Lowe syndrome is a rare X-linked disease characterized by congenital
cataracts, defects in renal tubule cell function, and
mental retardation. Mutations in the OCRL1 gene, which encodes ocrl1, a phosphatidylinositol-4,5-bisphosphate (
PtdIns(4,5)
P(2)) 5-phosphatase, are the cause of
Lowe syndrome.
PtdIns(4,5)
P(2), a substrate of ocrl1, is an important signaling molecule within the cell. OCRL1 is ubiquitously expressed and co-localizes with the trans-Golgi network (TGN) and endosomal
proteins. The ocrl1
protein contains two recognizable domains, one a conserved Ptd(4,5)
P(2) 5-phosphatase domain and the other with homology to
Rho GTPase activating
proteins (RhoGAPs). The objective of our study was to further characterize the ocrl1
RhoGAP-homology domain by analyzing the effect of two missense mutations in this domain, I751N and A780P, which were previously reported in
Lowe syndrome patients. Both
mutant proteins were expressed at levels similar to wild-type but their
enzyme activity was reduced by 85-90%, indicating that the
RhoGAP-homology domain is important for the enzymatic function of ocrl1. Study of a C-terminal region of wild-type ocrl1 containing this domain detected no GAP activity, eliminating the possibility of an effect by mutations in this domain on
GTPase activation. Because members of the Arf family of
small G-proteins are directly involved in (Ptd(4,5)
P(2)) signaling and localize to the TGN like ocrl1, we analyzed by immunoprecipitation the interaction of ocrl1 with Arf1 and Arf6 via its
RhoGAP-homology domain. Wild-type ocrl1, but not the I751N
mutant protein, co-immunoprecipitated with these two Arf
proteins. These results indicate that wild-type ocrl1 and Arf
proteins can interact and that this interaction is disrupted by the mutation. It remains unknown whether a disrupted interaction between Arf and ocrl1 plays a role in the
Lowe syndrome phenotype.