Nandadasa S, Kraft CM, Wang LW, O'Donnell A, Patel R, Gee HY, Grobe K, Cox TC, Hildebrandt F, Apte SS.
PMID: 30814516 | DOI: 10.1038/s41467-019-08520-7
Although hundreds of cytosolic or transmembrane molecules form the primary cilium, few secreted molecules are known to contribute to ciliogenesis. Here, homologous secreted metalloproteases ADAMTS9 and ADAMTS20 are identified as ciliogenesis regulators that act intracellularly. Secreted and furin-processed ADAMTS9 bound heparan sulfate and was internalized by LRP1, LRP2 and clathrin-mediated endocytosis to be gathered in Rab11 vesicles with a unique periciliary localization defined by super-resolution microscopy. CRISPR-Cas9 inactivation of ADAMTS9 impaired ciliogenesis in RPE-1 cells, which was restored by catalytically active ADAMTS9 or ADAMTS20 acting in trans, but not by their proteolytically inactive mutants. Their mutagenesis in mice impaired neural and yolk sac ciliogenesis, leading to morphogenetic anomalies resulting from impaired hedgehog signaling, which is transduced by primary cilia. In addition to their cognate extracellular proteolytic activity, ADAMTS9 and ADAMTS20 thus have an additional proteolytic role intracellularly, revealing an unexpected regulatory dimension in ciliogenesis.
J Invest Dermatol. 2015 Mar;135(3):701-9.
Chen J, Laclef C, Moncayo A, Snedecor ER, Yang N, Li L, Takemaru K, Paus R, Schneider-Maunoury S, Clark RA.
PMID: 25398052 | DOI: 10.1038/jid.2014.483.
The primary cilium is essential for skin morphogenesis through regulating the Notch, Wnt, and hedgehog signaling pathways. Prior studies on the functions of primary cilia in the skin were based on the investigations of genes that are essential for cilium formation. However, none of these ciliogenic genes has been linked to ciliopathy, a group of disorders caused by abnormal formation or function of cilia. To determine whether there is a genetic and molecular link between ciliopathies and skin morphogenesis, we investigated the role of RPGRIP1L, a gene mutated in Joubert (JBTS) and Meckel (MKS) syndromes, two severe forms of ciliopathy, in the context of skin development. We found that RPGRIP1L is essential for hair follicle morphogenesis. Specifically, disrupting the Rpgrip1l gene in mice resulted in reduced proliferation and differentiation of follicular keratinocytes, leading to hair follicle developmental defects. These defects were associated with significantly decreased primary cilium formation and attenuated hedgehog signaling. In contrast, we found that hair follicle induction and polarization and the development of interfollicular epidermis were unaffected. This study indicates that RPGRIP1L, a ciliopathy gene, is essential for hair follicle morphogenesis likely through regulating primary cilia formation and the hedgehog signaling pathway.
Li L, Grausam KB, Wang J, Lun MP, Ohli J, Lidov HG, Calicchio ML, Zeng E, Salisbury JL, Wechsler-Reya RJ, Lehtinen MK, Schüller U, Zhao H.
PMID: 26999738 | DOI: 10.1038/ncb3327
Aberrant Notch signalling has been linked to many cancers including choroid plexus (CP) tumours, a group of rare and predominantly paediatric brain neoplasms. We developed animal models of CP tumours, by inducing sustained expression of Notch1, that recapitulate properties of human CP tumours with aberrant NOTCH signalling. Whole-transcriptome and functional analyses showed that tumour cell proliferation is associated with Sonic Hedgehog (Shh) in the tumour microenvironment. Unlike CP epithelial cells, which have multiple primary cilia, tumour cells possess a solitary primary cilium as a result of Notch-mediated suppression of multiciliate differentiation. A Shh-driven signalling cascade in the primary cilium occurs in tumour cells but not in epithelial cells. Lineage studies show that CP tumours arise from monociliated progenitors in the roof plate characterized by elevated Notch signalling. Abnormal SHH signalling and distinct ciliogenesis are detected in human CP tumours, suggesting the SHH pathway and cilia differentiation as potential therapeutic avenues.
Lav, R;Krivanek, J;Anthwal, N;Tucker, AS;
PMID: 36931279 | DOI: 10.1016/j.stemcr.2023.02.004
Stem cell regulation plays a crucial role during development and homeostasis. Here, an essential source of Wnts from Gli1+ stem/progenitor cells was identified in the murine molar. Loss of Wnt production in Gli1+ apical stem/progenitor cells led to loss of Axin2 at the root apex, mis-regulation of SOX9, loss of BMP and Hh signaling, and truncation of root development. In the absence of Wnt signals, the root epithelium lost its integrity and epithelial identity. This phenotype could be partially mimicked by loss of Sox9 in the Gli1 population. Stabilization of Wnt signaling in the apical papilla led to rapid unordered differentiation of hard tissues and fragmentation of the epithelial root sheath. Wnt signaling from Gli1+ stem/progenitor cells, therefore, orchestrates root development, coordinating mesenchymal and epithelial interactions via SOX9 to regulate stem/progenitor cell expansion and differentiation. Our results demonstrate that disparate stem/progenitor cell populations are unified in their fundamental signaling interactions.
Xie, Y;Kuan, AT;Wang, W;Herbert, ZT;Mosto, O;Olukoya, O;Adam, M;Vu, S;Kim, M;Tran, D;Gómez, N;Charpentier, C;Sorour, I;Lacey, TE;Tolstorukov, MY;Sabatini, BL;Lee, WA;Harwell, CC;
PMID: 35196485 | DOI: 10.1016/j.celrep.2022.110416
Neuron-glia interactions play a critical role in the regulation of synapse formation and circuit assembly. Here we demonstrate that canonical Sonic hedgehog (Shh) pathway signaling in cortical astrocytes acts to coordinate layer-specific synaptic connectivity. We show that the Shh receptor Ptch1 is expressed by cortical astrocytes during development and that Shh signaling is necessary and sufficient to promote the expression of genes involved in regulating synaptic development and layer-enriched astrocyte molecular identity. Loss of Shh in layer V neurons reduces astrocyte complexity and coverage by astrocytic processes in tripartite synapses; conversely, cell-autonomous activation of Shh signaling in astrocytes promotes cortical excitatory synapse formation. Furthermore, Shh-dependent genes Lrig1 and Sparc distinctively contribute to astrocyte morphology and synapse formation. Together, these results suggest that Shh secreted from deep-layer cortical neurons acts to specialize the molecular and functional features of astrocytes during development to shape circuit assembly and function.
Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research
Coveney, CR;Samvelyan, HJ;Miotla-Zarebska, J;Carnegie, J;Chang, E;Corrin, CJ;Coveney, T;Stott, B;Parisi, I;Duarte, C;Vincent, TL;Staines, KA;Wann, AKT;
PMID: 35038201 | DOI: 10.1002/jbmr.4502
In comparison to our understanding of endochondral ossification, much less is known about the coordinated arrest of growth defined by the narrowing and fusion of the cartilaginous growth plate. Throughout the musculoskeletal system, appropriate cell and tissue responses to mechanical force delineate morphogenesis and ensure lifelong health. It remains unclear how mechanical cues are integrated into many biological programmes including those coordinating the ossification of the adolescent growth plate at the cessation of growth. Primary cilia are microtubule-based organelles tuning a range of cell activities, including signalling cascades activated or modulated by extracellular biophysical cues. Cilia have been proposed to directly facilitate cell mechanotransduction. To explore the influence of primary cilia in the mouse adolescent limb, we conditionally targeted the ciliary gene Intraflagellar transport protein 88 (Ift88fl/fl ) in the juvenile and adolescent skeleton using a cartilage-specific, inducible, Cre (AggrecanCreERT2 Ift88fl/fl ). Deletion of IFT88 in cartilage, which reduced ciliation in the growth plate, disrupted chondrocyte differentiation, cartilage resorption and mineralisation. These effects were largely restricted to peripheral tibial regions beneath the load-bearing compartments of the knee. These regions were typified by an enlarged population of hypertrophic chondrocytes. While normal patterns of hedgehog signalling were maintained, targeting IFT88 inhibited hypertrophic chondrocyte VEGF expression and downstream vascular recruitment, osteoclastic activity and the replacement of cartilage with bone. In control mice, increases to physiological loading also impair ossification in the peripheral growth plate, mimicking the effects of IFT88 deletion. Limb immobilisation inhibited changes to VEGF expression and epiphyseal morphology in Ift88cKO mice, indicating the effects of depletion of IFT88 in the adolescent growth plate are mechano-dependent. We propose that during this pivotal phase in adolescent skeletal maturation, ciliary IFT88 protects uniform, coordinated ossification of the growth plate from an otherwise disruptive heterogeneity of physiological mechanical forces. This article is protected by
Modern pathology : an official journal of the United States and Canadian Academy of Pathology, Inc
Menge, TD;Durgin, JS;Hrycaj, SM;Brent, AA;Patel, RM;Harms, PW;Fullen, DR;Chan, MP;Bresler, SC;
PMID: 37391171 | DOI: 10.1016/j.modpat.2023.100265
Basal cell carcinoma (BCC) is the most common human malignancy and is a leading cause of non-melanoma skin cancer-related morbidity. BCC has several histologic mimics which may have treatment and prognostic implications. Furthermore, BCC may show alternative differentiation toward a variety of cutaneous structures. The vast majority of BCCs harbor mutations in the hedgehog signaling pathway, resulting in increased expression of the GLI family of transcription factors. GLI1 immunohistochemistry has been shown to discriminate between several tumor types but demonstrates high background signal and lack of specificity. In this study we evaluate the utility of GLI1 RNA chromogenic in situ hybridization (CISH) as a novel method of distinguishing between BCC and other epithelial neoplasms. Expression of GLI1 by RNA CISH was retrospectively evaluated in a total of 220 cases including 60 BCCs, 37 squamous cell carcinomas (SCCs) including conventional, basaloid, and HPV-associated tumors, 16 sebaceous neoplasms, 10 Merkel cell carcinomas (MCCs), 58 benign follicular tumors, and 39 ductal tumors. The threshold for positivity was determined to be greater than or equal to 3 GLI1 signals in at least 50% of tumor cells. Positive GLI1 expression was identified in 57/60 BCCs including metastatic BCC, collision lesions with SCC, and BCCs with squamous, ductal, or clear cell differentiation or with other unusual features, as compared to 1/37 SCCs, 0/11 sebaceous carcinomas, 0/5 sebaceomas, 1/10 MCCs, 0/39 ductal tumors, and 28/58 follicular tumors. With careful evaluation, GLI1 RNA CISH is highly sensitive (95%) and specific (98%) in distinguishing between BCC and non-follicular epithelial neoplasms. However, GLI1 CISH is not specific for distinguishing BCC from most benign follicular tumors. Overall, detection of GLI1RNA by CISH may be a useful tool for precise classification of histologically challenging basaloid tumors, particularly in the setting of small biopsy specimens, metaplastic differentiation, or metastatic disease.
Sánchez-Danés A, Larsimont JC, Liagre M, Muñoz-Couselo E, Lapouge G, Brisebarre A, Dubois C, Suppa M, Sukumaran V, Del Marmol V, Tabernero J, Blanpain C.
PMID: 30297799 | DOI: 10.1038/s41586-018-0603-3
Basal cell carcinoma (BCC) is the most frequent cancer in humans and results from constitutive activation of the Hedgehog pathway1. Several Smoothened inhibitors are used to treat Hedgehog-mediated malignancies, including BCC and medulloblastoma2. Vismodegib, a Smoothened inhibitor, leads to BCC shrinkage in the majority of patients with BCC3, but the mechanism by which it mediates BCC regression is unknown. Here we used two genetically engineered mouse models of BCC4 to investigate the mechanisms by which inhibition of Smoothened mediates tumour regression. We found that vismodegib mediates BCC regression by inhibiting a hair follicle-like fate and promoting the differentiation of tumour cells. However, a small population of tumour cells persists and is responsible for tumour relapse following treatment discontinuation, mimicking the situation found in humans5. In both mouse and human BCC, this persisting, slow-cycling tumour population expresses LGR5 and is characterized by active Wnt signalling. Combining Lgr5 lineage ablation or inhibition of Wnt signalling with vismodegib treatment leads to eradication of BCC. Our results show that vismodegib induces tumour regression by promoting tumour differentiation, and demonstrates that the synergy between Wnt and Smoothened inhibitors is a clinically relevant strategy for overcoming tumour relapse in BCC.
Kaucka, M;Joven Araus, A;Tesarova, M;Currie, JD;Boström, J;Kavkova, M;Petersen, J;Yao, Z;Bouchnita, A;Hellander, A;Zikmund, T;Elewa, A;Newton, PT;Fei, JF;Chagin, AS;Fried, K;Tanaka, EM;Kaiser, J;Simon, A;Adameyko, I;
PMID: 36376278 | DOI: 10.1038/s41467-022-34266-w
There are major differences in duration and scale at which limb development and regeneration proceed, raising the question to what extent regeneration is a recapitulation of development. We address this by analyzing skeletal elements using a combination of micro-CT imaging, molecular profiling and clonal cell tracing. We find that, in contrast to development, regenerative skeletal growth is accomplished based entirely on cartilage expansion prior to ossification, not limiting the transversal cartilage expansion and resulting in bulkier skeletal parts. The oriented extension of salamander cartilage and bone appear similar to the development of basicranial synchondroses in mammals, as we found no evidence for cartilage stem cell niches or growth plate-like structures during neither development nor regeneration. Both regenerative and developmental ossification in salamanders start from the cortical bone and proceeds inwards, showing the diversity of schemes for the synchrony of cortical and endochondral ossification among vertebrates.
Manti, PG;Darbellay, F;Leleu, M;Coughlan, AY;Moret, B;Cuennet, J;Droux, F;Stoudmann, M;Mancini, GF;Hautier, A;Sordet-Dessimoz, J;Vincent, SD;Testa, G;Cossu, G;Barrandon, Y;
PMID: 36289911 | DOI: 10.3390/biomedicines10102647
Prdm1 mutant mice are one of the rare mutant strains that do not develop whisker hair follicles while still displaying a pelage. Here, we show that Prdm1 is expressed at the earliest stage of whisker development in clusters of mesenchymal cells before placode formation. Its conditional knockout in the murine soma leads to the loss of expression of Bmp2, Shh, Bmp4, Krt17, Edar, and Gli1, though leaving the β-catenin-driven first dermal signal intact. Furthermore, we show that Prdm1 expressing cells not only act as a signaling center but also as a multipotent progenitor population contributing to the several lineages of the adult whisker. We confirm by genetic ablation experiments that the absence of macro vibrissae reverberates on the organization of nerve wiring in the mystacial pads and leads to the reorganization of the barrel cortex. We demonstrate that Lef1 acts upstream of Prdm1 and identify a primate-specific deletion of a Lef1 enhancer named Leaf. This loss may have been significant in the evolutionary process, leading to the progressive defunctionalization and disappearance of vibrissae in primates.
Proceedings of the National Academy of Sciences of the United States of America
Chen, L;Li, Y;Song, Z;Xue, S;Liu, F;Chang, X;Wu, Y;Duan, X;Wu, H;
PMID: 35969743 | DOI: 10.1073/pnas.2202821119
Sonic hedgehog (Shh) signaling plays a critical role in regulating cerebellum development by maintaining the physiological proliferation of granule neuron precursors (GNPs), and its dysregulation leads to the oncogenesis of medulloblastoma. O-GlcNAcylation (O-GlcNAc) of proteins is an emerging regulator of brain function that maintains normal development and neuronal circuitry. Here, we demonstrate that O-GlcNAc transferase (OGT) in GNPs mediate the cerebellum development, and the progression of the Shh subgroup of medulloblastoma. Specifically, OGT regulates the neurogenesis of GNPs by activating the Shh signaling pathway via O-GlcNAcylation at S355 of GLI family zinc finger 2 (Gli2), which in turn promotes its deacetylation and transcriptional activity via dissociation from p300, a histone acetyltransferases. Inhibition of OGT via genetic ablation or chemical inhibition improves survival in a medulloblastoma mouse model. These data uncover a critical role for O-GlcNAc signaling in cerebellar development, and pinpoint a potential therapeutic target for Shh-associated medulloblastoma.
Minatoguchi, S;Saito, S;Furuhashi, K;Sawa, Y;Okazaki, M;Shimamura, Y;Kaihan, AB;Hashimoto, Y;Yasuda, Y;Hara, A;Mizutani, Y;Ando, R;Kato, N;Ishimoto, T;Tsuboi, N;Esaki, N;Matsuyama, M;Shiraki, Y;Kobayashi, H;Asai, N;Enomoto, A;Maruyama, S;
PMID: 35354870 | DOI: 10.1038/s41598-022-09331-5
Perivascular mesenchymal cells (PMCs), which include pericytes, give rise to myofibroblasts that contribute to chronic kidney disease progression. Several PMC markers have been identified; however, PMC heterogeneity and functions are not fully understood. Here, we describe a novel subset of renal PMCs that express Meflin, a glycosylphosphatidylinositol-anchored protein that was recently identified as a marker of fibroblasts essential for cardiac tissue repair. Tracing the lineage of Meflin+ PMCs, which are found in perivascular and periglomerular areas and exhibit renin-producing potential, showed that they detach from the vasculature and proliferate under disease conditions. Although the contribution of Meflin+ PMCs to conventional α-SMA+ myofibroblasts is low, they give rise to fibroblasts with heterogeneous α-SMA expression patterns. Genetic ablation of Meflin+ PMCs in a renal fibrosis mouse model revealed their essential role in collagen production. Consistent with this, human biopsy samples showed that progressive renal diseases exhibit high Meflin expression. Furthermore, Meflin overexpression in kidney fibroblasts promoted bone morphogenetic protein 7 signals and suppressed myofibroblastic differentiation, implicating the roles of Meflin in suppressing tissue fibrosis. These findings demonstrate that Meflin marks a PMC subset that is functionally distinct from classic pericytes and myofibroblasts, highlighting the importance of elucidating PMC heterogeneity.
