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ADAMTSL-4 improves microfibril of Marfan syndrome derived cells
Abstracts
IHC decreased with age. ADAMTS aggrecanases are known to degrade aggrecan, one of the major extracellular matrix components in cartilage. ADAMTS-9 is reportedly highly expressed throughout mouse development including skeletogenesis. The results of the current study suggested that ADAMTS-9 might have a role in the matrix degradation during the course of chondrocyte proliferation and hypertrophic differentiation in the growth plate.
S25
present. However, this network was less efficient at protein trafficking, indicating that the phenotype is likely caused by defects in protein secretion. The human disorder Achondrogenesis type 1A (ACG1A, Houston–Harris type) is characterized by neonatal lethality due to thoracic insufficiency, shortlimbed dwarfism and delayed vertebral body ossification. ACG1A chondrocytes also display ER swelling. We screened TRIP11 in ACG1A patients and identified nonsense, frameshift, and splice-site mutations in the majority of probands.
doi:10.1016/j.matbio.2008.09.279 doi:10.1016/j.matbio.2008.09.281
65 ADAMTSL-4 improves microfibril of Marfan syndrome derived cells
67 Abnormal differentiation of cardiac valves in Ltbp-1L null mice
Masahiro Saitoa, Tomoko Wadaa, Ko Tsutsuib, Naoto Sudac, Ganburged Ganjargalc, Kiyotoshi Sekiguchid, Toshiyuki Yonedaa a Department Mol Cell Biochem, Osaka Univ Grad Sch Dent, Osaka, Japan b Inst for Protein Res, Osaka Univ, Osaka, Japan c Maxillo. Ortho, Tokyo Medi and Dent Univ, Tokyo, Japan
Vesna Todorovic, Erin Finnegan, Daniel Rifkin Cell Biology Department, NYU Medical Center, New York, NY 10016L, United States
Marfan syndrome (MFS) is a systemic disorder affecting connective tissues that is caused by mutations of the FBN1 gene encoding fibrillin-1, a major microfibril component. Prior observation suggested that MFS is associated with increasing susceptibility to severe periodontitis which associated with irreversible damage of periodontal ligament (PDL). However, the molecular mechanisms of microfibrils assembly in PDL formation remain largely unknown. Here, we report that ADAMTSL-4β, a novel microfibril binding protein, not only promotes fibrillin-1 microfibril assembly in PDL but also improves microfibril disorganization in cultured PDL cells obtained from MFS patient (M-HPDL). Expression patterning analysis revealed that adamtsl4β mRNA is strongly expressed in the dental follicle, the origin of the PDL, and ADAMTSL-4β protein is colocalized with the fibrillin-1 microfibril in the course of microfibril maturation during PDL development. In contrast, mice homozygous for a targeted hypomorphic allele (mgR/mgR) of Fbn1, which served as a mice model of MFS, showed disorganization of PDL in association with progressive fragmentation of ADAMTSL-4β microfibrils. M-HPDL able to form insufficient fibrillin-1 microfibril, nevertheless overexpression of ADAMTSL-4β in M-HPDL markedly improved fibrillin-1 microfibril assembly. Our results suggest that ADAMTSL-4β regulates microfibril assembly of fibrillin-1 during PDL development, and could be a novel therapeutic target for the damaged PDL tissue in patients with MFS.
TBP-1 belongs to the LTBP/fibrillin family of extracellular proteins. Due to usage of different promoters, LTBP-1 exists in two major forms: long (L) and short (S), each expressed in a temporally and spatially unique fashion. Both LTBP-1 molecules covalently interact with latent TGF-β and regulate its function. Lack of Ltbp-1L results in perinatal death due to abnormal heart development, with improper septation of the cardiac outflow tract and remodeling of the associated vessels. These cardiac anomalies present as persistent truncus arteriosus and interrupted aortic arch, which are associated with faulty cardiac neural crest function (Todorovic et al., Development 2007,134:3723–32). However, 40% of Ltbp-1L-null embryos die during fetal development due to abnormal valve differentiation. Cardiac valves derive from endocardial cushion tissue and Ltbp-1L is expressed in the myocardium underlying the regions where endocardial–mesenchymal transformation (EMT), which produces the cushions, takes place. During endocardial cushion differentiation and development, Ltbp-1L expression widens to the whole cushion. While ex vitro results suggested a crucial role for Ltbp-1 in cardiac EMT, our results show that lack of Ltbp-1L does not affect cardiac EMT but rather endocardial cushion fusion and remodeling. The malformed valve leaflets are misshaped, hyperplastic and thick, causing improper heart function and heart failure. Absence of Ltbp-1L during valve development causes decreased TGF-β activity in the remodeling mesenchyme, revealing a critical role for Ltbp-1L as an extracellular regulator of TGF-β activity during valve formation.
doi:10.1016/j.matbio.2008.09.280 doi:10.1016/j.matbio.2008.09.282
66 TRIP11 is essential for skeletal development Patrick Smitsa, Andrew Boltonb, Minh Honga, Lei Luc, Andrea Superti-Furgaf, Shiro Ikegawae, Daniel Cohnd, Tom Kirchhausenc, Matthew Warmana, Dave Beierb a Department Orthopaedic Surgery, Children's Hospital, Boston, United States b Department Genetics, Brigham and Women's Hospital, Boston, United States c Department Cell Biology, Harvard Medical Institute, Boston, United States d Cedars-Sinai Medical Center, Los Angeles, United States e RIKEN, Tokyo, Japan f Department Pediatrics, University of Freiburg, Germany Trip11 (Thyroid hormone interacting protein 11) null mice display an autosomal recessive, perinatal lethal, skeletal dysplasia characterized by short limbs, small thoraces and a failure to ossify vertebral bodies and skull bones, indicating impaired endochondral and intra-membranous ossification. Growth plates showed abnormalities in chondrocyte proliferation and maturation; mutant chondrocytes also had increased apoptosis rates. A reduction in Cbfa1 expression in the presumptive skull bones was consistent with impaired intra-membranous bone formation. All Trip11 null cells displayed an absence of Golgi stacks, however only chondrocytes and osteoblasts contained swollen endoplasmic reticulum (ER). Although mutant fibroblasts lack Golgi stacks, an oriented cis- and trans-Golgi network was
68 ADAMTS proteases regulate BMP-mediated cell death Daniel R. McCullocha, Laura Collinsa, Courtney M. Nelsona, Takako Sasakib, Marion A. Cooleyc, W. Scott Argravesc, Suneel S. Aptea a Department of Biomedical Engineering, Cleveland Clinic, Cleveland, OH USA b Shriners Hospital for Children, Portland, OR, USA c Medical University of South Carolina, Charleston, SC, USA BMP-mediated apoptosis of interdigital mesenchyme regulates web regression in mammals. Failure of resorption leads to interdigital webbing, termed soft-tissue syndactyly (STS). We show that combinatorial ADAMTS protease deficient mice (Adamts5 + Adamts20 (bt) and Adamts5 + Adamts9) have STS with greater penetrance and severity than deletion of these genes singly. Apoptosis is reduced in the affected webs. Bmp2 and Bmp4 mRNA expression is unaffected in mutant limbs, but application of BMP-4 beads rescues apoptosis, suggesting that these ADAMTS proteases act upstream of BMP signaling. Adamts5, Adamts9 and Adamts20 expression overlaps with the distribution of the proteoglycan versican, which they all cleave, and with fibulin-1, a co-factor for ADAMTS1 proteolysis of aggrecan. Versican cleavage is markedly reduced in the interdigital mesenchyme of Adamts5−/−, Adamts20−/− mice. Versican cleavage by ADAMTS5 is enhanced four-fold by fibulin-1, and Adamts20−/− mice haploinsufficient for either versican or
IHC decreased with age. ADAMTS aggrecanases are known to degrade aggrecan, one of the major extracellular matrix components in cartilage. ADAMTS-9 is reportedly highly expressed throughout mouse development including skeletogenesis. The results of the current study suggested that ADAMTS-9 might have a role in the matrix degradation during the course of chondrocyte proliferation and hypertrophic differentiation in the growth plate.
S25
present. However, this network was less efficient at protein trafficking, indicating that the phenotype is likely caused by defects in protein secretion. The human disorder Achondrogenesis type 1A (ACG1A, Houston–Harris type) is characterized by neonatal lethality due to thoracic insufficiency, shortlimbed dwarfism and delayed vertebral body ossification. ACG1A chondrocytes also display ER swelling. We screened TRIP11 in ACG1A patients and identified nonsense, frameshift, and splice-site mutations in the majority of probands.
doi:10.1016/j.matbio.2008.09.279 doi:10.1016/j.matbio.2008.09.281
65 ADAMTSL-4 improves microfibril of Marfan syndrome derived cells
67 Abnormal differentiation of cardiac valves in Ltbp-1L null mice
Masahiro Saitoa, Tomoko Wadaa, Ko Tsutsuib, Naoto Sudac, Ganburged Ganjargalc, Kiyotoshi Sekiguchid, Toshiyuki Yonedaa a Department Mol Cell Biochem, Osaka Univ Grad Sch Dent, Osaka, Japan b Inst for Protein Res, Osaka Univ, Osaka, Japan c Maxillo. Ortho, Tokyo Medi and Dent Univ, Tokyo, Japan
Vesna Todorovic, Erin Finnegan, Daniel Rifkin Cell Biology Department, NYU Medical Center, New York, NY 10016L, United States
Marfan syndrome (MFS) is a systemic disorder affecting connective tissues that is caused by mutations of the FBN1 gene encoding fibrillin-1, a major microfibril component. Prior observation suggested that MFS is associated with increasing susceptibility to severe periodontitis which associated with irreversible damage of periodontal ligament (PDL). However, the molecular mechanisms of microfibrils assembly in PDL formation remain largely unknown. Here, we report that ADAMTSL-4β, a novel microfibril binding protein, not only promotes fibrillin-1 microfibril assembly in PDL but also improves microfibril disorganization in cultured PDL cells obtained from MFS patient (M-HPDL). Expression patterning analysis revealed that adamtsl4β mRNA is strongly expressed in the dental follicle, the origin of the PDL, and ADAMTSL-4β protein is colocalized with the fibrillin-1 microfibril in the course of microfibril maturation during PDL development. In contrast, mice homozygous for a targeted hypomorphic allele (mgR/mgR) of Fbn1, which served as a mice model of MFS, showed disorganization of PDL in association with progressive fragmentation of ADAMTSL-4β microfibrils. M-HPDL able to form insufficient fibrillin-1 microfibril, nevertheless overexpression of ADAMTSL-4β in M-HPDL markedly improved fibrillin-1 microfibril assembly. Our results suggest that ADAMTSL-4β regulates microfibril assembly of fibrillin-1 during PDL development, and could be a novel therapeutic target for the damaged PDL tissue in patients with MFS.
TBP-1 belongs to the LTBP/fibrillin family of extracellular proteins. Due to usage of different promoters, LTBP-1 exists in two major forms: long (L) and short (S), each expressed in a temporally and spatially unique fashion. Both LTBP-1 molecules covalently interact with latent TGF-β and regulate its function. Lack of Ltbp-1L results in perinatal death due to abnormal heart development, with improper septation of the cardiac outflow tract and remodeling of the associated vessels. These cardiac anomalies present as persistent truncus arteriosus and interrupted aortic arch, which are associated with faulty cardiac neural crest function (Todorovic et al., Development 2007,134:3723–32). However, 40% of Ltbp-1L-null embryos die during fetal development due to abnormal valve differentiation. Cardiac valves derive from endocardial cushion tissue and Ltbp-1L is expressed in the myocardium underlying the regions where endocardial–mesenchymal transformation (EMT), which produces the cushions, takes place. During endocardial cushion differentiation and development, Ltbp-1L expression widens to the whole cushion. While ex vitro results suggested a crucial role for Ltbp-1 in cardiac EMT, our results show that lack of Ltbp-1L does not affect cardiac EMT but rather endocardial cushion fusion and remodeling. The malformed valve leaflets are misshaped, hyperplastic and thick, causing improper heart function and heart failure. Absence of Ltbp-1L during valve development causes decreased TGF-β activity in the remodeling mesenchyme, revealing a critical role for Ltbp-1L as an extracellular regulator of TGF-β activity during valve formation.
doi:10.1016/j.matbio.2008.09.280 doi:10.1016/j.matbio.2008.09.282
66 TRIP11 is essential for skeletal development Patrick Smitsa, Andrew Boltonb, Minh Honga, Lei Luc, Andrea Superti-Furgaf, Shiro Ikegawae, Daniel Cohnd, Tom Kirchhausenc, Matthew Warmana, Dave Beierb a Department Orthopaedic Surgery, Children's Hospital, Boston, United States b Department Genetics, Brigham and Women's Hospital, Boston, United States c Department Cell Biology, Harvard Medical Institute, Boston, United States d Cedars-Sinai Medical Center, Los Angeles, United States e RIKEN, Tokyo, Japan f Department Pediatrics, University of Freiburg, Germany Trip11 (Thyroid hormone interacting protein 11) null mice display an autosomal recessive, perinatal lethal, skeletal dysplasia characterized by short limbs, small thoraces and a failure to ossify vertebral bodies and skull bones, indicating impaired endochondral and intra-membranous ossification. Growth plates showed abnormalities in chondrocyte proliferation and maturation; mutant chondrocytes also had increased apoptosis rates. A reduction in Cbfa1 expression in the presumptive skull bones was consistent with impaired intra-membranous bone formation. All Trip11 null cells displayed an absence of Golgi stacks, however only chondrocytes and osteoblasts contained swollen endoplasmic reticulum (ER). Although mutant fibroblasts lack Golgi stacks, an oriented cis- and trans-Golgi network was
68 ADAMTS proteases regulate BMP-mediated cell death Daniel R. McCullocha, Laura Collinsa, Courtney M. Nelsona, Takako Sasakib, Marion A. Cooleyc, W. Scott Argravesc, Suneel S. Aptea a Department of Biomedical Engineering, Cleveland Clinic, Cleveland, OH USA b Shriners Hospital for Children, Portland, OR, USA c Medical University of South Carolina, Charleston, SC, USA BMP-mediated apoptosis of interdigital mesenchyme regulates web regression in mammals. Failure of resorption leads to interdigital webbing, termed soft-tissue syndactyly (STS). We show that combinatorial ADAMTS protease deficient mice (Adamts5 + Adamts20 (bt) and Adamts5 + Adamts9) have STS with greater penetrance and severity than deletion of these genes singly. Apoptosis is reduced in the affected webs. Bmp2 and Bmp4 mRNA expression is unaffected in mutant limbs, but application of BMP-4 beads rescues apoptosis, suggesting that these ADAMTS proteases act upstream of BMP signaling. Adamts5, Adamts9 and Adamts20 expression overlaps with the distribution of the proteoglycan versican, which they all cleave, and with fibulin-1, a co-factor for ADAMTS1 proteolysis of aggrecan. Versican cleavage is markedly reduced in the interdigital mesenchyme of Adamts5−/−, Adamts20−/− mice. Versican cleavage by ADAMTS5 is enhanced four-fold by fibulin-1, and Adamts20−/− mice haploinsufficient for either versican or