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.

We conducted a genomewide linkage screen of a simple heavy-smoking quantitative trait, the maximum number of cigarettes smoked in a 24-h period, using two independent samples: 289 Australian and 155 Finnish nuclear multiplex families, all of which were of European ancestry and were targeted for DNA analysis by use of probands with a heavy-smoking phenotype. We analyzed the trait, using a regression of identity-by-descent allele sharing on the sum and difference of the trait values for relative pairs. Suggestive linkage was detected on chromosome 22 at 27-29 cM in each sample, with a LOD score of 5.98 at 26.96 cM in the combined sample. After additional markers were used to localize the signal, the LOD score was 5.21 at 25.46 cM. To assess the statistical significance of the LOD score in the combined sample, 1,000 simulated genomewide screens were conducted, resulting in an empirical P value of .006 for the LOD score of 5.21. This linkage signal is driven mainly by the microsatellite marker D22S315 (22.59 cM), which had a single-point LOD score of 5.41 in the combined sample and an empirical P value <.001 from 1,000 simulated genomewide screens. This marker is located within an intron of the gene ADRBK2, encoding the beta-adrenergic receptor kinase 2. Fine mapping of this linkage region may reveal variants contributing to heaviness of smoking, which will lead to a better understanding of the genetic mechanisms underlying nicotine dependence.

The significant worldwide health burden introduced by tobacco smoking highlights the importance of studying the genetic determinants of smoking behavior and the key factor sustaining compulsive smoking, that is, nicotine dependence (ND). We have here addressed the genetic background of smoking in a special study sample of twins, harmonized for early life events and specifically ascertained for smoking from the nationwide twin cohort of the genetically unique population of Finland. The twins and their families were carefully examined for extensive phenotype profiles and a genomewide scan was performed to identify loci behind the smoking status, ND and the comorbid phenotype of ND and alcohol use in 505 individuals from 153 families. We replicated previous linkage findings on 10q (max logarithm of the odds (LOD) 3.12) for a smoker phenotype, and on 7q and 11p (max LOD 2.50, and 2.25, respectively) for the ND phenotype. The loci linked for ND also showed evidence for linkage for the comorbid phenotype. Our study provides confirmatory evidence for the involvement of these genome regions in the genetic etiology of smoking behavior and ND and for the first time associates drinking and smoking to a shared locus on 10q.