Interaction of Aryl Hydrocarbon Receptor-interacting Protein-like 1 with the Farnesyl Moiety

Anurima Majumder,Kota N Gopalakrishna,Pallavi Cheguru,Lokesh Gakhar,Nikolai O Artemyev

doi:10.1074/jbc.m113.476242

Abstract

Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) is a photoreceptor specific chaperone of the visual effector enzyme phosphodiesterase-6 (PDE6). AIPL1 has been shown to bind the farnesylated PDE6A subunit. Mutations in AIPL1 are thought to destabilize PDE6 and thereby cause Leber congenital amaurosis type 4 (LCA4), a severe form of childhood blindness. Here, we examined the solution structure of AIPL1 by small angle x-ray scattering. A structural model of AIPL1 with the best fit to the scattering data features two independent FK506-binding protein (FKBP)-like and tetratricopeptide repeat domains. Guided by the model, we tested the hypothesis that AIPL1 directly binds the farnesyl moiety. Our studies revealed high affinity binding of the farnesylated-Cys probe to the FKBP-like domain of AIPL1, thus uncovering a novel function of this domain. Mutational analysis of the potential farnesyl-binding sites on AIPL1 identified two critical residues, Cys-89 and Leu-147, located in close proximity in the structure model. The L147A mutation and the LCA-linked C89R mutation prevented the binding of the farnesyl-Cys probe to AIPL1. Furthermore, Cys-89 and Leu-147 flank the unique insert region of AIPL1, deletion of which also abolished the farnesyl interaction. Our results suggest that the binding of PDE6A farnesyl is essential to normal function of AIPL1 and its disruption is one of the mechanisms underlying LCA.

Highlights

Mutations in Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1), a chaperone of the lipidated visual effector phosphodiesterase-6, cause severe childhood blindness
Analysis of AIPL1 Structure by SAXS—To guide our analysis of the interaction of AIPL1 with the farnesyl moiety, we examined the structure of mouse AIPL1 in solution by SAXS and generated a molecular model of the protein that agrees the best with the scattering data
AIPL1 is a specific chaperone for the visual effector enzyme, PDE6

Summary

Background

Mutations in AIPL1, a chaperone of the lipidated visual effector phosphodiesterase-6, cause severe childhood blindness. Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) is a photoreceptor specific chaperone of the visual effector enzyme phosphodiesterase-6 (PDE6). Our studies revealed high affinity binding of the farnesylated-Cys probe to the FKBP-like domain of AIPL1, uncovering a novel function of this domain. Our results suggest that the binding of PDE6A farnesyl is essential to normal function of AIPL1 and its disruption is one of the mechanisms underlying LCA. LCA-linked AIPL1 mutations are found in all three domains: the FKBP-like, TPR, and the proline-rich regions (Fig. 1A). Our analysis demonstrated high affinity binding of the farnesylated-Cys probe to the FKBP-like domain of AIPL1, thereby revealing a novel function of this domain. An LCA4-causing mutation C89R was found to abrogate the probe binding, suggesting an essential role for the farnesyl-mediated PDE6A interaction in normal function of AIPL1

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION