Alternative titles; symbolsARF-LIKE 6BBS3 GENEHGNC Approved Gene Symbol: ARL6Cytogenetic location: 3q11.2 Genomic coordinates (GRCh38): 3:97,762,580-97,812,5...
Alternative titles; symbols
HGNC Approved Gene Symbol: ARL6
Cytogenetic location: 3q11.2 Genomic coordinates (GRCh38): 3:97,762,580-97,812,584 (from NCBI)
ADP-ribosylation factor (ARF)-like-6 (ARL6) is a member of a subgroup of the ARF family, proteins that regulate diverse cellular functions, including regulation of intracellular traffic. The ARF group of proteins are part of the Ras superfamily, which is subdivided into ARF and ARF-like (ARL) subgroups (Pasqualato et al., 2002).
▼ Cloning and Expression
Ingley et al. (1999) cloned mouse Arl6. The deduced 186-amino acid protein has a calculated molecular mass of 20.9 kD. Arl6 contains conserved features of the ARF family, including an N-terminal myristoylation site followed by a hydrophobic alpha helix and a GTP-binding site. The 3-prime UTR of the Arl6 transcript contains 10 copies of the AT3 motif implicated in mRNA destabilization. Northern blot analysis of several mouse tissues detected a 1.3-kb transcript expressed at highest levels in brain and kidney, with low expression in other tissues. Subcellular fractionation and confocal microscopy of Arl6-transfected COS cells indicated that most of the Arl6 protein was cytosolic. The amount of Arl6 associated with membranes increased in the presence of a nonhydrolyzable GTP analog.
By searching for genes in the region of chromosome 3 associated with Bardet-Biedl syndrome-3 (BBS3; 600151), Chiang et al. (2004) identified ARL6. The deduced protein contains 186 amino acids. Database analysis revealed ARL6 orthologs in a wide range of species from mammals to invertebrates, yeast, and plants.
By EST database analysis, Pretorius et al. (2010) identified a BBS3 splice variant that they called BBS3-long (BBS3L). This EST was found only in retina or whole eye libraries. The BBS3L transcript includes an additional 13-bp coding exon near its 3-prime end that shifts the ORF. The deduced 193-amino acid BBS3L protein has 15 unique C-terminal amino acids that replace the 8 C-terminal amino acids of the 186-amino acid BBS3 protein. Immunohistochemical analysis of mouse and human retina using an antibody that recognized both BBS3 and BBS3L revealed expression in ganglion cell layer, nerve fiber layer, and photoreceptor cells.
Using a high-resolution immunohistochemical technique with human retinal pigment epithelium (RPE) cells, Jin et al. (2010) localized ARL6 to small punctae flanking the microtubule axoneme in cilia. ARL6 colocalized at the ciliary membrane with a stable protein complex containing BBS1 (209901) and BBIP10 (613605).
▼ Gene Function
Using a protein pull-down assay with homogenized bovine retina, Jin et al. (2010) showed that ARL6 bound a ciliary membrane protein complex containing 7 BBS proteins and BBIP10. Depletion of ARL6 in human RPE cells did not affect assembly of the complex, but it blocked its localization to cilia. Targeting of ARL6 and the protein complex to cilia required GTP binding by ARL6, but not ARL6 GTPase activity. When in the GTP-bound form, the N-terminal amphipathic helix of ARL6 bound brain lipid liposomes and recruited the BBS protein complex. Upon recruitment, the complex appeared to polymerize into an electron-dense planar coat, and it functioned in lateral transport of test cargo proteins to ciliary membranes.
▼ Gene Structure
Chiang et al. (2004) determined that the ARL6 gene contains 9 exons and spans 33.8 kb. The first 3 exons are noncoding. Pretorius et al. (2010) identified an additional exon near the 3-prime end of the ARL6 gene that is subject to alternative splicing.
The ARL6 gene maps within the BBS3 (see 600151) critical region, 3p12-q13 (Fan et al., 2004).
Gross (2014) mapped the ARL6 gene to chromosome 3q11.2 based on an alignment of the ARL6 sequence (GenBank BC024239) with the genomic sequence (GRCh37).
▼ Molecular Genetics
Bardet-Biedl Syndrome 3
To identify the gene mutant in Bardet-Biedl syndrome-3 (BBS3; 600151), which had been mapped in a large Israeli Bedouin kindred to chromosome 3p13-p12 (Sheffield et al., 1994), Chiang et al. (2004) performed comparative genomic analysis to prioritize BBS candidate genes for mutation screening. Known BBS proteins were compared with the translated genomes of model organisms to identify a subset of organisms in which these proteins were conserved. By including multiple organisms with relatively small genome sizes in the analysis, the number of candidate genes was reduced, and a few genes mapping to the BBS3 interval emerged as the best candidates for the disorder. One of these genes, ARL6, was found to contain a homozygous arg122-to-ter mutation (R122X; 608845.0001) that segregated completely with the disease in the Israeli Bedouin kindred in which the BBS3 locus was originally mapped, thus identifying ARL6 as the BBS3 gene.
Fan et al. (2004) identified ARL6 as a likely candidate for the BBS3 gene because of its position in the critical linkage mapping region for type 3 BBS and particularly because of its homology to a C. elegans gene containing an X-box sequence in its promoter region characteristic of genes known to be strictly expressed in ciliated cells. They found mutations in the ARL6 gene that segregated with BBS3 in 4 independent families. Comparison of amino acid sequences of ARL6 from divergent organisms showed that gly169, mutated in 1 family (608845.0002), is invariant. The residues in the other 2 mutations, thr31 (608845.0003, 608845.0005) and leu170 (608845.0004), are highly conserved. Fan et al. (2004) reported another family in which heterozygous mutation in BBS3 (608845.0002) appeared to modify the expression of a homozygous mutation in the BBS1 gene, M390R (209901.0001). The sister with the additional mutation in BBS3 was more severely affected than the other.
Bardet-Biedl syndrome is thought to result largely from ciliary dysfunction, because loss-of-function mutations in the genes of C. elegans homologous to BBS7 (607590) and BBS8 (608132) compromise cilia structure and function, and RNA interference of Chlamydomonas BBS5 (603650) results in the loss of flagella. 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. Fan et al. (2004) hypothesized that the C. elegans ortholog of the human BBS3 gene would also contain this regulatory element, which would allow them to identify candidates from among the more than 90 genes that map to the BBS3 critical interval. One of 3 genes containing the X box in their promoters that determine exclusive expression in cells with cilia was ARL6, making it a good candidate for the BBS3 gene. Fan et al. (2004) indeed found mutations in ARL6 segregating with BBS in 4 independent families.
In a study of 7 Saudi Arabian BBS families, Safieh et al. (2010) demonstrated that homozygosity mapping was an efficient approach to identifying causative mutations, because it allowed them to sequence only 1 gene per family and find 7 novel mutations, respectively: 3 in the BBS1 gene, 3 in the BBS3 gene, and 1 in the BBS4 gene. Six of the families displayed the typical constellation of findings for BBS, which varied in frequency between families but were highly consistent within families, suggesting that modifiers appear to play only a minor role in the expressivity of BBS. In the remaining family, previously reported by Aldahmesh et al. (2009), a homozygous BBS3 mutation (608845.0006) segregated with nonsyndromic autosomal recessive RP (RP55; 613575). Compared with earlier reports, Safieh et al. (2010) stated that their data were consistent with a trend towards milder severity in patients with BBS3 mutations, since all cases of documented normal male fertility or lack of cognitive impairment belonged to this category. In addition, atopy appeared to be a common clinical feature that was not restricted to a specific genotype, and none of their patients reported a history of hyposmia, suggesting that this is an uncommon finding.
Retinitis Pigmentosa 55
In a consanguineous Saudi Arabian family segregating autosomal recessive nonsyndromic retinitis pigmentosa (RP55; 613575) but with no other recognizable primary or secondary features of BBS, Aldahmesh et al. (2009) identified homozygosity for a missense mutation in the ARL6 gene (608845.0006), which was not found in 192 Saudi controls and in more than 50 additional RP patients, suggesting that ARL6 mutation is not a frequent cause of nonsyndromic RP.
In 3 Mexican sibs with RP, born of an endogamous marriage, Zenteno et al. (2020) identified homozygosity for a 1-bp duplication in the ARL6 gene (608845.0007).
▼ Animal Model
Using zebrafish embryos, Pretorius et al. (2010) showed that morpholino-based knockdown of bbs3, but not bbs3l, resulted in a BBS-like phenotype, including defects to the ciliated Kupffer vesicle and delayed retrograde melanosome transport. Bbs3l-specific knockdown impaired visual function. The overall architecture of the mutant retina appeared normal, but it showed mislocalization of the photopigment green cone opsin (OPN1MW; 300821). Expression of human BBS3L, but not BBS3, rescued both the vision defect and green opsin mislocalization in zebrafish retina. Bbs3l -/- mice lacked the obesity seen in Bbs -/- mice, and they showed disrupted photoreceptor inner segment architecture. The defect in Bbs3l -/- mice appeared milder than that seen in Bbs3l-knockdown zebrafish, suggesting that Bbs3 may partially compensate for loss of Bbs3l in mice.
▼ ALLELIC VARIANTS ( 7 Selected Examples):
.0001 BARDET-BIEDL SYNDROME 3
In affected members of the Israeli Bedouin family with Bardet-Biedl syndrome-3 (BBS3; 600151) originally described by Kwitek-Black et al. (1993) (family 2) and linked to chromosome 3 by Sheffield et al. (1994), Chiang et al. (2004) identified a homozygous C-to-T transition in exon 7 of the ARL6 gene, resulting in an arg122-to-ter (R122X) mutation with truncation of the protein from 186 to 121 amino acids. The R122X mutation was not found in 100 Arab control individuals from the Middle East or in 90 additional control individuals of diverse ethnicity.
.0002 BARDET-BIEDL SYNDROME 3
BARDET-BIEDL SYNDROME 1, MODIFIER OF, INCLUDED
In an affected individual from a well characterized family from Newfoundland with BBS3 (600151) (Young et al., 1998), Fan et al. (2004) found a homozygous 859G-C transversion in exon 8 of the ARL6 gene, resulting in a gly-to-ala substitution at residue 169 (G169A). Fan et al. (2004) also found the G169A mutation in heterozygous state in 1 of 2 sisters homozygous for the met390-to-arg (M390R) mutation of the BBS1 gene (209901.0001). The sister with the additional mutation in BBS3 was more severely affected than the sister without the mutation.
.0003 BARDET-BIEDL SYNDROME 3
In affected members of a Saudi Arabian family with BBS3 (600151), Fan et al. (2004) identified a homozygous missense mutation, thr31 to met (T31M), resulting from a C-to-T transition at nucleotide 445 in exon 3 of the ARL6 gene.
.0004 BARDET-BIEDL SYNDROME 3
In affected members of a North American family of suspected consanguinity with BBS3 (600151), Fan et al. (2004) identified a homozygous transversion in exon 8 of the ARL6 gene, 862T-G, resulting in a leu170-to-trp (L170W) mutation.
.0005 BARDET-BIEDL SYNDROME 3
In affected members of a consanguineous Irish family with BBS3 (600151), Fan et al. (2004) identified a homozygous 445C-G transversion in exon 3 of the ARL6 gene that resulted in a thr31-to-arg (T31R) mutation.
.0006 RETINITIS PIGMENTOSA 55
In affected sibs from a consanguineous Saudi Arabian family (DGU-F15) segregating autosomal recessive nonsyndromic retinitis pigmentosa (RP55; 613575), Aldahmesh et al. (2009) identified homozygosity for a 266C-T transition in the ARL6 gene, resulting in an ala89-to-val (A89V) substitution at a highly conserved residue. The mutation was not found in 192 Saudi controls. Thorough reexamination of the 4 affected sibs by Safieh et al. (2010) revealed no recognizable primary or secondary features of Bardet-Biedl syndrome (600151).
Pretorius et al. (2011) found that alanine-89 is highly conserved in vertebrates and that the A89V variant is expressed in both BBS3 and BBS3L isoforms. To analyze the function of A89V, Pretorius et al. (2011) used knockdown of bbs3 embryos coupled with RNA rescue in zebrafish. They found that A89V can rescue melanosome transport defects but not the vision impairment in these embryos.
.0007 RETINITIS PIGMENTOSA 55
ARL6, 1-BP DUP, 373A
In a Mexican patient (patient 1521) and 2 affected sibs with nonsyndromic retinitis pigmentosa (RP55; 613575), Zenteno et al. (2020) identified homozygosity for a 1-bp duplication (c.373dupA, NM_032146.4) in the ARL6 gene, causing a frameshift predicted to result in a premature termination codon (Ile125AsnfsTer7). Sanger sequencing confirmed familial segregation of the variant with disease.