The endosomal/lysosomal transmembrane protein CLN3 is mutated in the Batten 
disease (juvenile neuronal ceroid lipofuscinosis, JNCL). However, the molecular 
mechanism of JNCL pathogenesis and the exact function of the CLN3 protein have 
remained unclear. Previous studies have shown that deletion of BTN1, the yeast 
orthologue of CLN3, leads to increased expression of BTN2. BTN2 encodes Btn2p, a 
proposed homologue to a novel microtubule-binding protein Hook1, which regulates 
endocytosis in Drosophila. We analysed here the putative interconnection between 
CLN3 and Hook1 in the mammalian cells and discovered that overexpression of 
human CLN3 induces aggregation of Hook1 protein, potentially by mediating its 
dissociation from the microtubules. Using in vitro binding assay we were able to 
demonstrate a weak interaction between Hook1 and the cytoplasmic segments of 
CLN3. We also found receptor-mediated endocytosis to be defective in 
CLN3-deficient JNCL fibroblasts, connecting CLN3, Hook1 and endocytosis in the 
mammalian system. Moreover, co-immunoprecipitation experiments showed that Hook1 
physically interacts with endocytic Rab7, Rab9 and Rab11, hence delineating a 
manifold role for mammalian Hook1 in membrane trafficking events. These novel 
interactions between the microtubule-binding Hook1 and the large family of Rab 
GTPases also suggest a link between CLN3 function, microtubule cytoskeleton and 
endocytic membrane trafficking.