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Chantal Desdouets - One of the best experts on this subject based on the ideXlab platform.
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The intraflagellar transport component IFT88/polaris is a centrosomal protein regulating G1-S transition in non-ciliated cells
Journal of Cell Science, 2007Co-Authors: Aude Robert, Germain Margall-ducos, Jacques-emmanuel Guidotti, Olivier Brégerie, Claude Celati, Christian Bréchot, Chantal DesdouetsAbstract:Loss of normal primary cilia function in mammals is linked to proliferative diseases, such as polycystic kidney disease, suggesting a regulatory relationship between cilia and cell cycle. The primary cilium expressed by most mammalian cells is nucleated from the elder centriole of the centrosome. The relationship between centrosome and cilia suggests that these structures share functions and components. We now show that IFT88/polaris, a component of the intraflagellar transport, remains associated to the centrosome in a proliferative state. IFT88/polaris is tightly associated with the centrosome throughout the cell cycle in a microtubule- and dynein-independent manner. IFT88/polaris tetratricopeptide repeat motifs are essential for this localization. Overexpression of IFT88/polaris prevents G1-S transition and induces apoptotic cell death. By contrast, IFT88/polaris depletion induced by RNA interference promotes cell-cycle progression to S, G2, and M phases. Finally, we demonstrate that IFT88/polaris interacts with Che-1, an Rb-binding protein that inhibits the Rb growth suppressing function. We propose that IFT88/polaris, a protein essential for ciliogenesis, is also crucial for G1-S transition in non-ciliated cells.
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the intraflagellar transport component IFT88 polaris is a centrosomal protein regulating g1 s transition in non ciliated cells
Journal of Cell Science, 2007Co-Authors: Aude Robert, Jacques-emmanuel Guidotti, Olivier Brégerie, Claude Celati, Christian Bréchot, Germain Margallducos, Chantal DesdouetsAbstract:Loss of normal primary cilia function in mammals is linked to proliferative diseases, such as polycystic kidney disease, suggesting a regulatory relationship between cilia and cell cycle. The primary cilium expressed by most mammalian cells is nucleated from the elder centriole of the centrosome. The relationship between centrosome and cilia suggests that these structures share functions and components. We now show that IFT88/polaris, a component of the intraflagellar transport, remains associated to the centrosome in a proliferative state. IFT88/polaris is tightly associated with the centrosome throughout the cell cycle in a microtubule- and dynein-independent manner. IFT88/polaris tetratricopeptide repeat motifs are essential for this localization. Overexpression of IFT88/polaris prevents G1-S transition and induces apoptotic cell death. By contrast, IFT88/polaris depletion induced by RNA interference promotes cell-cycle progression to S, G2, and M phases. Finally, we demonstrate that IFT88/polaris interacts with Che-1, an Rb-binding protein that inhibits the Rb growth suppressing function. We propose that IFT88/polaris, a protein essential for ciliogenesis, is also crucial for G1-S transition in non-ciliated cells.
Esben Lorentzen - One of the best experts on this subject based on the ideXlab platform.
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crystal structure of intraflagellar transport protein 80 reveals a homo dimer required for ciliogenesis
eLife, 2018Co-Authors: Michael Taschner, Andre Mourao, Anna Lorentzen, Toby Collins, Grace Freke, Dale Moulding, Jerome Basquin, Dagan Jenkins, Esben LorentzenAbstract:Oligomeric assemblies of intraflagellar transport (IFT) particles build cilia through sequential recruitment and transport of ciliary cargo proteins within cilia. Here we present the 1.8 A resolution crystal structure of the Chlamydomonas IFT-B protein IFT80, which reveals the architecture of two N-terminal β-propellers followed by an α-helical extension. The N-terminal β-propeller tethers IFT80 to the IFT-B complex via IFT38 whereas the second β-propeller and the C-terminal α-helical extension result in IFT80 homo-dimerization. Using CRISPR/Cas to create biallelic Ift80 frameshift mutations in IMCD3 mouse cells, we demonstrate that IFT80 is absolutely required for ciliogenesis. Structural mapping and rescue experiments reveal that human disease-causing missense mutations do not cluster within IFT80 and form functional IFT particles. Unlike missense mutant forms of IFT80, deletion of the C-terminal dimerization domain prevented rescue of ciliogenesis. Taken together our results may provide a first insight into higher order IFT complex formation likely required for IFT train formation.
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intraflagellar transport protein ift52 recruits ift46 to the basal body and flagella
Journal of Cell Science, 2017Co-Authors: Lei Wan, Michael Taschner, Esben Lorentzen, Xi Cheng, Kaiyao HuangAbstract:ABSTRACT Cilia are microtubule-based organelles and perform motile, sensing and signaling functions. The assembly and maintenance of cilia depend on intraflagellar transport (IFT). Besides ciliary localization, most IFT proteins accumulate at basal bodies. However, little is known about the molecular mechanism of basal body targeting of IFT proteins. We first identified the possible basal body-targeting sequence in IFT46 by expressing IFT46 truncation constructs in an ift46-1 mutant. The C-terminal sequence between residues 246–321, termed BBTS3, was sufficient to target YFP to basal bodies in the ift46-1 strain. Interestingly, BBTS3 is also responsible for the ciliary targeting of IFT46. BBTS3::YFP moves bidirectionally in flagella and interacts with other IFT complex B (IFT-B) proteins. Using IFT and motor mutants, we show that the basal body localization of IFT46 depends on IFT52, but not on IFT81, IFT88, IFT122, FLA10 or DHC1b. IFT52 interacts with IFT46 through residues L285 and L286 of IFT46 and recruits it to basal bodies. Ectopic expression of the C-terminal domain of IFT52 in the nucleus resulted in accumulation of IFT46 in nuclei. These data suggest that IFT52 and IFT46 can preassemble as a complex in the cytoplasm, which is then targeted to basal bodies.
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IFT proteins spatially control the geometry of cleavage furrow ingression and lumen positioning
Nature Communications, 2017Co-Authors: Nicolas Taulet, Michael Taschner, Esben Lorentzen, Audrey Douanier, Benjamin Vitre, Christelle Anguille, Murielle Rocancourt, Arnaud Echard, Benedicte DelavalAbstract:Cytokinesis mediates the physical separation of dividing cells and, in 3D epithelia, provides a spatial landmark for lumen formation. Here, we unravel an unexpected role in cytokinesis for proteins of the intraflagellar transport (IFT) machinery, initially characterized for their ciliary role and their link to polycystic kidney disease. Using 2D and 3D cultures of renal cells, we show that IFT proteins are required to correctly shape the central spindle, to control symmetric cleavage furrow ingression and to ensure central lumen positioning. Mechanistically, IFT88 directly interacts with the kinesin MKLP2 and is essential for the correct relocalization of the Aurora B/MKLP2 complex to the central spindle. IFT88 is thus required for proper centralspindlin distribution and central spindle microtubule organization. Overall, this work unravels a novel non-ciliary mechanism for IFT proteins at the central spindle, which could contribute to kidney cyst formation by affecting lumen positioning.
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intraflagellar transport proteins 172 80 57 54 38 and 20 form a stable tubulin binding ift b2 complex
The EMBO Journal, 2016Co-Authors: Michael Taschner, Sagar Bhogaraju, Kristina Weber, Andre Mourao, Melanie Vetter, Mayanka Awasthi, Marc Stiegler, Esben LorentzenAbstract:Intraflagellar transport (IFT) relies on the IFT complex and is required for ciliogenesis. The IFT-B complex consists of 9-10 stably associated core subunits and six "peripheral" subunits that were shown to dissociate from the core structure at moderate salt concentration. We purified the six "peripheral"IFT-B subunits of Chlamydomonas reinhardtiias recombinant proteins and show that they form a stable complex independently of the IFT-B core. We suggest a nomenclature of IFT-B1 (core) and IFT-B2 (peripheral) for the two IFT-B subcomplexes. We demonstrate that IFT88, together with the N-terminal domain of IFT52, is necessary to bridge the interaction between IFT-B1 and B2. The crystal structure of IFT52N reveals highly conserved residues critical for IFT-B1/IFT-B2 complex formation. Furthermore, we show that of the three IFT-B2 subunits containing a calponin homology (CH) domain (IFT38, 54, and 57), only IFT54 binds αβ-tubulin as a potential IFT cargo, whereas the CH domains of IFT38 and IFT57 mediate the interaction with IFT80 and IFT172, respectively. Crystal structures of IFT54 CH domains reveal that tubulin binding is mediated by basic surface-exposed residues.
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crystal structures of ift70 52 and ift52 46 provide insight into intraflagellar transport b core complex assembly
Journal of Cell Biology, 2014Co-Authors: Michael Taschner, Fruzsina Kotsis, Philipp Braeuer, Wolfgang E Kuehn, Esben LorentzenAbstract:Cilia are microtubule-based organelles that assemble via intraflagellar transport (IFT) and function as signaling hubs on eukaryotic cells. IFT relies on molecular motors and IFT complexes that mediate the contacts with ciliary cargo. To elucidate the architecture of the IFT-B complex, we reconstituted and purified the nonameric IFT-B core from Chlamydomonas reinhardtii and determined the crystal structures of C. reinhardtii IFT70/52 and Tetrahymena IFT52/46 subcomplexes. The 2.5-A resolution IFT70/52 structure shows that IFT52330–370 is buried deeply within the IFT70 tetratricopeptide repeat superhelix. Furthermore, the polycystic kidney disease protein IFT88 binds IFT52281–329 in a complex that interacts directly with IFT70/IFT52330–381 in trans. The structure of IFT52C/IFT46C was solved at 2.3 A resolution, and we show that it is essential for IFT-B core integrity by mediating interaction between IFT88/70/52/46 and IFT81/74/27/25/22 subcomplexes. Consistent with this, overexpression of mammalian IFT52C in MDCK cells is dominant-negative and causes IFT protein mislocalization and disrupted ciliogenesis. These data further rationalize several ciliogenesis phenotypes of IFT mutant strains.
Michael Taschner - One of the best experts on this subject based on the ideXlab platform.
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IFT proteins interact with HSET to promote supernumerary centrosome clustering in mitosis
EMBO Reports, 2020Co-Authors: Benjamin Vitre, Nicolas Taulet, Audrey Douanier, Christelle Anguille, Justine Maurin, Audrey Guesdon, Aurelie Dosdane, Melanie Cisneros, Sabrina Hettinger, Michael TaschnerAbstract:Centrosome amplification is a hallmark of cancer, and centrosome clustering is essential for cancer cell survival. The mitotic kinesin HSET is an essential contributor to this process. Recent studies have highlighted novel functions for intraflagellar transport (IFT) proteins in regulating motors and mitotic processes. Here, using siRNA knock-down of various IFT proteins or AID-inducible degradation of endogenous IFT88 in combination with small-molecule inhibition of HSET, we show that IFT proteins together with HSET are required for efficient centrosome clustering. We identify a direct interaction between the kinesin HSET and IFT proteins, and we define how IFT proteins contribute to clustering dynamics during mitosis using high-resolution live imaging of centrosomes. Finally, we demonstrate the requirement of IFT88 for efficient centrosome clustering in a variety of cancer cell lines naturally harboring supernumerary centrosomes and its importance for cancer cell proliferation. Overall, our data unravel a novel role for the IFT machinery in centrosome clustering during mitosis in cells harboring supernumerary centrosomes.
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crystal structure of intraflagellar transport protein 80 reveals a homo dimer required for ciliogenesis
eLife, 2018Co-Authors: Michael Taschner, Andre Mourao, Anna Lorentzen, Toby Collins, Grace Freke, Dale Moulding, Jerome Basquin, Dagan Jenkins, Esben LorentzenAbstract:Oligomeric assemblies of intraflagellar transport (IFT) particles build cilia through sequential recruitment and transport of ciliary cargo proteins within cilia. Here we present the 1.8 A resolution crystal structure of the Chlamydomonas IFT-B protein IFT80, which reveals the architecture of two N-terminal β-propellers followed by an α-helical extension. The N-terminal β-propeller tethers IFT80 to the IFT-B complex via IFT38 whereas the second β-propeller and the C-terminal α-helical extension result in IFT80 homo-dimerization. Using CRISPR/Cas to create biallelic Ift80 frameshift mutations in IMCD3 mouse cells, we demonstrate that IFT80 is absolutely required for ciliogenesis. Structural mapping and rescue experiments reveal that human disease-causing missense mutations do not cluster within IFT80 and form functional IFT particles. Unlike missense mutant forms of IFT80, deletion of the C-terminal dimerization domain prevented rescue of ciliogenesis. Taken together our results may provide a first insight into higher order IFT complex formation likely required for IFT train formation.
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intraflagellar transport protein ift52 recruits ift46 to the basal body and flagella
Journal of Cell Science, 2017Co-Authors: Lei Wan, Michael Taschner, Esben Lorentzen, Xi Cheng, Kaiyao HuangAbstract:ABSTRACT Cilia are microtubule-based organelles and perform motile, sensing and signaling functions. The assembly and maintenance of cilia depend on intraflagellar transport (IFT). Besides ciliary localization, most IFT proteins accumulate at basal bodies. However, little is known about the molecular mechanism of basal body targeting of IFT proteins. We first identified the possible basal body-targeting sequence in IFT46 by expressing IFT46 truncation constructs in an ift46-1 mutant. The C-terminal sequence between residues 246–321, termed BBTS3, was sufficient to target YFP to basal bodies in the ift46-1 strain. Interestingly, BBTS3 is also responsible for the ciliary targeting of IFT46. BBTS3::YFP moves bidirectionally in flagella and interacts with other IFT complex B (IFT-B) proteins. Using IFT and motor mutants, we show that the basal body localization of IFT46 depends on IFT52, but not on IFT81, IFT88, IFT122, FLA10 or DHC1b. IFT52 interacts with IFT46 through residues L285 and L286 of IFT46 and recruits it to basal bodies. Ectopic expression of the C-terminal domain of IFT52 in the nucleus resulted in accumulation of IFT46 in nuclei. These data suggest that IFT52 and IFT46 can preassemble as a complex in the cytoplasm, which is then targeted to basal bodies.
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IFT proteins spatially control the geometry of cleavage furrow ingression and lumen positioning
Nature Communications, 2017Co-Authors: Nicolas Taulet, Michael Taschner, Esben Lorentzen, Audrey Douanier, Benjamin Vitre, Christelle Anguille, Murielle Rocancourt, Arnaud Echard, Benedicte DelavalAbstract:Cytokinesis mediates the physical separation of dividing cells and, in 3D epithelia, provides a spatial landmark for lumen formation. Here, we unravel an unexpected role in cytokinesis for proteins of the intraflagellar transport (IFT) machinery, initially characterized for their ciliary role and their link to polycystic kidney disease. Using 2D and 3D cultures of renal cells, we show that IFT proteins are required to correctly shape the central spindle, to control symmetric cleavage furrow ingression and to ensure central lumen positioning. Mechanistically, IFT88 directly interacts with the kinesin MKLP2 and is essential for the correct relocalization of the Aurora B/MKLP2 complex to the central spindle. IFT88 is thus required for proper centralspindlin distribution and central spindle microtubule organization. Overall, this work unravels a novel non-ciliary mechanism for IFT proteins at the central spindle, which could contribute to kidney cyst formation by affecting lumen positioning.
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intraflagellar transport proteins 172 80 57 54 38 and 20 form a stable tubulin binding ift b2 complex
The EMBO Journal, 2016Co-Authors: Michael Taschner, Sagar Bhogaraju, Kristina Weber, Andre Mourao, Melanie Vetter, Mayanka Awasthi, Marc Stiegler, Esben LorentzenAbstract:Intraflagellar transport (IFT) relies on the IFT complex and is required for ciliogenesis. The IFT-B complex consists of 9-10 stably associated core subunits and six "peripheral" subunits that were shown to dissociate from the core structure at moderate salt concentration. We purified the six "peripheral"IFT-B subunits of Chlamydomonas reinhardtiias recombinant proteins and show that they form a stable complex independently of the IFT-B core. We suggest a nomenclature of IFT-B1 (core) and IFT-B2 (peripheral) for the two IFT-B subcomplexes. We demonstrate that IFT88, together with the N-terminal domain of IFT52, is necessary to bridge the interaction between IFT-B1 and B2. The crystal structure of IFT52N reveals highly conserved residues critical for IFT-B1/IFT-B2 complex formation. Furthermore, we show that of the three IFT-B2 subunits containing a calponin homology (CH) domain (IFT38, 54, and 57), only IFT54 binds αβ-tubulin as a potential IFT cargo, whereas the CH domains of IFT38 and IFT57 mediate the interaction with IFT80 and IFT172, respectively. Crystal structures of IFT54 CH domains reveal that tubulin binding is mediated by basic surface-exposed residues.
Iain A. Drummond - One of the best experts on this subject based on the ideXlab platform.
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Assaying sensory ciliopathies using calcium biosensor expression in zebrafish ciliated olfactory neurons
Cilia, 2018Co-Authors: Judith G. M. Bergboer, Cameron Wyatt, Christina Austin-tse, Emre Yaksi, Iain A. DrummondAbstract:Background Primary cilia mediate signal transduction by acting as an organizing scaffold for receptors, signalling proteins and ion channels. Ciliated olfactory sensory neurons (OSNs) organize olfactory receptors and ion channels on cilia and generate a calcium influx as a primary signal in odourant detection. In the zebrafish olfactory placode, ciliated OSNs and microvillus OSNs constitute the major OSN cell types with distinct odourant sensitivity. Methods Using transgenic expression of the calcium biosensor GCaMP5 in OSNs, we analysed sensory cilia-dependent odour responses in live zebrafish, at individual cell resolution. oval/IFT88 mutant and ift172 knockdown zebrafish were compared with wild-type siblings to establish ciliated OSN sensitivity to different classes of odourants. Results oval/IFT88 mutant and ift172 knockdown zebrafish showed fewer and severely shortened OSN cilia without a reduction in OSN number. The fraction of responding OSNs and response amplitudes to bile acids and food odour, both sensed by ciliated OSNs, were significantly reduced in IFT88 mutants and ift172 -deficient embryos, while the amino acids responses were not significantly changed. Conclusions Our approach presents a quantitative model for studying sensory cilia signalling using zebrafish OSNs. Our results also implicate ift172 -deficiency as a novel cause of hyposmia, a reduced sense of smell, highlighting the value of directly assaying sensory cilia signalling in vivo and supporting the idea that hyposmia can be used as a diagnostic indicator of ciliopathies.
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Assaying sensory ciliopathies using calcium biosensor expression in zebrafish ciliated olfactory neurons.
Cilia, 2018Co-Authors: Judith G. M. Bergboer, Cameron Wyatt, Christina Austin-tse, Emre Yaksi, Iain A. DrummondAbstract:Primary cilia mediate signal transduction by acting as an organizing scaffold for receptors, signalling proteins and ion channels. Ciliated olfactory sensory neurons (OSNs) organize olfactory receptors and ion channels on cilia and generate a calcium influx as a primary signal in odourant detection. In the zebrafish olfactory placode, ciliated OSNs and microvillus OSNs constitute the major OSN cell types with distinct odourant sensitivity. Using transgenic expression of the calcium biosensor GCaMP5 in OSNs, we analysed sensory cilia-dependent odour responses in live zebrafish, at individual cell resolution. oval/IFT88 mutant and ift172 knockdown zebrafish were compared with wild-type siblings to establish ciliated OSN sensitivity to different classes of odourants. oval/IFT88 mutant and ift172 knockdown zebrafish showed fewer and severely shortened OSN cilia without a reduction in OSN number. The fraction of responding OSNs and response amplitudes to bile acids and food odour, both sensed by ciliated OSNs, were significantly reduced in IFT88 mutants and ift172-deficient embryos, while the amino acids responses were not significantly changed. Our approach presents a quantitative model for studying sensory cilia signalling using zebrafish OSNs. Our results also implicate ift172-deficiency as a novel cause of hyposmia, a reduced sense of smell, highlighting the value of directly assaying sensory cilia signalling in vivo and supporting the idea that hyposmia can be used as a diagnostic indicator of ciliopathies.
Rosa Serra - One of the best experts on this subject based on the ideXlab platform.
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Normal mammary development and function in mice with IFT88 deleted in MMTV- and K14-Cre expressing cells
Cilia, 2014Co-Authors: Elizabeth H Mitchell, Rosa SerraAbstract:Background Primary cilia (PC) are non-motile microtubule based organelles present on almost every cell type and are known to serve as critical organizing centers for several signaling pathways crucial to embryonic and postnatal development. Alterations in the Hh pathway, the most studied signaling pathway regulated by PC, affect mammary gland development as well as maintenance of the stem and progenitor cell populations. Results We developed mouse models with deletion of PC in mammary luminal epithelial, basal epithelial, and stromal cells for evaluation of the function of PC in mammary development via MMTV-Cre, K14-Cre, and Prx1-Cre mediated deletion, respectively. The activity of Cre was confirmed using ROSA26 reporters. Mammary stem and progenitor cells were enriched through growth as mammospheres. Adenovirus-Cre mediated deletion of IFT88 was used to determine a role for PC in this population of cells. Disruption of IFT88 and PC were confirmed in using PCR and immunofluorescent methods. Prx1-Cre; IFT88 ^Del mice demonstrated defects in terminal end buds during puberty. However, these IFT88 ^Del glands exhibited typical terminal end bud formation as well as normal ductal histology when transplanted into wild type hosts, indicating that the phenotype observed was not intrinsic to the mammary gland. Furthermore, no discernable alterations to mammary development were observed in MMTV-Cre- or K14-Cre; IFT88 ^Del lines. These mice were able to feed and support several litters of pups even though wide spread depletion of PC was confirmed. Cells grown in mammosphere culture were enriched for PC containing cells suggesting PC are preferentially expressed on mammary stem and progenitor cells. Deletion of IFT88 in mammary epithelial cells resulted in a significant reduction in the number of primary mammospheres established; however, there was no effect on outgrowth of secondary mammospheres in PC-depleted cells. Conclusions PC regulate systemic factors that can affect mammary development in early puberty. PC on MMTV- or K14-expressing epithelial cells are not required for normal mammary development or function. PC are expressed at high levels on cells in mammosphere cultures. PC may be required for cells to establish mammospheres in culture; however, PC are not required for renewal of the cultures.
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IFT88 regulates hedgehog signaling sfrp5 expression and β catenin activity in post natal growth plate
Journal of Orthopaedic Research, 2013Co-Authors: Ching-fang Chang, Rosa SerraAbstract:Primary cilia are present on most cell types including chondrocytes. Dysfunction of primary cilia results in pleiotropic symptoms including skeletal dysplasia. Previously, we showed that deletion of IFT88 and subsequent depletion of primary cilia from chondrocytes resulted in disorganized columnar structure and early loss of growth plate. To understand underlying mechanisms whereby IFT88 regulates growth plate function, we compared gene expression profiles in normal and IFT88 deleted growth plates. Pathway analysis indicated that Hedgehog (Hh) signaling was the most affected pathway in mutant growth plate. Expression of the Wnt antagonist, Sfrp5, was also down-regulated. In addition, Sfrp5 was up-regulated by Shh in rib chondrocytes and regulation of Sfrp5 by Shh was attenuated in mutant cells. This result suggests Sfrp5 is a downstream target of Hh and that IFT88 regulates its expression. Sfrp5 is an extracellular antagonist of Wnt signaling. We observed an increase in Wnt/β-catenin signaling specifically in flat columnar cells of the growth plate in IFT88 mutant mice as measured by increased expression of Axin2 and Lef1 as well as increased nuclear localization of β-catenin. We propose that IFT88 and primary cilia regulate expression of Sfrp5 and Wnt signaling pathways in growth plate via regulation of Ihh signaling.
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IFT88 regulates Hedgehog signaling, Sfrp5 expression, and β‐catenin activity in post‐natal growth plate
Journal of orthopaedic research : official publication of the Orthopaedic Research Society, 2012Co-Authors: Ching-fang Chang, Rosa SerraAbstract:Primary cilia are present on most cell types including chondrocytes. Dysfunction of primary cilia results in pleiotropic symptoms including skeletal dysplasia. Previously, we showed that deletion of IFT88 and subsequent depletion of primary cilia from chondrocytes resulted in disorganized columnar structure and early loss of growth plate. To understand underlying mechanisms whereby IFT88 regulates growth plate function, we compared gene expression profiles in normal and IFT88 deleted growth plates. Pathway analysis indicated that Hedgehog (Hh) signaling was the most affected pathway in mutant growth plate. Expression of the Wnt antagonist, Sfrp5, was also down-regulated. In addition, Sfrp5 was up-regulated by Shh in rib chondrocytes and regulation of Sfrp5 by Shh was attenuated in mutant cells. This result suggests Sfrp5 is a downstream target of Hh and that IFT88 regulates its expression. Sfrp5 is an extracellular antagonist of Wnt signaling. We observed an increase in Wnt/β-catenin signaling specifically in flat columnar cells of the growth plate in IFT88 mutant mice as measured by increased expression of Axin2 and Lef1 as well as increased nuclear localization of β-catenin. We propose that IFT88 and primary cilia regulate expression of Sfrp5 and Wnt signaling pathways in growth plate via regulation of Ihh signaling.
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depletion of primary cilia in articular chondrocytes results in reduced gli3 repressor to activator ratio increased hedgehog signaling and symptoms of early osteoarthritis
Osteoarthritis and Cartilage, 2012Co-Authors: Ching-fang Chang, Girish Ramaswamy, Rosa SerraAbstract:Summary Objective Primary cilia are present in almost every cell type including chondrocytes. Studies have shown that defects in primary cilia result in skeletal dysplasia. The purpose of this study was to understand how loss of primary cilia affects articular cartilage. Design IFT88 encodes a protein that is required for intraflagellar transport and formation of primary cilia. In this study, we used Col2aCre;IFT88 fl/fl transgenic mice in which primary cilia were deleted in chondrocytes. Col2aCre;IFT88 fl/fl articular cartilage was characterized by histological staining, real time RT-PCR, and microindentation. Hedgehog (Hh) signaling was measured by expression of Ptch1 and Gli1 mRNA. The levels of Gli3 proteins were determined by western blot. Results Col2aCre;IFT88 fl/fl articular cartilage was thicker and had increased cell density, likely due to decreased apoptosis during cartilage remodeling. Mutant articular cartilage also showed increased expression of osteoarthritis (OA) markers including Mmp13 , Adamts5 , ColX , and Runx2 . OA was also evident by reduced stiffness in mutant cartilage as measured by microindentation. Up-regulation of Hh signaling, which has been associated with OA, was present in mutant articular cartilage as measured by expression of Ptch1 and Gli1 . Col2aCre;IFT88 fl/fl cartilage also demonstrated reduced Gli3 repressor to activator ratio. Conclusion Our results indicate that primary cilia are required for normal development and maintenance of articular cartilage. It was shown that primary cilia are required for processing full length Gli3 to the truncated repressor form. We propose that OA symptoms in Col2aCre;IFT88 fl/fl cartilage are due to reduced Hh signal repression by Gli3.
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Role for primary cilia in the regulation of mouse ovarian function.
Developmental dynamics : an official publication of the American Association of Anatomists, 2008Co-Authors: Ellen T. Johnson, Bradley K Yoder, Courtney J. Haycraft, Teodora Nicola, Kevin Roarty, Rosa SerraAbstract:IFT88 is a component of the intraflagellar transport complex required for formation and maintenance of cilia. Disruption of IFT88 results in depletion of cilia. The goal of the current study was to determine the role of primary cilia in ovarian function. Deletion of IFT88 in ovary using Cre-Lox recombination in mice resulted in a severe delay in mammary gland development including lack of terminal end bud structures, alterations in the estrous cycle, and impaired ovulation. Because estrogen drives the formation of end buds and Cre was expressed in the granulosa cells of the ovary, we tested the hypothesis that addition of estradiol to the mutant mice would compensate for defects in ovarian function and rescue the mammary gland phenotype. Mammary gland development including the formation of end bud structures resumed in mutant mice that were injected with estradiol. Together the results suggest that cilia are required for ovarian function.
