Molecular identification and characterization of mammalian homologs of proteins important in Drosophila photoreceptor function

Paulo Alexandre M. P Ferreira, Purdue University

Abstract

The overall goal of this project is the molecular identification and characterization of bovine retina-specific homologs of two Drosophila melanogaster photoreceptor machinery components, norpA and ninaA. Conserved regions of norpA cDNA were used to isolate bovine cDNAs that would encode four alternative forms of phospholipase C of the $\beta$ class that are highly homologous to the norpA protein and expressed preferentially in the retina. Two of the variants are highly unusual in that they lack much of the N-terminal region present in all other known phospholipases C. The sequence conservation between these proteins and the norpA protein is higher than that between any other known phospholipases C. GTPase sequence motifs found in proteins of the GTPase superfamily are found conserved in all four variants of the bovine retinal protein as well as the norpA protein but not in other phospholipases C. The norpA-homologous bovine retinal PI-PLCs, though found in other retinal neurons as well, are found specifically in cones but not in rods. The results suggest that these proteins together with the norpA protein constitute a distinctive subfamily of phospholipases C that are closely related in structure, function, and tissue distribution. Furthermore, immunocytochemical results suggest that the phototransduction in cones is not identical to rods and may utilize phospholipase C in addition to phosphodiesterase. Mutations in the norpA gene in addition to blocking phototransduction, cause light-dependent degeneration of photoreceptors. In view of the strong similarity in structure and tissue distribution, a defect in these proteins may have similar consequences in the mammalian retina. The Drosophila ninaA gene encodes a photoreceptor-specific cyclophilin (peptidyl-prolyl cis-trans isomerase (PPlase); cyclosporin A (CsA)-binding protein) thought to play a critical role in rhodopsin folding during its synthesis or maturation. Cyclophilins comprise a highly conserved family of proteins which are the primary targets of the potent immunosuppressive drug, cyclosporin A (CsA). However, with the exception of ninaA protein, the in vivo functions of the cyclophilins are poorly understood. A probe derived from the ninaA cDNA was used to isolate bovine cDNAs that would encode at least two major alternative forms of cyclophilins that are specifically expressed in the retina. These proteins represent new classes of cyclophilins with novel structural features and with strongly reduced PPlase and CsA-binding activities in contrast to other known mammalian cyclophilins. Immunocytochemical results show that the ninaA-like proteins, though found in other retinal neurons as well, are found specifically in cones and not in rod photoreceptors. (Abstract shortened by UMI.)

Degree

Ph.D.

Advisors

Pak, Purdue University.

Subject Area

Neurology|Molecular biology

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