FRINK Linked Data Fragments server

Matches in SemOpenAlex for { <https://semopenalex.org/work/W2912306611> ?p ?o ?g. }

• W2912306611 abstract "Report18 February 2019Open Access Source DataTransparent process MET mutation causes muscular dysplasia and arthrogryposis Hang Zhou Hang Zhou Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Provincial Key Laboratory of Orthopedics and Traumatology, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Province Center for Peripheral Nerve Tissue Engineering and Technology Research, Guangzhou, Guangdong, China Guangdong Province Engineering Laboratory for Soft Tissue Biofabrication, Guangzhou, Guangdong, China Search for more papers by this author Chengjie Lian Chengjie Lian Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Provincial Key Laboratory of Orthopedics and Traumatology, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Province Center for Peripheral Nerve Tissue Engineering and Technology Research, Guangzhou, Guangdong, China Guangdong Province Engineering Laboratory for Soft Tissue Biofabrication, Guangzhou, Guangdong, China Search for more papers by this author Tingting Wang Tingting Wang Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Xiaoming Yang Xiaoming Yang Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Caixia Xu Caixia Xu Research Centre for Translational Medicine, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Deying Su Deying Su Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Shuhui Zheng Shuhui Zheng Research Centre for Translational Medicine, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Xiangyu Huang Xiangyu Huang Department of Stomatology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong, China Search for more papers by this author Zhiheng Liao Zhiheng Liao Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Taifeng Zhou Taifeng Zhou Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Xianjian Qiu Xianjian Qiu Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Yuyu Chen Yuyu Chen Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Bo Gao Bo Gao Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Yongyong Li Yongyong Li Research Centre for Translational Medicine, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Xudong Wang Xudong Wang Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Guoling You Guoling You Department of Laboratory Medicine, Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, Shanghai, China Search for more papers by this author Qihua Fu Qihua Fu Department of Laboratory Medicine, Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, Shanghai, China Search for more papers by this author Christina Gurnett Christina Gurnett Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA Department of Neurology, Washington University, St. Louis, MO, USA Department of Pediatrics, Washington University, St. Louis, MO, USA Search for more papers by this author Dongsheng Huang Dongsheng Huang Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Peiqiang Su Corresponding Author Peiqiang Su [email protected] orcid.org/0000-0001-5391-7558 Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Provincial Key Laboratory of Orthopedics and Traumatology, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Province Center for Peripheral Nerve Tissue Engineering and Technology Research, Guangzhou, Guangdong, China Guangdong Province Engineering Laboratory for Soft Tissue Biofabrication, Guangzhou, Guangdong, China Search for more papers by this author Hang Zhou Hang Zhou Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Provincial Key Laboratory of Orthopedics and Traumatology, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Province Center for Peripheral Nerve Tissue Engineering and Technology Research, Guangzhou, Guangdong, China Guangdong Province Engineering Laboratory for Soft Tissue Biofabrication, Guangzhou, Guangdong, China Search for more papers by this author Chengjie Lian Chengjie Lian Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Provincial Key Laboratory of Orthopedics and Traumatology, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Province Center for Peripheral Nerve Tissue Engineering and Technology Research, Guangzhou, Guangdong, China Guangdong Province Engineering Laboratory for Soft Tissue Biofabrication, Guangzhou, Guangdong, China Search for more papers by this author Tingting Wang Tingting Wang Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Xiaoming Yang Xiaoming Yang Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Caixia Xu Caixia Xu Research Centre for Translational Medicine, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Deying Su Deying Su Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Shuhui Zheng Shuhui Zheng Research Centre for Translational Medicine, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Xiangyu Huang Xiangyu Huang Department of Stomatology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong, China Search for more papers by this author Zhiheng Liao Zhiheng Liao Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Taifeng Zhou Taifeng Zhou Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Xianjian Qiu Xianjian Qiu Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Yuyu Chen Yuyu Chen Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Bo Gao Bo Gao Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Yongyong Li Yongyong Li Research Centre for Translational Medicine, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Xudong Wang Xudong Wang Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Guoling You Guoling You Department of Laboratory Medicine, Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, Shanghai, China Search for more papers by this author Qihua Fu Qihua Fu Department of Laboratory Medicine, Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, Shanghai, China Search for more papers by this author Christina Gurnett Christina Gurnett Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA Department of Neurology, Washington University, St. Louis, MO, USA Department of Pediatrics, Washington University, St. Louis, MO, USA Search for more papers by this author Dongsheng Huang Dongsheng Huang Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Search for more papers by this author Peiqiang Su Corresponding Author Peiqiang Su [email protected] orcid.org/0000-0001-5391-7558 Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Provincial Key Laboratory of Orthopedics and Traumatology, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China Guangdong Province Center for Peripheral Nerve Tissue Engineering and Technology Research, Guangzhou, Guangdong, China Guangdong Province Engineering Laboratory for Soft Tissue Biofabrication, Guangzhou, Guangdong, China Search for more papers by this author Author Information Hang Zhou1,2,3,4,‡, Chengjie Lian1,2,3,4,‡, Tingting Wang1, Xiaoming Yang1, Caixia Xu5, Deying Su1, Shuhui Zheng5, Xiangyu Huang6, Zhiheng Liao1, Taifeng Zhou1, Xianjian Qiu7, Yuyu Chen1, Bo Gao7, Yongyong Li5, Xudong Wang7, Guoling You8, Qihua Fu8, Christina Gurnett9,10,11, Dongsheng Huang7 and Peiqiang Su *,1,2,3,4,12 1Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China 2Guangdong Provincial Key Laboratory of Orthopedics and Traumatology, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China 3Guangdong Province Center for Peripheral Nerve Tissue Engineering and Technology Research, Guangzhou, Guangdong, China 4Guangdong Province Engineering Laboratory for Soft Tissue Biofabrication, Guangzhou, Guangdong, China 5Research Centre for Translational Medicine, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China 6Department of Stomatology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong, China 7Department of Spine Surgery, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China 8Department of Laboratory Medicine, Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, Shanghai, China 9Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA 10Department of Neurology, Washington University, St. Louis, MO, USA 11Department of Pediatrics, Washington University, St. Louis, MO, USA 12Present address: Department of Orthopaedic Surgery, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China ‡These authors contributed equally to this work *Corresponding author. Tel: +862087755766 6236; E-mail: [email protected] EMBO Mol Med (2019)11:e9709https://doi.org/10.15252/emmm.201809709 PDFDownload PDF of article text and main figures. Peer ReviewDownload a summary of the editorial decision process including editorial decision letters, reviewer comments and author responses to feedback. ToolsAdd to favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract Arthrogryposis is a group of phenotypically and genetically heterogeneous disorders characterized by congenital contractures of two or more parts of the body; the pathogenesis and the causative genes of arthrogryposis remain undetermined. We examined a four-generation arthrogryposis pedigree characterized by camptodactyly, limited forearm supination, and loss of myofibers in the forearms and hands. By using whole-exome sequencing, we confirmed MET p.Y1234C mutation to be responsible for arthrogryposis in this pedigree. MET p.Y1234C mutation caused the failure of activation of MET tyrosine kinase. A Met p.Y1232C mutant mouse model was established. The phenotypes of homozygous mice included embryonic lethality and complete loss of muscles that originated from migratory precursors. Heterozygous mice were born alive and showed reduction of the number of myofibers in both appendicular and axial muscles. Defective migration of muscle progenitor cells and impaired proliferation of secondary myoblasts were proven to be responsible for the skeletal muscle dysplasia of mutant mice. Overall, our study shows MET to be a causative gene of arthrogryposis and MET mutation could cause skeletal muscle dysplasia in human beings. Synopsis In this study, MET was identified as a causative gene for arthrogryposis, characterized by congenital contractures of two or more parts of the body. The mutant MET p.Y1234C impaired activation of MET tyrosine kinase, resulting in muscular dysplasia of the limbs. MET p.Y1234C mutation was identified in a four-generation arthrogryposis pedigree. Muscular dysplasia of the upper arms and hands was observed in the arthrogryposis patients. In vitro, MET p.Y1234C was responsible for the failure in MET tyrosine kinase activation. In vivo, Met p.Y1232C led to defective migration of muscle progenitor cells and impaired proliferation of secondary myoblasts. Introduction Arthrogryposis is a group of disorders characterized by congenital joint contractures that mainly involve two or more areas of the body (Bamshad et al, 2009; Bayram et al, 2016) and affects approximately 1 in 3,000 newborns (Bayram et al, 2016). Structural and functional disorders of skeletal muscles are the most common reason for arthrogryposis (Toydemir et al, 2006). Arthrogryposis is a group of disorders with high clinical and genetic heterogeneity. Variants in more than 220 genes have been found to be associated with arthrogryposis (Narkis et al, 2007; Drury et al, 2014; Hunter et al, 2015; Bayram et al, 2016). However, the molecular etiology still remains unclear in a large number of cases of arthrogryposis. Further studies to identify causative genes and pathogenic mechanisms are needed. MET belongs to the receptor tyrosine kinase family, and it is encoded by the MET proto-oncogene, receptor tyrosine kinase (MET, MIM:164860) gene. The biological effects exerted by MET are triggered by the stimulation of its only ligand, hepatocyte growth factor (HGF; Trusolino et al, 2010). Upon ligand binding, MET is autophosphorylated on tyrosine (Y)-1234/1235 in the activation loop of the MET catalytic domain. Y-1234/1235 phosphorylation is required for the activation of MET kinase and subsequent phosphorylation of other tyrosine sites of MET, including Y-1003 in the juxtamembrane domain, and Y-1349 and Y-1356 in the carboxyl terminus (Sangwan et al, 2008). Phosphorylated Y-1349 and Y-1356 serve as multifunctional binding sites for GAB1, GRB2, PI3K, and other downstream substrates (Birchmeier et al, 2003). The HGF-MET signal plays a vital role in regulating the development of skeletal muscle, placenta, and liver during embryogenesis (Birchmeier et al, 1997; Haines et al, 2004; Ueno et al, 2013). In skeletal muscle development, MET has been demonstrated not only to be crucial for the migration of muscle progenitor cells into the limbs, tongue, and diaphragm, but also to be necessary for the proliferation of secondary myoblasts in the trunk (Maina et al, 1996). Herein, we recruited a rare four-generation Chinese arthrogryposis pedigree with only upper limb involvement, and we found the MET c.A3701G (p.Y1234C; Refseq NM_000245.2) mutation to be responsible for the pathogenesis of arthrogryposis in this pedigree. MET p.Y1234C mutation was shown to cause the dysfunction of phosphorylation and tyrosine kinase activity of MET in vitro. We established a Met c.A3695G (p.Y1232C; Refseq NM_008591.2) mutant mouse model, and the defective migration of myogenic progenitor cells and impaired proliferation of secondary myoblasts were demonstrated to be responsible for the disturbed muscle development. Results Clinical presentation of patients from a large arthrogryposis family A four-generation Chinese family presented with completely penetrant, autosomal dominant arthrogryposis characterized mainly by camptodactyly (Fig 1A). All patients in this family had camptodactyly, and seven patients had camptodactyly, absent flexion crease, and limited forearm supination (Fig 1B; Table EV1). Signs of lower limb, and facial and spinal involvement were absent. Since interphalangeal joints and carpal joints were both affected in seven individuals, a diagnosis of arthrogryposis involving only the upper limbs was made. Figure 1. MET p.Y1234C mutation caused arthrogryposis in a four-generation Chinese family A. The MET p.Y1234C mutation segregated with disease phenotypes in the arthrogryposis family. Filled symbols denote affected individuals, open symbols indicate unaffected individuals, and symbols with slashes represent decreased individuals. Asterisks indicate a mutation is present, # means wild-type. B. Phenotypes of affected individuals. Camptodactyly, absent flexion crease, and limited forearm supination were observed. C–E. T1-weighted MRI scan on upper limbs of subject IV:7. (C) The pronator quadratus absence of affected side was indicated by a red arrow. (D) No difference was found in palmar muscles. (E) Loss of lumbricalis and interosseous muscles of fifth finger of affected side was indicated by a red arrow. F–H. T1-weighted MRI scan on upper limbs of subject IV:8. (F) Increased epimysial fat was indicated by a red arrow. (G) Completely loss of thenar eminences of both hands was indicated by red arrows. (H) Loss of radial lumbricalis and interosseous muscles of both hands was indicated by red arrows. I. The MET variants by Sanger sequencing were indicated by a red arrow. J. 293T cells were transfected with FLAG-tagged MET/METMut/Vector plasmids, and 48 h post-transfection, cells were treated with 10 ng/ml recombinant human HGF for 1 h. Then, MET/METMut protein purification and tyrosine kinase assay were conducted. Western blot pictures representative of n = 3 experiments. METMut means p.Y1234C mutant MET. EGFR means epidermal growth factor receptor and serves as a positive control. NC means negative control. Source data are available online for this figure. Source Data for Figure 1J [emmm201809709-sup-0004-SDataFig1J.pdf] Download figure Download PowerPoint Subject IV:7 is a patient with unilateral camptodactyly, absent flexion crease, and limited forearm supination. Severe pronator quadratus aplasia of affected forearm was observed through magnetic resonance imaging (MRI; Fig 1C). For the palmar muscles, loss of lumbricalis and interosseous muscles of fifth finger of affected side was found (Fig 1D and E). Subject IV:8 is a patient with severe bilateral camptodactyly, absent flexion crease, and limited forearm supination. MRI scan showed increased epimysial fat among muscle compartments (Fig 1F), complete loss of thenar eminences, the radial lumbricalis, and interosseous muscles of both hands (Fig 1G and H). The lumbricalis muscle of subject IV:8 showed varying fiber size and more centrally located nuclei than control lumbricalis muscle from an age- and gender-matched person without muscular dysplasia (Fig EV1A). No bone abnormality was observed in arthrogryposis patients of this pedigree (Fig EV1B). Overall, a diagnosis of arthrogryposis involving only the upper limbs was made, and muscular dysplasia was observed in the affected forearms and hands of these patients. Click here to expand this figure. Figure EV1. Histological study and X-ray scan of arthrogryposis patients A. HE staining of lumbrical samples from patient IV:8 and the age-/gender-matched control was conducted. Centrally located muscles were indicated by a black arrow. Scale bars, 100 μm. B. X-ray scan of the patients’ hands was conducted. Download figure Download PowerPoint Whole-exome sequencing identified MET as a disease-causing gene of arthrogryposis To identify arthrogryposis-predisposing variants, whole-exome sequencing was initially performed on four affected individuals and one healthy member of this arthrogryposis pedigree (Appendix Table S1). As previously reported (Gao et al, 2017), we annotated and filtered variants, and kept variants that were novel in dbSNP. Polyphen-2, Mutation Taster, and Genomic Evolutionary Rate Profiling (GERP) were then used to predict the potential functional effects of these mutations, which yielded two candidate SNVs, c.A3701G in the MET; c.G2074A (Refseq NM_006019) in TCIRG1 (MIM:604592). By using Sanger sequencing, we excluded the SNV on TCIRG1 because MET c.A3701G turned out to be the only one which co-segregated with disease phenotypes in this family (Fig 1I, Appendix Table S2). MET p.Y1234C mutation caused dysfunction of the phosphorylation and tyrosine kinase activity of MET The influence of p.Y1234C mutation on the function of MET was studied (Fig EV2A), and HGF treatment was shown to be unable to phosphorylate the Y-1234/1235, Y-1349, and Y-1356 sites of mutant MET receptor (Fig EV2B–D), suggesting MET mutation impaired the activation of MET receptor. Moreover, the tyrosine kinase activity of mutant MET was shown to decrease dramatically (Fig 1J). Click here to expand this figure. Figure EV2. MET p.Y1234C mutation caused the dysfunction of the phosphorylation of MET protein A. Upper panel shows the protein structure of MET. Green bars represent tyrosine phosphorylation sites, and red arrow indicates the mutation that was observed in our arthrogryposis pedigree. Lower panel shows phylogenetic conservation of mutated residues and homology among different species of MET gene, and cons means conservation. B–D. 293T cells were transfected with pCMV-C-FLAG-MET, pCMV-C-FLAG-METMut, or pCMV-C-FLAG-Vector, and 48 h post-transfection, cells were treated with 10 ng/ml recombinant human HGF for 1 h. Immunoprecipitation was carried out with anti-FLAG antibody and followed by immunoblotting with anti-FLAG antibody and anti-p-METY1234/1235 (B), anti-p-METY1349 (C), and anti-p-METY1356 (D) antibodies. Western blot pictures representative of n = 3 experiments. Source data are available online for this figure. Download figure Download PowerPoint Met mutation resulted in the reduction of limb myofibers in transgenic mouse model To determine the mechanism by which MET mutation causes arthrogryposis, a Met p.Y1232C (which was identical to p.Y1234C in human beings) mutant mouse model was constructed. No homozygous newborns were found. The ratio of homozygous embryos started to decline since E14.5, and E16.5 was the latest time that homozygous embryos could survive, which was consistent with Met null mutants (Schmidt et al, 1995). The failure of placental development in homozygotes might be responsible for the death of embryos in utero (Ueno et al, 2013). Heterozygotes were smaller than wild-type individuals at birth (Fig 2A and B). Compared with wild-type newborns, the mean number of myofibers of paraspinal muscles, forelimbs, hindlimbs, and hands of heterozygotes reduced by 14, 55, 29, and 93%, respectively, while the foot muscles remained normal (Fig 2C–H). To figure out whether the myofibrils were affected, the gastrocnemius of wild-type and heterozygous newborns was tested with transmission electron microscope (TEM). However, no abnormality of the structure of myofibrils was found (Fig 2I). Figure 2. Met mutation caused the reduction of both appendicular and paraspinal muscles of P0 heterozygotes A. Gross appearance of wild-type and heterozygous newborns. B. Graph showing weight of P0 mice. Bars show mean ± SD. Sample size: WT (n = 4) and Hetero (n = 5). *P < 0.05, by one-way ANOVA and followed by Dunnett's post hoc test. C–G. Sections of spine, forelimbs, hindlimbs, hand, and foot from wild types and heterozygotes at P0 were conducted with HE staining and immunofluorescence staining using anti-myosin heavy chain antibody. Scale bars, 100 μm. H. The mean number of myofibers in the muscles of the spine, forelimb, hindlimb, hand, and foot was qualified. Bars show mean ± SD. NS means no statistic significance. *P < 0.05, **P < 0.01, by two-tailed independent Student's t-test. I. Transmission electron microscope analysis of gastrocnemius from wild-type and heterozygous newborns. Black arrows denote mitochondria. Scale bars, 2 μm. WT means wild types, Hetero means heterozygotes. Data information: In (C–H), n = 3. Download figure Download PowerPoint Met mutation affected the migration of muscle progenitor cells To determine how Met mutation affected muscle development, its effect on muscle progenitor cells’ migrating out of dermomyotome was examined firstly. Using in situ hybridization, expression of Pax3 (MIM: 606597) and Met was assessed in embryonic limbs and dermomyotome at the end of migration (E10.5). In dermomyotome, heterozygous and homozygous embryos showed more Pax3-positive (Pax3+) and Met-positive (Met+) cells than wild types (Figs 3A and EV3). Pax3+ and Met+ cells were absent from homozygous limbs, while the number of Pax3+ and Met+ cells was markedly lower in heterozygous limbs, suggesting Met mutation impaired muscle progenitor cells’ migration out of dermomyotome to the limb (Figs 3A and EV3). Figure 3. Met mutation led to the defects in the migration of muscle progenitor cells and impaired proliferation of secondary myoblasts A. In situ hybridization of E10.5 embryos using Pax3 probe. Pax3 expression (brown signal) was observed in limb bud and dermomyotome (DM), respectively. Cells labeled with Pax3 were indicated by a red arrow. Scale bars, 200 μm. B. Gross appearance of E14.5 embryos of indicated genotype. Scale bars, 2.5 mm. C. HE staining of forelimb, hindlimb, and paraspinal muscle of E14.5 embryos. Scale bars, 100 μm. D. Gross appearance of E16.5 embryos of indicated genotype. Scale bars, 2.5 mm. E. HE staining of forelimb, hindlimb, and paraspinal muscle of E16.5 embryos. n = 3, scale bars, 200 μm. F. Anti-Ki67 antibody was used to label proliferative myoblasts (red fluorescence) with DAPI-labeled nuclei (blue fluorescence) in paraspinal muscle. Scale bar, 25 μm. G. Bar graph showing statistical analysis of positive rate of Ki67-labeled nuclei, n = 3 with more than 150 cells analyzed per n, **P < 0.01, by chi-square test (χ2 test). Bars show mean ± SD. WT means wild types, Hetero means heterozygotes, and Homo means homozygotes. Download figure Download PowerPoint Click here to expand this figure. Figure EV3. In situ hybridization of E10.5 embryos using Met probeMet expression (brown signal) was detected through in situ hybridization in dermomyotome (DM) and limb bud of E10.5 embryos from all three genotypes, n = 3. Scale bars, 200 μm. WT means wild types, Hetero means heterozygotes, and Homo means homozygotes. Download figure Download PowerPoint Met mutation had no effect on primary myogenesis of embryonic muscle development The decreased number of muscle fibers in axial muscles of P0 heterozygotes suggested that Met mutation might also affect the myoblasts which do not undergo migration. The body size and weight of E14.5 (the very end of primary myogenesis) homozygous embryos were lower than that of wild types (Figs 3B and EV4A). For muscles that derive from migratory precursors, remarkable decrease of muscle fibers was observed in heterozygotes, while the homozygotes showed a complete loss of muscle fibers in these areas (limbs, front tongue, and diaphragm, Figs 3C and EV4B). For muscles that do not originate from the migratory precursors, no obvious difference was observed among three genotypes at E14.5 (paraspinal muscle and intercostal muscle, Figs 3C and EV4B). To evaluate the proliferation and apoptosis of primary myoblasts, Ki67 staining and TUNEL assay were conducted in paraspinal muscle, and no significant difference was found among all three genotypes (Fig EV4C–F), suggesting that Met mutation had no effect on primary myogenesis. Click here to expand this figure. Figure EV4. Met mutation had no effect on the formation of primary muscle fiber A. Graph indicates the weight of E14.5 embryos of three genotypes. Bars show mean ± SD. Sample size: WT (n = 5), Hetero (n = 5), and Homo (n = 5). *P < 0.05, by one-way ANOVA and followed by Dunnett's post hoc test. B. HE staining of front tongue, diaphragm, and rib from E14.5 embryos was performed. Front tongue was denoted by black arrow, and diaphragm was indicated by a red arrow. Scale bars, 100 μm. C. Anti-Ki67 antibody was used to label proliferative myoblasts (red fluorescence) with DAPI-labeled nuclei (blue fluorescence) in paraspinal muscles of E14.5 embryos. Scale bars, 25 μm. D. TUNEL assay was conducted to reveal apoptotic myoblasts (green fluorescence) with DAPI-labeled nuclei (blue fluorescence) in paraspinal muscles of E14.5 embryos. Scale bars, 25 μm. E. Positive rate of Ki67-labeled nuclei in (C) was quantified. Bars show mean ± SD. F. Positive rate of TUNEL-labeled nuclei in" @default.
• W2912306611 created "2019-02-21" @default.
• W2912306611 creator A5000863572 @default.
• W2912306611 creator A5003760398 @default.
• W2912306611 creator A5005887396 @default.
• W2912306611 creator A5012395068 @default.
• W2912306611 creator A5019290469 @default.
• W2912306611 creator A5036046257 @default.
• W2912306611 creator A5036492073 @default.
• W2912306611 creator A5037878631 @default.
• W2912306611 creator A5043951146 @default.
• W2912306611 creator A5049344677 @default.
• W2912306611 creator A5063610143 @default.
• W2912306611 creator A5067215726 @default.
• W2912306611 creator A5072884120 @default.
• W2912306611 creator A5074200539 @default.
• W2912306611 creator A5076587915 @default.
• W2912306611 creator A5081093007 @default.
• W2912306611 creator A5085249009 @default.
• W2912306611 creator A5086359066 @default.
• W2912306611 creator A5087886690 @default.
• W2912306611 creator A5088321212 @default.
• W2912306611 date "2019-02-18" @default.
• W2912306611 modified "2023-10-15" @default.
• W2912306611 title "<i><scp>MET</scp></i>mutation causes muscular dysplasia and arthrogryposis" @default.
• W2912306611 cites W1499575114 @default.
• W2912306611 cites W1944377017 @default.
• W2912306611 cites W1990244564 @default.
• W2912306611 cites W2022527563 @default.
• W2912306611 cites W2038103542 @default.
• W2912306611 cites W2049731453 @default.
• W2912306611 cites W2057391791 @default.
• W2912306611 cites W2069421644 @default.
• W2912306611 cites W2073083344 @default.
• W2912306611 cites W2089502371 @default.
• W2912306611 cites W2094745160 @default.
• W2912306611 cites W2097316073 @default.
• W2912306611 cites W2107061457 @default.
• W2912306611 cites W2131073955 @default.
• W2912306611 cites W2132831751 @default.
• W2912306611 cites W2133535400 @default.
• W2912306611 cites W2136390065 @default.
• W2912306611 cites W2139685274 @default.
• W2912306611 cites W2142289251 @default.
• W2912306611 cites W2171370093 @default.
• W2912306611 cites W2233137135 @default.
• W2912306611 cites W2476789590 @default.
• W2912306611 cites W2734858404 @default.
• W2912306611 cites W2746520029 @default.
• W2912306611 cites W2890618054 @default.
• W2912306611 doi "https://doi.org/10.15252/emmm.201809709" @default.
• W2912306611 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/6404111" @default.
• W2912306611 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/30777867" @default.
• W2912306611 hasPublicationYear "2019" @default.
• W2912306611 type Work @default.
• W2912306611 sameAs 2912306611 @default.
• W2912306611 citedByCount "2" @default.
• W2912306611 countsByYear W29123066112020 @default.
• W2912306611 countsByYear W29123066112022 @default.
• W2912306611 crossrefType "journal-article" @default.
• W2912306611 hasAuthorship W2912306611A5000863572 @default.
• W2912306611 hasAuthorship W2912306611A5003760398 @default.
• W2912306611 hasAuthorship W2912306611A5005887396 @default.
• W2912306611 hasAuthorship W2912306611A5012395068 @default.
• W2912306611 hasAuthorship W2912306611A5019290469 @default.
• W2912306611 hasAuthorship W2912306611A5036046257 @default.
• W2912306611 hasAuthorship W2912306611A5036492073 @default.
• W2912306611 hasAuthorship W2912306611A5037878631 @default.
• W2912306611 hasAuthorship W2912306611A5043951146 @default.
• W2912306611 hasAuthorship W2912306611A5049344677 @default.
• W2912306611 hasAuthorship W2912306611A5063610143 @default.
• W2912306611 hasAuthorship W2912306611A5067215726 @default.
• W2912306611 hasAuthorship W2912306611A5072884120 @default.
• W2912306611 hasAuthorship W2912306611A5074200539 @default.
• W2912306611 hasAuthorship W2912306611A5076587915 @default.
• W2912306611 hasAuthorship W2912306611A5081093007 @default.
• W2912306611 hasAuthorship W2912306611A5085249009 @default.
• W2912306611 hasAuthorship W2912306611A5086359066 @default.
• W2912306611 hasAuthorship W2912306611A5087886690 @default.
• W2912306611 hasAuthorship W2912306611A5088321212 @default.
• W2912306611 hasBestOaLocation W29123066111 @default.
• W2912306611 hasConcept C104317684 @default.
• W2912306611 hasConcept C105702510 @default.
• W2912306611 hasConcept C126322002 @default.
• W2912306611 hasConcept C185592680 @default.
• W2912306611 hasConcept C2779030066 @default.
• W2912306611 hasConcept C2781111119 @default.
• W2912306611 hasConcept C501734568 @default.
• W2912306611 hasConcept C502942594 @default.
• W2912306611 hasConcept C54355233 @default.
• W2912306611 hasConcept C71924100 @default.
• W2912306611 hasConcept C86803240 @default.
• W2912306611 hasConcept C95444343 @default.
• W2912306611 hasConceptScore W2912306611C104317684 @default.
• W2912306611 hasConceptScore W2912306611C105702510 @default.
• W2912306611 hasConceptScore W2912306611C126322002 @default.
• W2912306611 hasConceptScore W2912306611C185592680 @default.
• W2912306611 hasConceptScore W2912306611C2779030066 @default.
• W2912306611 hasConceptScore W2912306611C2781111119 @default.
• W2912306611 hasConceptScore W2912306611C501734568 @default.

• next