LIG_OCRL_FandH_1

Three known human proteins possess a small 11-13-mer peptide determined by a strictly conserved phenylalanine and histidine residue, the F and H motif. These proteins are APPL1 (also known as DCC-interacting protein DIP-13alpha) (Q9UKG1) (2EJ8) and the two Ses ("Sesquipedalian") proteins Ses1 and 2 (also called IPIP27 A and B) (Q8N4B1; Q6ICB4) (Swan,2010; Noakes,2011). This F and H motif binds into the groove of an ASH-RhoGAP-like domain complex of the inositol polyphosphate 5-phosphatases OCRL1 (Q01968) and INPP5B (P32019). The endocytic adaptor proteins APPL1 and Ses form temporally and spatially defined sub-complexes with OCRL1 or INPP5B through the F and H motif (Coon,2012). Nevertheless, it seems to be essential for a range of cellular processes involved in membrane trafficking, including endocytosis and endosomal maturation, phagocytosis, receptor recycling and primary cilia assembly.
Among the trans-Golgi-network (TNG) OCRL1 is localized to clathrin-coated vesicles, early endosomes and macropinosomes (Vicinanza,2011). Mutations in OCRL1 that disrupt the interaction with the F and H motif containing proteins, but not with other substrates of OCRL, like Rab or clathrin, impair OCRL1's endosomal localization and cause defects in endocytic trafficking and signalling (Erdmann,2007, Choudhury,2009). Essentially, the interaction of 5-phosphatases with F and H proteins participate in regulating the level and pattern of inositol polyphosphates in a highly temporally and spatially controlled manner that is crucial for endosomal morphology and function (Noakes,2011). APPL1 co-localizes with OCRL1 in a different subset of OCRL1-positive endosomes than Ses. Immature multivesicular bodies called signalling endosomes that harbor the phospholipids PtdIns(4,5)P2 and PtdIns(3,4,5)P3 are associated with APPL1. On the other hand sorting endosomes, a form of early endosomes which predominantly host PtdIns(3)P, are associated with Ses (Ooms,2009, Swan,2010). In addition, APPL1-positive macropinosomes exclusively convert into Ses-positive compartments (Swan,2010). Thus, APPL1 and Ses reside in distinct vesicle populations and associate sequentially with endosomes. The competitive interaction of OCRL1 with APPL1 and Ses through the F and H motif may provide a regulatory mechanism of vesicle trafficking by generating two distinct stages for vesicle maturation. In this model, the APPL1-OCRL1 interaction represents the earliest stage of endocytic traffic for a subset of endosomal vesicles derived from clathrin-coated pits that for example internalize growth factor receptors and macropinosomes (Swan,2010). The maturation of APPL1-positive signalling endosomes and their transformation into sorting endosomes is controlled by a PtdIns (PI) switch to PtdIns(3)P performed by 5-phosphatases (Zoncu,2009). Ses proteins seem to contribute to loss of APPL1 and termination of APPL1 signalling. Therefore, Ses acts as negative regulators for APPL1 signalling by out-competiting APPL1 for OCRL1 binding. Since Ses exclusively localizes to sorting endosomes and share the F and H motif binding OCRL1 competitively with a higher affinity, Ses has the ability to displace APPL1 from the membrane to preserve the strict demarcation between the signalling and sorting endosomes (Noakes,2011).
Phagocytosis is a process by which particulate material is endocytosed, resulting in the formation of large intracellular membrane-bound vesicles called phagosomes. This process is important for example as a defense mechanism against pathogen invasion. During pathogen invasion a serine/threonine kinase called Akt is activated, which inhibits apoptosis and needs to be closely regulated to achieve an appropriated immune response (Bohdanowicz,2012). APPL1 is a Rab5 effector that binds the activated GTP-bound form of Rab5 and interacts with Akt (Miaczynska,2004). Since Akt is activated by PtdIns(3,4,5)P3, which is formed at phagosomes, and Rab5 recruits APPL1 to that site, APPL1 modulates Akt signaling via OCRL1 and INPP5B (Bohdanowicz,2012). The 5-phosphatases hydrolyze PtdIns(4,5)P2 and PtdIns(3,4,5)P3 and concomitantly terminate PtdIns(3,4,5)P3 synthesis, so that Akt loses its activation factor limiting the duration of Akt signaling. Thus, the interaction of APPL1 with OCRL1 and INPP5B in phagosomes results in the premature termination of Akt activity (Bohdanowicz,2012).
Additionally, APPL1 interacts with the oligomeric scaffold protein GIPC (Gα-interacting protein-interacting protein C-terminus). Both APPL1 and GIPC bind to the nerve growth factor receptor TrkA (tropomyosin receptor kinase A) (Erdmann,2007; Ooms,2009). Through the F and H motif APPL1 links OCRL1 to TrkA, which is important for vesicular trafficking and recycling of the receptor. Disturbance of the interaction could be complicit in neurological disorder and cognitive deficits of Lowe Syndrome (McCrea,2008). Moreover, through the F and H motif of APPL1 OCRL1 is connected to GIPC, which directly binds the scavenger receptor megalin mediating the uptake of low molecular weight proteins in kidney (McCrea,2008). Depletion of OCRL1 impairs the recycling of megalin in proximal tubule cells (Vicinanza,2011). These findings provide a molecular network that links OCRL to the reabsorption machinery of the proximal tubule in the kidney, which is also deficient in Lowe disease patient (Erdmann,2007). Additionally, the Ses proteins appear to be important for transferrin receptor (TfR) homeostasis, since Ses depletion impairs trafficking from endosomes to the TGN (Noakes,2011). Ses seems to link OCRL to sorting and recycling endosomes to achieve receptor recycling (Noakes,2011).
Coon,2012 showed that OCRL1 mediated endosomal trafficking is also essentially involved in primary cilia assembly. Assembly depends on the interaction of OCRL1 with Rab8 and the F and H motif containing endocytic proteins affecting the endocytic routes for primary cilia protein localization. OCRL1 knockdown experiments revealed a crucial role in ciliogenesis for OCRL1 resulting in defects in the assembly of primary cilia. Remarkably, defects could be rescued by OCRL1/wt or (partially) by the OCRL1 homolog INPP5B, demonstrating a partial functional redundancy. Moreover, Ses depletion inhibited ciliogenesis, whereas Ses overexpression mended the OCRL1/kd phenotype. Indeed, mutations that abolish the interaction of OCRL1 with F and H motif containing proteins impair primary cilia assembly (Coon,2012).
The F and H motif in APPL1 provides potential phosphorylation sites, which in their phosphorylated state abolish the interaction with OCRL1. Overexpression of protein kinase A revealed a reduced interaction between OCRL1 and APPL1, whereas phosphomimetic mutations and in vitro phosphorylation lead to a complete loss of binding (Erdmann,2007, Pirruccello,2011). The phosphorylation-dependent regulation of the F and H motif supports its physiological significance (Erdmann,2007). Finally, the interaction of APPL1 with OCRL1/INPP5B is influenced by the presence of Rab5-GTP because of the dependency of APPL1's membrane localization on the GTP-bound form of Rab5; it is therefore sensitive to GTP hydrolysis (Miaczynska,2004). Remarkably, the vast majority of known Lowe syndrome patient mutations abolish the interaction of OCRL1 with APPL1 and Ses (Loi,2006, Pirruccello,2011). The disruption of these interactions may play a major role in the pathogenesis of Lowe syndrome and the related Dent's disease.