How do rare endosomal pools of phosphoinositide lipids lead to the congenital disorder Oculocerebral renal syndrome of Lowe (Lowe Syndrome)?

Oculocerebrorenal Syndrome of Lowe (OCRL) is an enzyme, which acts to remove a phosphate from the D-5 position of the inositol ring of phosphoinositides. There are seven phosphoinositides, which are lipids that specify subcellular membrane identity and are crucial for regulating processes including vesicular trafficking. Mutations in OCRL have been associated with the X-linked disorder, Lowe syndrome. This disease is characterised by renal tubular dysfunction, behavioural difficulties, developmental delay and congenital cataracts and has an estimated prevalence of approximately 1 in 500,000 in the general population. This enzyme possesses four domains, starting with the N-terminal Pleckstrin Homology (PH) domain, a central 5-phosphatase domain that acts against PI(4,5)P2 and PI(3,4,5)P3 and a C-terminal ASPM-SPD2-hydin (ASH) domain linked by a hydrophobic interface to a catalytically inactive Rho GTPase activating (RhoGAP) domain. OCRL is critical for the process of membrane trafficking and localises to multiple stations of the endocytic pathway. Evidence has arisen that the activity of OCRL against PI(4,5)P2 is even critical on the Golgi, endosomes and lysosomes, all of which possess very low levels of PI(4,5)P2. This project will involve studying various mutations within OCRL linked to Lowe syndrome and firstly will involve characterising the process of endocytosis in OCRL mutants, mainly using fluid-phase dye uptake. This aspect of the project will involve utilising Dictyostelium discoideum, which possesses a single OCRL-like protein, known as Dd5P4, which is involved in the regulation of phagocytosis. The loss of Dd5P4 is also not lethal, making Dictyostelium highly convenient for this study. The second aspect of this project would involve studying and understanding the role of the three proteins that interact with OCRL through usage of a small 11-13 amino acid peptide motif called the F&H motif, located on the RhoGAP domain. The three proteins in question are the APPL1 and Ses1/2 endocytic adaptor proteins and a Rho-guanine nucleotide exchange factor (GEF) called Frabin. Previous work has demonstrated that missense mutations within the ASH-RhoGAP domain abolish interactions with APPL1 and Ses1/2 proteins for example, which each contain an F&H motif. This project will overall provide a greater understanding of the mechanistic insights as to why these mutations within OCRL lead to the onset of Lowe syndrome. This work could also in the long-term lead to the development of drugs to treat this disease to alleviate the symptoms and improve the quality of life or extend the life span of individuals with Lowe syndrome beyond the current 40 years of age.