heads and bodies of adults. The deduced dpdi protein is 496 amino acids in length and contains two domains exhibiting high similarity to thioredoxin, two regions that are similar to the hormone binding domain of human estrogen receptor, and a sequence of four amino acids (KDEL) at the C-terminus which has been described by others as being responsible for retention of proteins in the endoplasmic reticulum. Overall, dpdi contains a higher similarity to rat protein disulfide isomerase (53% identical) than to rat PLC alpha (30% identical). However, it is unclear whether dpdi functions in vivo as a PDI or as a PLC, or both. Drosophila, with its well characterized genetics and the ability to generate mutants in a gene that has been cloned, provides an excellent system in which to resolve this issue.

Severe mutations within the norpA gene of Drosophila abolish the photoreceptor potential and render the fly blind by deleting phospholipase C, an essential component of the phototransduction pathway. To study the membrane association of phospholipase C, we have utilized biochemical assays of phospholipase C activity, which predominant measurable phospholipase C activity in head homogenates has been shown to be encoded by norpA, as well as antisera generated against the major gene product of norpA to examine its subcellular distribution before and during phototransduction. We find that both phospholipase C activity and the norpA protein are predominantly associated with membrane fractions in heads of both light- and dark-adapted flies. Moreover, phospholipase C activity as well as norpA protein can be easily extracted from membrane preparations of light- or dark-adapted flies using high salt, indicating that the norpA protein is peripherally localized on the membrane. These data suggest that the norpA encoded phospholipase C of Drosophila is a permanent peripheral membrane protein. If this is indeed the case, then it would mean that the reversible redistribution of phospholipase C from the cytosol to the membrane, as observed in epidermal growth factor receptor stimulation of mammalian phospholipase C gamma, is not a universal mechanism utilized by all types of phosphatidylinositol-specific phospholipase C.