Assay and functional analysis of the ARL3 effector RP2 involved in X-linked retinitis pigmentosa.
Mutations in RP2 cause X-linked retinitis pigmentosa and also macular and peripapillary atrophy. RP2 is a functional homologue of the tubulin folding cofactor, cofactor C, as it can replace the beta tubulin GTPase stimulating activity of cofactor C in an in vitro assay. An important difference between RP2 and cofactor C is their subcellular localization. RP2 is targeted to the cytoplasmic face of the plasma membrane by dual N-terminal acylation, and this post-translational modification is important for protein function. The activity of tubulin folding cofactors is modulated by certain ADP ribosylation factor-like (Arl) proteins. It has been shown that RP2 can interact directly with Arl3. Here we describe the methodologies that we have developed to analyze the interaction of RP2 with Arl3 and to investigate the effect of RP2 post-translational modifications on its subcellular and tissue localization.
Evans
R Jane
RJ
Institute of Ophthalmology, University College London, United Kingdom.
Chapple
J Paul
JP
Grayson
Celene
C
Hardcastle
Alison J
AJ
Cheetham
Michael E
ME
D020727:ADP-Ribosylation Factors / Q000032:analysis* / Q000502:physiology*; D002499:Centrifugation, Density Gradient; D003001:Cloning, Molecular; D005136:Eye Proteins / Q000032:analysis* / Q000502:physiology*; D019204:GTP-Binding Proteins; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D019814:Myristic Acid / Q000378:metabolism; D025941:Protein Interaction Mapping / Q000379:methods; D012160:Retina / Q000737:chemistry / Q000648:ultrastructure; D012174:Retinitis Pigmentosa / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D020798:Two-Hybrid System Techniques
D005136:Eye Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; C072178:RP2 protein, human; D019814:Myristic Acid; D019204:GTP-Binding Proteins; D020727:ADP-Ribosylation Factors; C490247:ARL3 protein, human