Bardet-Biedl syndrome (BBS) is primarily an autosomal recessive ciliopathy characterized by progressive retinal degeneration, obesity, cognitive impairment, polydactyly, and kidney anomalies. The disorder is genetically heterogeneous, with 11 BBS genes identified to date, which account for ~70% of affected families. We have combined single-nucleotide-polymorphism array homozygosity mapping with in silico analysis to identify a new BBS gene, BBS12. Patients from two Gypsy families were homozygous and haploidentical in a 6-Mb region of chromosome 4q27. FLJ35630 was selected as a candidate gene, because it was predicted to encode a protein with similarity to members of the type II chaperonin superfamily, which includes BBS6 and BBS10. We found pathogenic mutations in both Gypsy families, as well as in 14 other families of various ethnic backgrounds, indicating that BBS12 accounts for approximately 5% of all BBS cases. BBS12 is vertebrate specific and, together with BBS6 and BBS10, defines a novel branch of the type II chaperonin superfamily. These three genes are characterized by unusually rapid evolution and are likely to perform ciliary functions specific to vertebrates that are important in the pathophysiology of the syndrome, and together they account for about one-third of the total BBS mutational load. Consistent with this notion, suppression of each family member in zebrafish yielded gastrulation-movement defects characteristic of other BBS morphants, whereas simultaneous suppression of all three members resulted in severely affected embryos, possibly hinting at partial functional redundancy within this protein family.

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We identified the residues that are important for the binding of ␣-dendrotoxin (␣DTX) to Kv1 potassium channels on rat brain synaptosomal membranes, using a mutational approach based on site-directed mutagenesis and chemical synthesis. Twenty-six of its 59 residues were individually substituted by alanine. Substitutions of Lys 5 and Leu 9 decreased affinity more than 1000-fold, and substitutions of Arg 3 , Arg 4 , Leu 6 , and Ile 8 by 5-30-fold. Substitution of Lys 5 by norleucine or ornithine also greatly altered the binding properties of ␣DTX. All of these analogs displayed similar circular dichroism spectra as compared with the wild-type ␣DTX, indicating that none of these substitutions affect the overall conformation of the toxin. Substitutions of Ser 38 and Arg 46 also reduced the affinity of the toxin but, in addition, modified its dichroic properties, suggesting that these two residues play a structural role. The other residues were excluded from the recognition site because their substitutions caused no significant affinity change. Venomous animals from four distinct phyla produce small toxic proteins that block a variety of Kv1 voltage-gated potassium channels. These are the scorpions (1), sea anemones (2-4), marine cone snails (5), and snakes (6 -8), which are arthropods, cnidarians, molluscs, and chordates, respectively. At least four different folds, the sizes of which range from approximately 30 to 60 residues, are associated with these different potassium channel-blocking toxins. These are (i) the ␣/␤-toxin fold, which is found in scorpion toxins, such as charybdotoxin (9); (ii) the fold that comprises two short helices and is only adopted by toxins from sea anemone toxins such as ShK (10) and BgK (11); (iii) the -conotoxin fold, which has three ␤-sheet strands and is adopted by -conotoxin from cone snails (12, 13); and (iv) the BPTI 1 -type fold (14), composed of two short helices and a two-stranded ␤-sheet, which is adopted by the snake dendrotoxins (15-17) and probably by the sea anemone kalicludines (4).Although structurally unrelated, the Kv1 channel-blocking toxins produced by scorpions, snakes, sea anemones, and snails all are likely to bind to the peptide loop between the membranespanning segments S5 and S6 of Kv1 channels (18 -25, 11). Therefore, all of these toxins may possess a functional surface that is complementary to this loop, an observation that raises the question as to how similar these surfaces are from one toxin to another. The answer to such a question may not only shed light on the evolution of these toxins but should also help characterize the surface by which Kv1 channels interact with these toxins. Mutational analyses have finely delineated the functional sites of scorpion toxins (26) and sea anemone toxins (11,25). Although the sea anemone and scorpion toxins are not structurally related, their functional sites share some similarities. They are all flat surfaces of comparable size (ϳ700 Å 2 ) with five functionally important residues, including a similar critical function...

The extensive genetic heterogeneity of Bardet-Biedl syndrome (BBS) is documented by the identification, by classical linkage analysis complemented recently by comparative genomic approaches, of nine genes (BBS1-9) that account cumulatively for about 50% of patients. The BBS genes appear implicated in cilia and basal body assembly or function. In order to find new BBS genes, we performed SNP homozygosity mapping analysis in an extended consanguineous family living in a small Lebanese village. This uncovered an unexpectedly complex pattern of mutations, and led us to identify a novel BBS gene (BBS10). In one sibship of the pedigree, a BBS2 homozygous mutation was identified, while in three other sibships, a homozygous missense mutation was identified in a gene encoding a vertebrate-specific chaperonine-like protein (BBS10). The single patient in the last sibship was a compound heterozygote for the above BBS10 mutation and another one in the same gene. Although triallelism (three deleterious alleles in the same patient) has been described in some BBS families, we have to date no evidence that this is the case in the present family. The analysis of this family challenged linkage analysis based on the expectation of a single locus and mutation. The very high informativeness of SNP arrays was instrumental in elucidating this case, which illustrates possible pitfalls of homozygosity mapping in extended families, and that can be explained by the rather high prevalence of heterozygous carriers of BBS mutations (estimated at one in 50 in Europeans).