PURPOSE: Bardet-Biedl syndrome is a complex ciliopathy that usually manifests with some form of retinal degeneration, amongst other ciliary-related deficiencies. One of the genetic causes of this syndrome results from a defect in Bardet-Biedl Syndrome 5 (BBS5) p
rotein. BBS5 is one component of the BBSome, a complex of proteins that regulates the protein composition in cilia. In this study, we identify a smaller molecular mass form of BBS5 as a variant formed by alternative splicing and show that expression of this splice variant is restricted to the retina. METHODS: Reverse transcription PCR from RNA was used to isolate and identify potential alternative transcripts of Bbs5. A peptide unique to the C-terminus of the BBS5 splice variant was synthesized and used to prepare antibodies that selectively recognized the BBS5 splice variant. These antibodies were used on immunoblots of tissue extracts to determine the extent of expression of the alternative transcript and on tissue slices to determine the localization of expressed protein. Pull-down of fluorescently labeled arrestin1 by immunoprecipitation of the BBS5 splice variant was performed to assess functional interaction between the two proteins. RESULTS: PCR from mouse retinal cDNA using Bbs5-specific primers amplified a unique cDNA that was shown to be a splice variant of BBS5 resulting from the use of cryptic splicing sites in Intron 7. The resulting transcript codes for a truncated form of the BBS5 protein with a unique 24 amino acid C-terminus, and predicted 26.5 kD molecular mass. PCR screening of RNA isolated from various ciliated tissues and immunoblots of protein extracts from these same tissues showed that this splice variant was expressed in retina, but not brain, heart, kidney, or testes. Quantitative PCR showed that the splice variant transcript is 8.9-fold (+/- 1.1-fold) less abundant than the full-length transcript. In the retina, the splice variant of BBS5 appears to be most abundant in the connecting cilium of photoreceptors, where BBS5 is also localized. Like BBS5, the binding of BBS5L to arrestin1 can be modulated by phosphorylation through protein kinase C. CONCLUSIONS: In this study we have identified a novel splice variant of BBS5 that appears to be expressed only in the retina. The BBS5 splice variant is expressed at approximately 10% of full-length BBS5 level. No unique functional or localization properties could be identified for the splice variant compared to BBS5.
Cilia and flagella are microtubule-based structures nucleated by modified centrioles termed basal bodies. These biochemically complex organelles have more than 250 and 150 polypeptides, respectively. To identify the proteins involved in ciliary and basal body biogenesis and function, we undertook a
comparative genomics approach that subtracted the nonflagellated proteome of Arabidopsis from the shared proteome of the ciliated/flagellated organisms Chlamydomonas and human. We identified 688 genes that are present exclusively in organisms with flagella and basal bodies and validated these data through a series of in silico, in vitro, and in vivo studies. We then applied this resource to the study of human ciliation disorders and have identified BBS5, a novel gene for Bardet-Biedl syndrome. We show that this novel protein localizes to basal bodies in mouse and C. elegans, is under the regulatory control of daf-19, and is necessary for the generation of both cilia and flagella.
PURPOSE: Bardet-Biedl syndrome (BBS) is rare in the Philippines and only limited information on the prevalent subtypes is available as yet. The purpose of this study is to present the clinical characteristics of two Filipino siblings presenting with mutations in BBS5
00;'>BBS5. PATIENTS AND METHODS: The Filipino female siblings, aged 11 and 14 years underwent comprehensive ophthalmologic evaluation. Fundus photography, macular optical coherence tomography (OCT) and electroretinography (ERG) were also obtained. Systemic workup was performed including radiographic imaging of limb defects, renal ultrasound, blood chemistry, and transvaginal ultrasound. Targeted Bardet-Biedl sequence analysis and deletion/duplication analysis were performed to determine potential pathogenic mutations. RESULTS: Both children had poor visual acuity with a myopic refraction. There was a pigmentary retinopathy with retinal pigment epithelium changes and attenuation of vessels without waxy disc pallor. Generalized macular thinning and undetectable ERG responses were recorded. Physical examination revealed obesity, facial anomalies, brachydactyly, postaxial polydactyly, and clinodactyly of fifth digits. Both patients displayed cognitive developmental delay and hypogonadism. Molecular analysis revealed novel compound heterozygous mutations in BBS5 with c.143-1 G > A (splice acceptor) and c.925_931del (p.Gln309ilefs*14), each inherited from one asymptomatic parent. CONCLUSION: These are probably the first reported BBS5 mutations causing Bardet-Biedl syndrome in the Philippines. Patients were managed by a multi-disciplinary team and the parents were counseled regarding the prognosis and additional complications associated with the syndrome.
Imani S, etal., Biosci Rep. 2019 Mar 28;39(3):BSR20181544. doi: 10.1042/BSR20181544. Print 2019 Mar 29.
Bardet-Biedl syndrome (BBS) is a rare genetically heterogeneous ciliopathy which accompanies retinitis pigmentosa (RP). However, the BBS5 mutation remains unclear in Iranians with BBS. The purpose of study is to evaluate genetic analyses of a BBS Iranian family
using targetted exome sequencing (TES). A male 11-year-old proband and three related family members were recruited. Biochemical tests, electrocardiography and visual acuity testing, such as funduscopic, fundus photography (FP), optical coherence tomography (OCT), and standard electroretinography, were conducted. Molecular analysis and high-throughput DNA sequence analysis were performed. The proband was diagnosed with possible BBS based on the presence of three primary features and two secondary features. The TES analysis of the proband with BBS resulted in the identification of a novel, homozygous splicing variant c. 208+2T>C of the BBS5 gene (NM_152384.2) in this Iranian BBS family. This variant was confirmed and was completely co-segregated with the disease in this family by Sanger sequencing. Thus, we report a novel, homozygous splicing site variant c.208+2T>C in the BBS5 gene for the first time in the Iranian family.
