Identification of a novel Bardet-Biedl syndrome protein, BBS7, that shares structural features with BBS1 and BBS2

Abstract

Bardet-Biedl syndrome (BBS) is a genetically heterogeneous disorder, the primary features of which include obesity, retinal dystrophy, polydactyly, hypogenitalism, learning difficulties, and renal malformations. Conventional linkage and positional cloning have led to the mapping of six BBS loci in the human genome, four of which (BBS1, BBS2, BBS4, and BBS6) have been cloned. Despite these advances, the protein sequences of the known BBS genes have provided little or no insight into their function. To delineate functionally important regions in BBS2, we performed phylogenetic and genomic studies in which we used the human and zebrafish BBS2 peptide sequences to search dbEST and the translation of the draft human genome. We identified two novel genes that we initially named "BBS2L1" and "BBS2L2" and that exhibit modest similarity with two discrete, overlapping regions of BBS2. In the present study, we demonstrate that BBS2L1 mutations cause BBS, thereby defining a novel locus for this syndrome, BBS7, whereas BBS2L2 has been shown independently to be BBS1. The motif-based identification of a novel BBS locus has enabled us to define a potential functional domain that is present in three of the five known BBS proteins and, therefore, is likely to be important in the pathogenesis of this complex syndrome.

Figure 1

Mapping and genomic structure of BBS2L1 and its murine ortholog. The position and size of each exon of the human and mouse transcript are shown (black boxes). In the human transcript, two different splice variants of exon 18 are shown. In the short isoform, the 124-bp–long exon 18 is contiguous with an additional 417-bp 3′ UTR (box with diagonal dashes), whereas, in the long isoform, this exon is spliced with a new 726-bp exon that encodes an additional 44 residues and a different 3′ UTR (dotted box). We subsequently termed this locus “BBS7” because of the presence of pathogenic mutations that segregated with BBS.

Figure 2

Expression profile of BBS7. A, Northern blotting of a human 3′ UTR probe showing an ∼2.7-kb species expressed in low-to-moderate levels in multiple human tissues. B, RT-PCR analysis of the two alternatively spliced isoforms in 17 adult and 8 fetal tissues. The short isoform (sBBS2L1) is ubiquitously expressed, whereas the long isoform (lBBS2L1) is expressed in numerous, but not all, tissues.

Figure 3

Mutations in BBS7 associated with BBS. A, Recessive inheritance in BBS7. The sole affected individual, –05, in consanguineous family KK26 harbors a homozygous 4-bp deletion in exon 7 that is predicted to lead to premature termination in exon 9 (K237fsX296), thereby abolishing ∼64% of the wild-type protein. Likewise, in two unrelated pedigrees (AR25 and AR395), all patients are homozygous for an H323R allele. Individuals unavailable for collection are shown with dashes. B, Complex inheritance in BBS7. Consanguineous pedigree AR69 harbors a homozygous T211I missense mutation in BBS7 and a heterozygous E234K mutation in BBS1. All patients carry all three mutant alleles, and genetic analyses across BBS1 indicate that this family is unlikely to carry other mutations at that locus, since affected individuals –04 and –05 have inherited different paternal chromosomes. Note that asymptomatic sibling –03 has identical BBS1 haplotypes to patient –04, including the heterozygous E234K mutation.

Figure 4

A potential domain shared between multiple BBS proteins. A, Schematic representation of the region of overlap between the three proteins. The region of homology between BBS1 and BBS2 is shown in orange; the homology between BBS2 and BBS7 only is shown in red, whereas the area of overlap among all three proteins is depicted in light blue. Numbers correspond to amino acid residues; the predicted β-propeller, as well as the mutations in BBS1 and BBS7, are also shown (scale is approximate). Note that the potential BBS1 triallelic mutation E234K maps in a region shared between BBS1 and BBS2. B, Local alignment of BBS2 orthologs with homologous regions in BBS7 and BBS1. The positions of the BBS7 mutations are also shown; the threonine 211 residue is completely conserved and forms part of this potential domain, whereas the 4-bp deletion at the lysine 237 residue of BBS7 abolishes most of the region shared uniquely with BBS2. hs = Homo sapiens; mm = Mus musculus; rn = Rattus norvegicus; dr = Danio rerio; and ce = Caenorhabditis elegans.