Mutations in a member of the Ras superfamily of small GTP-binding proteins causes Bardet-Biedl syndrome

Abstract

RAB, ADP-ribosylation factors (ARFs) and ARF-like (ARL) proteins belong to the Ras superfamily of small GTP-binding proteins and are essential for various membrane-associated intracellular trafficking processes 1,2 . None of the B50 known members of this family are linked to human disease. Using a bioinformatic screen for ciliary genes in combination with mutational analyses, we identified ARL6 as the gene underlying Bardet-Biedl syndrome type 3, a multisystemic disorder characterized by obesity, blindness, polydactyly, renal abnormalities and cognitive impairment 3,4 . We uncovered four different homozygous substitutions in ARL6 in four unrelated families affected with Bardet-Biedl syndrome, two of which disrupt a threonine residue important for GTP binding 5 and function 5-7 of several related small GTP-binding proteins. Analysis of the Caenorhabditis elegans ARL6 homolog indicates that it is specifically expressed in ciliated cells, and that, in addition to the postulated cytoplasmic functions of ARL proteins, it undergoes intraflagellar transport. These findings implicate a small GTP-binding protein in ciliary transport and the pathogenesis of a pleiotropic disorder.Cilia and flagella are ancient, evolutionarily conserved eukaryotic organelles that project from cells and have been adapted by organisms to carry out diverse biological functions 8 . The assembly, maintenance and function of cilia and flagella depend on intraflagellar transport (IFT), and defects in this microtubule-based transport process and the function of cilia are associated with several human diseases, including Bardet-Biedl syndrome (BBS) [8][9][10] . Genes underlying seven of the eight loci known to be associated with BBS have been identified 4,11 ; only the gene mutated in BBS type 3 (called BBS3), previously mapped to 3p12 (refs. 12,13), remained unidentified. BBS is thought to result largely from ciliary dysfunction, because loss-of-function mutations in C. elegans bbs-7 and bbs-8 compromise cilia structure and function 14 and RNA interference of Chlamydomonas BBS5 results in the loss of flagella 11 . Notably, all known C. elegans bbs genes are expressed exclusively in cells with cilia, owing to the presence of a DAF-19 RFX transcription factor binding site (X box) in their promoters 10,11 . We hypothesized that the C. elegans ortholog of human BBS3 would also contain this regulatory element, which would allow us to identify candidates from the 490 genes that map to the BBS3 critical interval 12,13,15 . We generated a consensus X-box sequence from a training set of 14 C. elegans genes containing X boxes that are known to be strictly expressed in ciliated cells and used them to scan the C. elegans genome. We identified 368 genes with an X-box sequence within 1.5 kb of the start codon, 168 of which had a bona fide human ortholog (E value r 10 À6 ); three of these fell in the BBS3 critical interval (Fig. 1a). The first gene, ESRRBL1, is probably the human ortholog of C. elegans che-13. che-13 is expressed exclusively in ciliated neurons ...

Pitfalls of homozygosity mapping: an extended consanguineous Bardet–Biedl syndrome family with two mutant genes (BBS2, BBS10), three mutations, but no triallelism

“…13 This combination of homozygosity mapping and comparative genomics allowed the identification of BBS3, BBS5 and BBS9. 5,6,8,13 Single sibship consanguineous families that have enough informativeness to reach an lod score of 3 are quite rare, even in countries with high consanguinity and large sibship size (one requires, in a first cousin marriage at least 3 affected sibs, or 2 affected and X5 nonaffected). More distant consanguinity is potentially more informative, but the risk is then to miss the relevant homozygous region, if the genome scan is performed at the usual density of one marker every 10 cM.…”

“…24 This gene, like BBS6, was thus not included in the lists of candidates obtained by comparative genomic approaches. 5,6,8,13,47 One can wonder why BBS10, which is with BBS1 the most frequently mutated BBS gene, at least in patients of European origin, escaped positional cloning until now, whereas genes that are mutated in a much smaller proportion of patients have been identified. The absence of BBS10 on lists of candidate genes might explain why it was not detected by Nishimura et al 13 who had two small consanguineous families consistent with the chromosome 12 localization of BBS10.…”

“…1,2 Positional cloning studies complemented recently by comparative genomic approaches have proven the extensive genetic heterogeneity of this autosomal-recessive condition. Nine genes have been identified before 2006 BBS1 (11q13); 3 BBS2 (16q21); 4 BBS3/ARL6 (3p12 -13); 5,6 BBS4 (15q22.3 -q23); 7 BBS5 (2q31); 8 BBS6/MKKS (20 p12); 9,10 BBS7 (4q27); 11 BBS8/ TTC8(14q32.11) 12 and BBS9 /PTHB1 (7p14). 13 Mutations in these genes account only for about 50% of BBS patients, suggesting that there are several more genes to be found for this condition.…”

Caenorhabditis elegans as a model for obesity research

“…41 Likewise, the C. elegans ARL6 homolog arl-6 is restricted to ciliated cells and is involved in ciliary transport. 42 The reason for the association of obesity with BBS has been elusive. However, it has been discovered that tub-1 is a single gene mutation, which induces obesity that crosses species and results in changes consistent with Tubby mice, human BBS and C. elegans through increases in fat storage and decreases in fat oxidation.…”

BBS8 is rarely mutated in a cohort of 128 Bardet–Biedl syndrome families

“…To date, eight BBS genes have been identified: BBS1 (11q13; Mykytyn et al 2002), BBS2 (16q21; Nishimura et al 2001);BBS3 (3p13-p12;Fan et al 2004;Chiang et al 2004), BBS4 (15q22.3-q23;Mykytyn et al 2001); BBS5 (2q31; Li et al 2004);BBS6 (20 p12;Slaovotinek et al 2000;Katsanis et al 2000), BBS7 (4q27; Badano et al 2003) and BBS8 (14q32.11; Ansley et al 2003).…”