The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform

Val C Sheffield - One of the best experts on this subject based on the ideXlab platform.

  • Ectopic expression of BBS1 rescues male infertility, but not retinal degeneration, in a BBS1 mouse model
    Gene Therapy, 2021
    Co-Authors: Matthew R. Cring, Charles Searby, Kai Wang, Kacie J. Meyer, Adam Hedberg-buenz, Michael Cave, Michael G. Anderson, Val C Sheffield
    Abstract:

    Bardet–Biedl syndrome (BBS) is a rare ciliopathy for which there are no current effective treatments. BBS is a genetically heterogeneous disease, though the M390R mutation in BBS1 is involved in ~25% of all genetic diagnoses of BBS. The principle features of BBS include retinal degeneration, obesity, male infertility, polydactyly, intellectual disability, and renal abnormalities. Patients with mutations in BBS genes often present with night blindness within the first decade of life, which progresses to complete blindness. This is due to progressive loss of photoreceptor cells. Male infertility is caused by a lack of spermatozoa flagella, rendering them immobile. In this study, we have crossed the wild-type human BBS1 gene, driven by the CAG promoter, onto the BBS1 ^ M390R/M390R mouse model to determine if ectopic expression of BBS1 rescues male infertility and retinal degeneration. qRT-PCR indicates that the BBS1 transgene is expressed in multiple tissues throughout the mouse, with the highest expression seen in the testes, and much lower expression in the eye and hypothalamus. Immunohistochemistry of the transgene in the eye showed little if any expression in the photoreceptor outer nuclear layer. When male BBS1 ^ M30R/M390R ;BBS1 ^ TG+ mice are housed with WT females, they are able to sire offspring, indicating that the male infertility phenotype of BBS is rescued by the transgene. Using electroretinography (ERGs) to measure retinal function and optical coherence tomography to measure retinal thickness, we show that the transgene does not confer protection against retinal degeneration in BBS1 ^ M300R/M390R ;BBS1 ^ TG+ mice. The results of this study indicate that the male infertility aspect of BBS is an attractive target for gene therapy.

  • gene therapeutic reversal of peripheral olfactory impairment in bardet biedl syndrome
    Molecular Therapy, 2017
    Co-Authors: Corey L Williams, Darryl Y Nishimura, Cedric Uytingco, Warren W Green, Jeremy C Mcintyre, Kirill Ukhanov, Arthur D Zimmerman, Dana T Shively, Lian Zhang, Val C Sheffield
    Abstract:

    Olfactory dysfunction is a pervasive but underappreciated health concern that affects personal safety and quality of life. Patients with olfactory dysfunctions have limited therapeutic options, particularly those involving congenital diseases. Bardet-Biedl syndrome (BBS) is one such disorder, where olfactory loss and other symptoms manifest from defective cilium morphology and/or function in various cell types/tissues. Olfactory sensory neurons (OSNs) of BBS mutant mice lack the capacity to build/maintain cilia, rendering the cells incapable of odor detection. Here we examined OSN cilium defects in BBS1 mutant mice and assessed the utility of gene therapy to restore ciliation and function in young and adult mice. BBS1 mutant mice possessed short residual OSN cilia in which BBSome protein trafficking and odorant detection were defective. Gene therapy with an adenovirus-delivered wild-type BBS1 gene restored OSN ciliation, corrected BBSome cilium trafficking defects, and returned acute odor responses. Finally, using clinically approved AAV serotypes, we demonstrate, for the first time, the capacity of AAVs to restore ciliation and odor detection in OSNs of BBS1 mutants. Together, our data demonstrate that OSN ciliogenesis can be promoted in differentiated cells of young and adult BBS1 mutants and highlight the potential of gene therapy as a viable restorative treatment for congenital olfactory disorders.

  • BBS1 protein interacts with the LRb and regulates its trafficking to the cells surface.
    2016
    Co-Authors: Deng Fu Guo, Darryl Nishimura, Qihong Zhang, Val C Sheffield, Donald A Morgan, Daniel R. Thedens, Huxing Cui, Justin L. Grobe, Kamal Rahmouni
    Abstract:

    (A) Interaction between the BBS1 protein and the endogenous LRb in brain lysate. Ability of Flag-tagged BBS1 and LRb to pull down each other by co-immunoprecipitation assays using brain lysates of transgenic (Tg) mice expressing a Flag-BBS1 protein. IP: immunoprecipitation, IB: immunoblot (representative of n = 3, 1 male and 2 females). (B–C) shRNA-mediated silencing of BBS1 reduces the surface levels of Flag-tagged LRb in a dose-dependent manner in HEK 293 cells transiently co-transfected with Flag-LRb and BBS1-shRNA plasmids. This effect is specific as the surface expression of HA-tagged LRa (in separate experiments) and endogenous transferrin receptor (TrR, in same experiments as the Flag-LRb) were not altered (n = 6 per group). (D-E) Silencing of BBS2, another BBSome protein, lowers the surface levels of Flag-LRb, but not HA-LRa or endogenous TrR in HEK 293 cells (n = 6 per group). (F–G) Blockade of Rab8a using a dominant negative (DN) form decreases the surface expression of Flag-LRb, HA-LRa and endogenous TrR in HEK 293 cells (n = 6 per group). (H–I) siRNA-mediated silencing of Ift88 had no effect on the surface expression of Flag-LRb or endogenous TrR in HEK 293 cells (n = 8 per group). Data are means ± SEM, *P< 0.05 vs. control group.

  • Mice lacking the BBS1 gene in the nervous system develop obesity.
    2016
    Co-Authors: Deng Fu Guo, Darryl Nishimura, Qihong Zhang, Val C Sheffield, Donald A Morgan, Daniel R. Thedens, Huxing Cui, Justin L. Grobe, Kamal Rahmouni
    Abstract:

    (A-B) Selectivity of BBS1 gene deletion to the nervous system in NestinCre/BBS1fl/fl mice as determined by RT-PCR (A) and quantitative RT-PCR (B, n = 1 male and 2 females in group). BBS1 expression was absent in the brain (hypothalamus [Hypoth], cortex, hippocampus [Hippo] and brainstem [Brsm]) of NestinCre/BBS1fl/fl mice (Cre+) relative to controls (Cre-). However, BBS1 expression was not altered in white adipose tissue (WAT), liver, kidney and skeletal muscle (Sk Mus). (C) Analysis of the BBSome complex in the brain and testes of Nestincre/BBS1fl/fl mice. Inability of BBS2 to pull down BBS9 in the brain, but not in the testes, of NestinCre/BBS1fl/fl mice (Cre+) relative to controls (Cre-). (D) Weekly body weights of Nestincre/BBS1fl/fl and control mice (n = 12 males and 12 females for controls; 6 males and 6 females for Nestincre/BBS1fl/fl mice). (E-F) Comparison of body composition between Nestincre/BBS1fl/fl and control mice. Shown are coronal (top) and axial abdominal (bottom) representative MRI sections (E) and total fat mass and lean mass (F) of 25 weeks old control and Nestincre/BBS1fl/fl mice (n = 6 males and 6 females in each group). (G) Cumulative food intake of Nestincre/BBS1fl/fl and littermate controls (n = 6 males and 6 females for controls, and 6 males and 8 females for Nestincre/BBS1fl/fl mice). (H) Hypothalamic mRNA levels of AgRP, NPY and POMC in Nestincre/BBS1fl/fl and control mice (n = 3 males and 3 females in each group). Data are means ± SEM, *P< 0.05 vs control group.

  • abstract 083 metabolic and cardiovascular effects of BBS1 ablation from the pomc containing neurons
    Hypertension, 2015
    Co-Authors: Deng F. Guo, Val C Sheffield, Donald A Morgan, Justin L. Grobe, Kamal Rahmouni
    Abstract:

    Bardet-Biedl syndrome (BBS) is a pleiotropic autosomal recessive disorder associated with several features including obesity and hypertension. Deletion of Bbs genes globally, in the nervous system or in the leptin receptor-expressing cells recapitulated many of the BBS phenotypes including obesity and hypertension. Here, we assessed the effect of ablating the BBS1 gene from the neurons expressing proopiomelanocortin (POMC) neurons. Breeding BBS1flox mice with POMCCre mice created mice deficient in BBS1 gene only in the POMC-positive neurons (visualized by tdTomato expression). Importantly, POMCCre/BBS1fl/fl mice display an obesity phenotype as indicated by the increased (P<0.05) body weight (48.6±1.7 vs 34.9±0.8 g in controls) and fat pads (3.3±0.3 vs. 0.7±1.1 g for inguinal fat, 2.1±0.2 vs. 0.4±0.05 g for perirenal fat, 3.3±0.3 vs. 0.8±0.3 g for reproduction fat and 0.4±0.02 vs. 0.2±0.02 g for brow fat) associated with increased (P<0.05) food intake (3.79.±0.14 vs. 2.97±0.15 g in controls) in 25 weeks old mice. POMCCre/BBS1fl/fl mice displayed decreased (P<0.05) O2consumption (2.6±0.03 vs. 3±0.04 mL O2/100g/min in controls) and heat production (8.0±0.09 vs. 9.2±0.13 kcal/kg/hr in controls). These results indicate that hyperphagia and decreased energy expenditure contribute to the development of obesity in POMCCre/BBS1fl/fl mice. Next, we assessed the consequence on arterial pressure (AP) and sympathetic nerve activity (SNA) of ablating the BBS1 gene from POMC neurons. Interestingly, deletion of the BBS1 gene in POMC neurons did not recapitulate the hypertension phenotype of BBS as indicated by the slight, but not significant increase in mean AP (116±4.7 vs 109±6.1 mmHg in controls). However, conscious renal SNA was significantly higher in POMCCre/BBS1fl/fl mice relative to controls (132.4±11 vs 74.3±4.2 spikes/sec, P<0.05). Finally, the depressor effect of ganglionic blockade (hexamethonium) was exaggerated in POMCCre/BBS1fl/fl mice (-68.6±4.4 vs -45.2±9.4 mmHg in control, P=0.017). These findings demonstrate that BBS1 gene in the POMC neurons is critical for energy homeostasis, but not for arterial pressure regulation.

Charles Searby - One of the best experts on this subject based on the ideXlab platform.

  • Ectopic expression of BBS1 rescues male infertility, but not retinal degeneration, in a BBS1 mouse model
    Gene Therapy, 2021
    Co-Authors: Matthew R. Cring, Charles Searby, Kai Wang, Kacie J. Meyer, Adam Hedberg-buenz, Michael Cave, Michael G. Anderson, Val C Sheffield
    Abstract:

    Bardet–Biedl syndrome (BBS) is a rare ciliopathy for which there are no current effective treatments. BBS is a genetically heterogeneous disease, though the M390R mutation in BBS1 is involved in ~25% of all genetic diagnoses of BBS. The principle features of BBS include retinal degeneration, obesity, male infertility, polydactyly, intellectual disability, and renal abnormalities. Patients with mutations in BBS genes often present with night blindness within the first decade of life, which progresses to complete blindness. This is due to progressive loss of photoreceptor cells. Male infertility is caused by a lack of spermatozoa flagella, rendering them immobile. In this study, we have crossed the wild-type human BBS1 gene, driven by the CAG promoter, onto the BBS1 ^ M390R/M390R mouse model to determine if ectopic expression of BBS1 rescues male infertility and retinal degeneration. qRT-PCR indicates that the BBS1 transgene is expressed in multiple tissues throughout the mouse, with the highest expression seen in the testes, and much lower expression in the eye and hypothalamus. Immunohistochemistry of the transgene in the eye showed little if any expression in the photoreceptor outer nuclear layer. When male BBS1 ^ M30R/M390R ;BBS1 ^ TG+ mice are housed with WT females, they are able to sire offspring, indicating that the male infertility phenotype of BBS is rescued by the transgene. Using electroretinography (ERGs) to measure retinal function and optical coherence tomography to measure retinal thickness, we show that the transgene does not confer protection against retinal degeneration in BBS1 ^ M300R/M390R ;BBS1 ^ TG+ mice. The results of this study indicate that the male infertility aspect of BBS is an attractive target for gene therapy.

  • abstract 016 BBS1 gene deletion from the leptin receptor neurons causes obesity and hypertension in mice
    Hypertension, 2014
    Co-Authors: Deng F. Guo, Charles Searby, Val C Sheffield, Donald A Morgan, Darryl Y Nishimura, Justin L. Grobe, Kamal Rahmouni
    Abstract:

    Bardet-Biedl syndrome (BBS) is a pleiotropic autosomal recessive disorder associated with several features including obesity and hypertension. Deletion of BBS genes globally or in the nervous system recapitulated many of the BBS phenotypes including obesity and hypertension. Here, we assessed the effect of ablating the BBS1 gene from the neurons expressing the long signaling form of the leptin receptor (LepRb). Breeding BBS1flox with LepRbCre mice created mice deficient in the BBS1 gene only in LepRb-positive neurons (visualized by tdTomato expression) as indicated by loss of leptin activation of Stat3. Importantly, BBS1flox/LepRCre mice display an obesity phenotype as indicated by the increased (P<0.05) body weight (39±2 vs. 30±1 g in controls) and fat mass measured by MRI (14±3 vs. 4±1 g in controls) associated with increased (P<0.05) food intake (3.4±0.1 vs. 2.9±0.1 g in controls) in 25 weeks old mice. However, body weight and fat pads of pair-fed LRbCre/BBS1fl/fl mice remained significantly elevated compared to controls suggesting that LRbCre/BBS1fl/fl mice have reduced energy expenditure. Consistent with this possibility, LRbCre/BBS1fl/fl mice displayed decreased (P<0.05) O2 consumption (2.6±0.1 vs. 3.1±0.1 mL/100g/min in controls) and heat production (8.1±0.3 vs. 9.6±0.3 kcal/kg/h in controls). These results indicate that hyperphagia and decreased energy expenditure contribute to the development of obesity in BBS1flox/LepRCre mice. Next, we assessed the effect on arterial pressure (AP) and sympathetic nerve activity (SNA) of ablating the BBS1 gene from the LepR-containing neurons. Interestingly, deletion of the BBS1 gene in LepR neurons recapitulates the hypertension phenotype of BBS as indicated by elevated mean AP (125±4 vs 109±3 mmHg in controls, P=0.03). Conscious renal SNA was also elevated in LRbCre/BBS1fl/fl mice relative to controls (97±8 vs 62±10 spikes/sec, P<0.05). Finally, the depressor effect of ganglionic blockade (hexamethonium) was exaggerated in BBS1flox/LepRCre mice (-57±5 vs -38±5 mmHg in control, P=0.01). These findings demonstrate that the BBS1 gene in LepR neurons is critical for energy homeostasis and arterial pressure regulation.

  • The centriolar satellite protein AZI1 interacts with BBS4 and regulates ciliary trafficking of the BBSome.
    PLoS genetics, 2014
    Co-Authors: Xitiz Chamling, Charles Searby, Diane C Slusarski, Seongjin Seo, Gunhee Kim, Val C Sheffield
    Abstract:

    Bardet-Biedl syndrome (BBS) is a well-known ciliopathy with mutations reported in 18 different genes. Most of the protein products of the BBS genes localize at or near the primary cilium and the centrosome. Near the centrosome, BBS proteins interact with centriolar satellite proteins, and the BBSome (a complex of seven BBS proteins) is believed to play a role in transporting ciliary membrane proteins. However, the precise mechanism by which BBSome ciliary trafficking activity is regulated is not fully understood. Here, we show that a centriolar satellite protein, AZI1 (also known as CEP131), interacts with the BBSome and regulates BBSome ciliary trafficking activity. Furthermore, we show that AZI1 interacts with the BBSome through BBS4. AZI1 is not involved in BBSome assembly, but accumulation of the BBSome in cilia is enhanced upon AZI1 depletion. Under conditions in which the BBSome does not normally enter cilia, such as in BBS3 or BBS5 depleted cells, knock down of AZI1 with siRNA restores BBSome trafficking to cilia. Finally, we show that azi1 knockdown in zebrafish embryos results in typical BBS phenotypes including Kupffer's vesicle abnormalities and melanosome transport delay. These findings associate AZI1 with the BBS pathway. Our findings provide further insight into the regulation of BBSome ciliary trafficking and identify AZI1 as a novel BBS candidate gene.

  • The Centriolar Satellite Protein AZI1 Interacts with BBS4 and Regulates Ciliary Trafficking of the BBSome
    2014
    Co-Authors: Xitiz Chamling, Charles Searby, Diane C Slusarski, Seongjin Seo, Gunhee Kim, Val C Sheffield
    Abstract:

    Bardet-Biedl syndrome (BBS) is a well-known ciliopathy with mutations reported in 18 different genes. Most of the protein products of the BBS genes localize at or near the primary cilium and the centrosome. Near the centrosome, BBS proteins interact with centriolar satellite proteins, and the BBSome (a complex of seven BBS proteins) is believed to play a role in transporting ciliary membrane proteins. However, the precise mechanism by which BBSome ciliary trafficking activity is regulated is not fully understood. Here, we show that a centriolar satellite protein, AZI1 (also known as CEP131), interacts with the BBSome and regulates BBSome ciliary trafficking activity. Furthermore, we show that AZI1 interacts with the BBSome through BBS4. AZI1 is not involved in BBSome assembly, but accumulation of the BBSome in cilia is enhanced upon AZI1 depletion. Under conditions in which the BBSome does not normally enter cilia, such as in BBS3 or BBS5 depleted cells, knock down of AZI1 with siRNA restores BBSome trafficking to cilia. Finally, we show that azi1 knockdown in zebrafish embryos results in typical BBS phenotypes including Kupffer’s vesicle abnormalities and melanosome transport delay. These findings associate AZI1 with the BBS pathway. Our findings provide further insight into the regulation of BBSome ciliar

  • AZI1 knockdown reduces ciliogenesis but increases ciliary localization of the BBSome.
    2014
    Co-Authors: Xitiz Chamling, Charles Searby, Diane C Slusarski, Seongjin Seo, Gunhee Kim, Val C Sheffield
    Abstract:

    A) RPE-1 cells were transfected with siRNA against BBS4, PCM1, and AZI1. Approximately 500 cells per sample were counted. B) AZI1 depletion increases the ciliary localization of BBS9 compared to the control knockdown. Red staining represents BBS9, and cilia are stained with acetylated α-tubulin. Nuclei are stained blue with DAPI. C) Graph showing a significant increase in cells with ciliary BBS9 upon AZI1 knockdown by different siRNAs. D) BBS8 (red) is used as a BBSome marker to confirm increased ciliary localization of the BBSome upon AZI1 knockdown. Cilia (green) in the insets of figures B and D are slightly shifted to show ciliary localization of BBS proteins. E) The graph shows a significant increase in the number of ciliated cells with BBS8 or BBS9 upon AZI1 knockdown, and decrease in ciliary BBS8 or BBS9 localization upon AZI1 overexpression. Approximately 250 ciliated cells were counted in control as well as AZI1 knockdown culture of RPE-1 cells. All data are presented in mean +/− SEM. Significance is calculated using Student's t-test for C and E, and one way ANOVA for A. P

Darryl Y Nishimura - One of the best experts on this subject based on the ideXlab platform.

  • gene therapeutic reversal of peripheral olfactory impairment in bardet biedl syndrome
    Molecular Therapy, 2017
    Co-Authors: Corey L Williams, Darryl Y Nishimura, Cedric Uytingco, Warren W Green, Jeremy C Mcintyre, Kirill Ukhanov, Arthur D Zimmerman, Dana T Shively, Lian Zhang, Val C Sheffield
    Abstract:

    Olfactory dysfunction is a pervasive but underappreciated health concern that affects personal safety and quality of life. Patients with olfactory dysfunctions have limited therapeutic options, particularly those involving congenital diseases. Bardet-Biedl syndrome (BBS) is one such disorder, where olfactory loss and other symptoms manifest from defective cilium morphology and/or function in various cell types/tissues. Olfactory sensory neurons (OSNs) of BBS mutant mice lack the capacity to build/maintain cilia, rendering the cells incapable of odor detection. Here we examined OSN cilium defects in BBS1 mutant mice and assessed the utility of gene therapy to restore ciliation and function in young and adult mice. BBS1 mutant mice possessed short residual OSN cilia in which BBSome protein trafficking and odorant detection were defective. Gene therapy with an adenovirus-delivered wild-type BBS1 gene restored OSN ciliation, corrected BBSome cilium trafficking defects, and returned acute odor responses. Finally, using clinically approved AAV serotypes, we demonstrate, for the first time, the capacity of AAVs to restore ciliation and odor detection in OSNs of BBS1 mutants. Together, our data demonstrate that OSN ciliogenesis can be promoted in differentiated cells of young and adult BBS1 mutants and highlight the potential of gene therapy as a viable restorative treatment for congenital olfactory disorders.

  • abstract 016 BBS1 gene deletion from the leptin receptor neurons causes obesity and hypertension in mice
    Hypertension, 2014
    Co-Authors: Deng F. Guo, Charles Searby, Val C Sheffield, Donald A Morgan, Darryl Y Nishimura, Justin L. Grobe, Kamal Rahmouni
    Abstract:

    Bardet-Biedl syndrome (BBS) is a pleiotropic autosomal recessive disorder associated with several features including obesity and hypertension. Deletion of BBS genes globally or in the nervous system recapitulated many of the BBS phenotypes including obesity and hypertension. Here, we assessed the effect of ablating the BBS1 gene from the neurons expressing the long signaling form of the leptin receptor (LepRb). Breeding BBS1flox with LepRbCre mice created mice deficient in the BBS1 gene only in LepRb-positive neurons (visualized by tdTomato expression) as indicated by loss of leptin activation of Stat3. Importantly, BBS1flox/LepRCre mice display an obesity phenotype as indicated by the increased (P<0.05) body weight (39±2 vs. 30±1 g in controls) and fat mass measured by MRI (14±3 vs. 4±1 g in controls) associated with increased (P<0.05) food intake (3.4±0.1 vs. 2.9±0.1 g in controls) in 25 weeks old mice. However, body weight and fat pads of pair-fed LRbCre/BBS1fl/fl mice remained significantly elevated compared to controls suggesting that LRbCre/BBS1fl/fl mice have reduced energy expenditure. Consistent with this possibility, LRbCre/BBS1fl/fl mice displayed decreased (P<0.05) O2 consumption (2.6±0.1 vs. 3.1±0.1 mL/100g/min in controls) and heat production (8.1±0.3 vs. 9.6±0.3 kcal/kg/h in controls). These results indicate that hyperphagia and decreased energy expenditure contribute to the development of obesity in BBS1flox/LepRCre mice. Next, we assessed the effect on arterial pressure (AP) and sympathetic nerve activity (SNA) of ablating the BBS1 gene from the LepR-containing neurons. Interestingly, deletion of the BBS1 gene in LepR neurons recapitulates the hypertension phenotype of BBS as indicated by elevated mean AP (125±4 vs 109±3 mmHg in controls, P=0.03). Conscious renal SNA was also elevated in LRbCre/BBS1fl/fl mice relative to controls (97±8 vs 62±10 spikes/sec, P<0.05). Finally, the depressor effect of ganglionic blockade (hexamethonium) was exaggerated in BBS1flox/LepRCre mice (-57±5 vs -38±5 mmHg in control, P=0.01). These findings demonstrate that the BBS1 gene in LepR neurons is critical for energy homeostasis and arterial pressure regulation.

  • bardet biedl syndrome 3 bbs3 knockout mouse model reveals common bbs associated phenotypes and bbs3 unique phenotypes
    Proceedings of the National Academy of Sciences of the United States of America, 2011
    Co-Authors: Qihong Zhang, Kevin Bugge, Charles Searby, Timothy W Vogel, Edwin M. Stone, Seongjin Seo, Donald A Morgan, Darryl Y Nishimura, Kamal Rahmouni, Val C Sheffield
    Abstract:

    Bardet-Biedl syndrome (BBS) is a heterogeneous disorder characterized by obesity, retinopathy, polydactyly, and congenital anomalies. The incidence of hypertension and diabetes are also increased in BBS patients. Mutation of 16 genes independently causes BBS, and seven BBS proteins form the BBSome that promotes ciliary membrane elongation. BBS3 (ARL6), an ADP ribosylation factor-like small GTPase, is not part of the BBSome complex. The in vivo function of BBS3 is largely unknown. Here we developed a Bbs3 knockout model and demonstrate that Bbs3−/− mice develop BBS-associated phenotypes, including retinal degeneration, male infertility, and increased body fat. Interestingly, Bbs3−/− mice develop some unique phenotypes not seen in other BBS knockout models: no overt obesity, severe hydrocephalus, and elevated blood pressure (shared by some but not all BBS gene knockout mice). We found that endogenous BBS3 and the BBSome physically interact and depend on each other for their ciliary localization. This finding explains the phenotypic similarity between Bbs3−/− mice and BBSome subunit knockout mice. Loss of Bbs3 does not affect BBSome formation but disrupts normal localization of melanin concentrating hormone receptor 1 to ciliary membranes and affects retrograde transport of Smoothened inside cilia. We also show that the endogenous BBSome and BBS3 associate with membranes and the membrane association of the BBSome and BBS3 are not interdependent. Differences between BBS mouse models suggest nonoverlapping functions to individual BBS protein.

  • bardet biedl syndrome in denmark report of 13 novel sequence variations in six genes
    Human Mutation, 2010
    Co-Authors: Tina Duelund Hjortshoj, Val C Sheffield, Darryl Y Nishimura, Karen Gronskov, Alisdair R Philp, Ruth Riise, Thomas Rosenberg, Karen Brondumnielsen
    Abstract:

    Bardet-Biedl syndrome (BBS) is an autosomal recessive disease characterized by retinal dystrophy, polydactyly, obesity, learning disabilities, renal involvement, and male hypogenitalism. BBS is genetically heterogeneous with mutations of 14 genes, accounting for approximately 70% of cases. Triallelic inheritance has been suggested in about 5% of cases. Forty-nine unrelated BBS patients were screened for mutations by DHPLC analysis in BBS1, BBS2, BBS4, BBS6/MKKS, BBS10, and BBS12. The selected genes either account for more than 5% of the mutational load or are commonly reported in triallelic inheritance. Eight patients with only one or no BBS mutation were further investigated by single nucleotide polymorphism (SNP) analysis. In total, mutations were detected in 44 patients. Twenty percent had two mutations in BBS1, 18% in BBS2, 4% in BBS9, 43% in BBS10, and 2% in BBS12. Five patients were heterozygous for a sequence variation in BBS6/MKKS. We found eight patients with three sequence variations in two genes, which could be explained by triallelic inheritance, by the prevalence of heterozygous carriers or the third sequence variations representing rare polymorphisms. All changes found in a second BBS gene were amino acid substitutions. Genotype–phenotype correlations suggest a milder phenotype for BBS1 compared to BBS2 and BBS10, which we ascribe to the hypomorphic p.Met390Arg-mutation. Hum Mutat 31:429–436, 2010. © 2010 Wiley-Liss, Inc.

  • Leptin resistance contributes to obesity and hypertension in mouse models of Bardet-Biedl syndrome.
    The Journal of clinical investigation, 2008
    Co-Authors: Kamal Rahmouni, Darryl Y Nishimura, Melissa A. Fath, Daniel R. Thedens, Christopher J. Berry, Robert M. Weiss, Val C Sheffield
    Abstract:

    Bardet-Biedl syndrome (BBS) is a heterogeneous genetic disorder characterized by many features, including obesity and cardiovascular disease. We previously developed knockout mouse models of 3 BBS genes: BBS2, BBS4, and BBS6. To dissect the mechanisms involved in the metabolic disorders associated with BBS, we assessed the development of obesity in these mouse models and found that BBS-null mice were hyperphagic, had low locomotor activity, and had elevated circulating levels of the hormone leptin. The effect of exogenous leptin on body weight and food intake was attenuated in BBS mice, which suggests that leptin resistance may contribute to hyperleptinemia. In other mouse models of obesity, leptin resistance may be selective rather than systemic; although mice became resistant to leptin's anorectic effects, the ability to increase renal sympathetic nerve activity (SNA) was preserved. Although all 3 of the BBS mouse models were similarly resistant to leptin, the sensitivity of renal SNA to leptin was maintained in Bbs4 -/- and Bbs6 -/- mice, but not in Bbs2 -/- mice. Consequently, Bbs4 -/- and Bbs6 -/- mice had higher baseline renal SNA and arterial pressure and a greater reduction in arterial pressure in response to ganglionic blockade. Furthermore, we found that BBS mice had a decreased hypothalamic expression of proopiomelanocortin, which suggests that BBS genes play an important role in maintaining leptin sensitivity in proopiomelanocortin neurons.

Kamal Rahmouni - One of the best experts on this subject based on the ideXlab platform.

  • abstract 136 the ventromedial hypothalamic bbsome is required for energy homeostasis and sympathetic nerve traffic control
    Hypertension, 2019
    Co-Authors: Deng Fu Guo, Donald A Morgan, Mohamed Rouabhi, Kamal Rahmouni
    Abstract:

    The BBSome, a protein complex of 8 Bardet-Biedl Syndrome (BBS) protein including BBS1, has emerged as an important regulator of metabolic and cardiovascular function. We previously demonstrated tha...

  • Mice lacking the BBS1 gene in the nervous system develop obesity.
    2016
    Co-Authors: Deng Fu Guo, Darryl Nishimura, Qihong Zhang, Val C Sheffield, Donald A Morgan, Daniel R. Thedens, Huxing Cui, Justin L. Grobe, Kamal Rahmouni
    Abstract:

    (A-B) Selectivity of BBS1 gene deletion to the nervous system in NestinCre/BBS1fl/fl mice as determined by RT-PCR (A) and quantitative RT-PCR (B, n = 1 male and 2 females in group). BBS1 expression was absent in the brain (hypothalamus [Hypoth], cortex, hippocampus [Hippo] and brainstem [Brsm]) of NestinCre/BBS1fl/fl mice (Cre+) relative to controls (Cre-). However, BBS1 expression was not altered in white adipose tissue (WAT), liver, kidney and skeletal muscle (Sk Mus). (C) Analysis of the BBSome complex in the brain and testes of Nestincre/BBS1fl/fl mice. Inability of BBS2 to pull down BBS9 in the brain, but not in the testes, of NestinCre/BBS1fl/fl mice (Cre+) relative to controls (Cre-). (D) Weekly body weights of Nestincre/BBS1fl/fl and control mice (n = 12 males and 12 females for controls; 6 males and 6 females for Nestincre/BBS1fl/fl mice). (E-F) Comparison of body composition between Nestincre/BBS1fl/fl and control mice. Shown are coronal (top) and axial abdominal (bottom) representative MRI sections (E) and total fat mass and lean mass (F) of 25 weeks old control and Nestincre/BBS1fl/fl mice (n = 6 males and 6 females in each group). (G) Cumulative food intake of Nestincre/BBS1fl/fl and littermate controls (n = 6 males and 6 females for controls, and 6 males and 8 females for Nestincre/BBS1fl/fl mice). (H) Hypothalamic mRNA levels of AgRP, NPY and POMC in Nestincre/BBS1fl/fl and control mice (n = 3 males and 3 females in each group). Data are means ± SEM, *P< 0.05 vs control group.

  • BBS1 protein interacts with the LRb and regulates its trafficking to the cells surface.
    2016
    Co-Authors: Deng Fu Guo, Darryl Nishimura, Qihong Zhang, Val C Sheffield, Donald A Morgan, Daniel R. Thedens, Huxing Cui, Justin L. Grobe, Kamal Rahmouni
    Abstract:

    (A) Interaction between the BBS1 protein and the endogenous LRb in brain lysate. Ability of Flag-tagged BBS1 and LRb to pull down each other by co-immunoprecipitation assays using brain lysates of transgenic (Tg) mice expressing a Flag-BBS1 protein. IP: immunoprecipitation, IB: immunoblot (representative of n = 3, 1 male and 2 females). (B–C) shRNA-mediated silencing of BBS1 reduces the surface levels of Flag-tagged LRb in a dose-dependent manner in HEK 293 cells transiently co-transfected with Flag-LRb and BBS1-shRNA plasmids. This effect is specific as the surface expression of HA-tagged LRa (in separate experiments) and endogenous transferrin receptor (TrR, in same experiments as the Flag-LRb) were not altered (n = 6 per group). (D-E) Silencing of BBS2, another BBSome protein, lowers the surface levels of Flag-LRb, but not HA-LRa or endogenous TrR in HEK 293 cells (n = 6 per group). (F–G) Blockade of Rab8a using a dominant negative (DN) form decreases the surface expression of Flag-LRb, HA-LRa and endogenous TrR in HEK 293 cells (n = 6 per group). (H–I) siRNA-mediated silencing of Ift88 had no effect on the surface expression of Flag-LRb or endogenous TrR in HEK 293 cells (n = 8 per group). Data are means ± SEM, *P< 0.05 vs. control group.

  • abstract 083 metabolic and cardiovascular effects of BBS1 ablation from the pomc containing neurons
    Hypertension, 2015
    Co-Authors: Deng F. Guo, Val C Sheffield, Donald A Morgan, Justin L. Grobe, Kamal Rahmouni
    Abstract:

    Bardet-Biedl syndrome (BBS) is a pleiotropic autosomal recessive disorder associated with several features including obesity and hypertension. Deletion of Bbs genes globally, in the nervous system or in the leptin receptor-expressing cells recapitulated many of the BBS phenotypes including obesity and hypertension. Here, we assessed the effect of ablating the BBS1 gene from the neurons expressing proopiomelanocortin (POMC) neurons. Breeding BBS1flox mice with POMCCre mice created mice deficient in BBS1 gene only in the POMC-positive neurons (visualized by tdTomato expression). Importantly, POMCCre/BBS1fl/fl mice display an obesity phenotype as indicated by the increased (P<0.05) body weight (48.6±1.7 vs 34.9±0.8 g in controls) and fat pads (3.3±0.3 vs. 0.7±1.1 g for inguinal fat, 2.1±0.2 vs. 0.4±0.05 g for perirenal fat, 3.3±0.3 vs. 0.8±0.3 g for reproduction fat and 0.4±0.02 vs. 0.2±0.02 g for brow fat) associated with increased (P<0.05) food intake (3.79.±0.14 vs. 2.97±0.15 g in controls) in 25 weeks old mice. POMCCre/BBS1fl/fl mice displayed decreased (P<0.05) O2consumption (2.6±0.03 vs. 3±0.04 mL O2/100g/min in controls) and heat production (8.0±0.09 vs. 9.2±0.13 kcal/kg/hr in controls). These results indicate that hyperphagia and decreased energy expenditure contribute to the development of obesity in POMCCre/BBS1fl/fl mice. Next, we assessed the consequence on arterial pressure (AP) and sympathetic nerve activity (SNA) of ablating the BBS1 gene from POMC neurons. Interestingly, deletion of the BBS1 gene in POMC neurons did not recapitulate the hypertension phenotype of BBS as indicated by the slight, but not significant increase in mean AP (116±4.7 vs 109±6.1 mmHg in controls). However, conscious renal SNA was significantly higher in POMCCre/BBS1fl/fl mice relative to controls (132.4±11 vs 74.3±4.2 spikes/sec, P<0.05). Finally, the depressor effect of ganglionic blockade (hexamethonium) was exaggerated in POMCCre/BBS1fl/fl mice (-68.6±4.4 vs -45.2±9.4 mmHg in control, P=0.017). These findings demonstrate that BBS1 gene in the POMC neurons is critical for energy homeostasis, but not for arterial pressure regulation.

  • Physical interaction between BBS proteins and IR.
    2015
    Co-Authors: Rachel D. Starks, Qihong Zhang, Val C Sheffield, Deng Fu Guo, Andreas M. Beyer, Lauren Boland, Kamal Rahmouni
    Abstract:

    A) Sedimentation analysis showing the β subunit of the IR in complex with the BBSome (corresponding to fraction 9). B-C) Reciprocal co-immunoprecipitation with GFP-tagged BBS17 and the β subunit of the IR in protein lysates from HEK293T cells. The interaction between the IRβ subunit and BBS17 can be detected whether IR antibody is used for immunoprecipitation (A) or GFP-BBS17 is used to precipitate IR (Ctrl line in C). Silencing the BBS1 gene with shRNA disrupt the interaction between BBS17 and IRβ (right line in C).

Seongjin Seo - One of the best experts on this subject based on the ideXlab platform.

  • accumulation of non outer segment proteins in the outer segment underlies photoreceptor degeneration in bardet biedl syndrome
    Proceedings of the National Academy of Sciences of the United States of America, 2015
    Co-Authors: Poppy Datta, Val C Sheffield, Arlene V. Drack, Sajag Bhattarai, Chantal Allamargot, Joseph S Hudson, Emily K Andersen, Seongjin Seo
    Abstract:

    Compartmentalization and polarized protein trafficking are essential for many cellular functions. The photoreceptor outer segment (OS) is a sensory compartment specialized for phototransduction, and it shares many features with primary cilia. As expected, mutations disrupting protein trafficking to cilia often disrupt protein trafficking to the OS and cause photoreceptor degeneration. Bardet–Biedl syndrome (BBS) is one of the ciliopathies associated with defective ciliary trafficking and photoreceptor degeneration. However, precise roles of BBS proteins in photoreceptor cells and the underlying mechanisms of photoreceptor degeneration in BBS are not well understood. Here, we show that accumulation of non-OS proteins in the OS underlies photoreceptor degeneration in BBS. Using a newly developed BBS mouse model [Leucine zipper transcription factor-like 1 (Lztfl1)/BBS17 mutant], isolated OSs, and quantitative proteomics, we determined 138 proteins that are enriched more than threefold in BBS mutant OS. In contrast, only eight proteins showed a more than threefold reduction. We found striking accumulation of Stx3 and Stxbp1/Munc18-1 and loss of polarized localization of Prom1 within the Lztfl1 and BBS1 mutant OS. Ultrastructural analysis revealed that large vesicles are formed in the BBS OS, disrupting the lamellar structure of the OS. Our findings suggest that accumulation (and consequent sequestration) of non-OS proteins in the OS is likely the primary cause of photoreceptor degeneration in BBS. Our data also suggest that a major function of BBS proteins in photoreceptors is to transport proteins from the OS to the cell body or to prevent entry of non-OS proteins into the OS.

  • The centriolar satellite protein AZI1 interacts with BBS4 and regulates ciliary trafficking of the BBSome.
    PLoS genetics, 2014
    Co-Authors: Xitiz Chamling, Charles Searby, Diane C Slusarski, Seongjin Seo, Gunhee Kim, Val C Sheffield
    Abstract:

    Bardet-Biedl syndrome (BBS) is a well-known ciliopathy with mutations reported in 18 different genes. Most of the protein products of the BBS genes localize at or near the primary cilium and the centrosome. Near the centrosome, BBS proteins interact with centriolar satellite proteins, and the BBSome (a complex of seven BBS proteins) is believed to play a role in transporting ciliary membrane proteins. However, the precise mechanism by which BBSome ciliary trafficking activity is regulated is not fully understood. Here, we show that a centriolar satellite protein, AZI1 (also known as CEP131), interacts with the BBSome and regulates BBSome ciliary trafficking activity. Furthermore, we show that AZI1 interacts with the BBSome through BBS4. AZI1 is not involved in BBSome assembly, but accumulation of the BBSome in cilia is enhanced upon AZI1 depletion. Under conditions in which the BBSome does not normally enter cilia, such as in BBS3 or BBS5 depleted cells, knock down of AZI1 with siRNA restores BBSome trafficking to cilia. Finally, we show that azi1 knockdown in zebrafish embryos results in typical BBS phenotypes including Kupffer's vesicle abnormalities and melanosome transport delay. These findings associate AZI1 with the BBS pathway. Our findings provide further insight into the regulation of BBSome ciliary trafficking and identify AZI1 as a novel BBS candidate gene.

  • The Centriolar Satellite Protein AZI1 Interacts with BBS4 and Regulates Ciliary Trafficking of the BBSome
    2014
    Co-Authors: Xitiz Chamling, Charles Searby, Diane C Slusarski, Seongjin Seo, Gunhee Kim, Val C Sheffield
    Abstract:

    Bardet-Biedl syndrome (BBS) is a well-known ciliopathy with mutations reported in 18 different genes. Most of the protein products of the BBS genes localize at or near the primary cilium and the centrosome. Near the centrosome, BBS proteins interact with centriolar satellite proteins, and the BBSome (a complex of seven BBS proteins) is believed to play a role in transporting ciliary membrane proteins. However, the precise mechanism by which BBSome ciliary trafficking activity is regulated is not fully understood. Here, we show that a centriolar satellite protein, AZI1 (also known as CEP131), interacts with the BBSome and regulates BBSome ciliary trafficking activity. Furthermore, we show that AZI1 interacts with the BBSome through BBS4. AZI1 is not involved in BBSome assembly, but accumulation of the BBSome in cilia is enhanced upon AZI1 depletion. Under conditions in which the BBSome does not normally enter cilia, such as in BBS3 or BBS5 depleted cells, knock down of AZI1 with siRNA restores BBSome trafficking to cilia. Finally, we show that azi1 knockdown in zebrafish embryos results in typical BBS phenotypes including Kupffer’s vesicle abnormalities and melanosome transport delay. These findings associate AZI1 with the BBS pathway. Our findings provide further insight into the regulation of BBSome ciliar

  • AZI1 knockdown increases ciliary localization of BBS4.
    2014
    Co-Authors: Xitiz Chamling, Charles Searby, Diane C Slusarski, Seongjin Seo, Gunhee Kim, Val C Sheffield
    Abstract:

    A) RPE-1 cells expressing GFP-BBS4 were depleted of AZI1, and the number of cells with ciliary GFP (BBS4) was counted. BBS4 is stained with GFP (green), Acetylated α-tubulin was used to detect cilia (red). B) Graph showing significant increases in ciliary localization of BBS4 upon AZI1 knockdown. C) Depletion of AZI1 in BBS3 and BBS5 depleted cells restores ciliary BBSome localization. RPE-1 cells were transfected with siRNA as indicated, and BBS9 (red) localization was analyzed. Cilia (green) in the insets of figures A and C are slightly shifted to show ciliary localization of BBS proteins. D) Cilia containing BBS9 at different conditions were counted and presented graphically. All data are presented in mean +/− SEM. Significance is calculated using the Student's t-test for C and E, and one way ANOVA for A. P

  • AZI1 knockdown reduces ciliogenesis but increases ciliary localization of the BBSome.
    2014
    Co-Authors: Xitiz Chamling, Charles Searby, Diane C Slusarski, Seongjin Seo, Gunhee Kim, Val C Sheffield
    Abstract:

    A) RPE-1 cells were transfected with siRNA against BBS4, PCM1, and AZI1. Approximately 500 cells per sample were counted. B) AZI1 depletion increases the ciliary localization of BBS9 compared to the control knockdown. Red staining represents BBS9, and cilia are stained with acetylated α-tubulin. Nuclei are stained blue with DAPI. C) Graph showing a significant increase in cells with ciliary BBS9 upon AZI1 knockdown by different siRNAs. D) BBS8 (red) is used as a BBSome marker to confirm increased ciliary localization of the BBSome upon AZI1 knockdown. Cilia (green) in the insets of figures B and D are slightly shifted to show ciliary localization of BBS proteins. E) The graph shows a significant increase in the number of ciliated cells with BBS8 or BBS9 upon AZI1 knockdown, and decrease in ciliary BBS8 or BBS9 localization upon AZI1 overexpression. Approximately 250 ciliated cells were counted in control as well as AZI1 knockdown culture of RPE-1 cells. All data are presented in mean +/− SEM. Significance is calculated using Student's t-test for C and E, and one way ANOVA for A. P