TY - JOUR
T1 - UDP xylose synthase 1 is required for morphogenesis and histogenesis of the craniofacial skeleton
AU - Eames, B. Frank
AU - Singer, Amy
AU - Smith, Gabriel A.
AU - Wood, Zachary A.
AU - Yan, Yi Lin
AU - He, Xinjun
AU - Polizzi, Samuel J.
AU - Catchen, Julian M.
AU - Rodriguez-Mari, Adriana
AU - Linbo, Tor
AU - Raible, David W.
AU - Postlethwait, John H.
N1 - Thanks to Ruth Bremiller and Amanda Rapp for assistance with histology and animal care, and to Jeremy Wegner and Yasuko Honjo for erm, gli1, gli2, and gli3 probes. G.A.S. submitted a portion of this work as an undergraduate thesis to the Clark Honors College, University of Oregon, and offers special thanks to Nathan Tublitz and Joseph Fracchia. B.F.E. would like to thank Charles Kimmel for his support. This work was supported by grant numbers 5 F32 DE016778-03 (B.F.E.), R01 DE13834 (C.B.K.), and 5R01RR020833 and P01 HD22486 (J.H.P.) from the National Institutes of Health. The contents of this study are solely the responsibility of the authors and do not necessarily represent the official views of NCRR or NIH.
PY - 2010/5
Y1 - 2010/5
N2 - UDP-xylose synthase (Uxs1) is strongly conserved from bacteria to humans, but because no mutation has been studied in any animal, we do not understand its roles in development. Furthermore, no crystal structure has been published. Uxs1 synthesizes UDP-xylose, which initiates glycosaminoglycan attachment to a protein core during proteoglycan formation. Crystal structure and biochemical analyses revealed that an R233H substitution mutation in zebrafish uxs1 alters an arginine buried in the dimer interface, thereby destabilizing and, as enzyme assays show, inactivating the enzyme. Homozygous uxs1 mutants lack Alcian blue-positive, proteoglycan-rich extracellular matrix in cartilages of the neurocranium, pharyngeal arches, and pectoral girdle. Transcripts for uxs1 localize to skeletal domains at hatching. GFP-labeled neural crest cells revealed defective organization and morphogenesis of chondrocytes, perichondrium, and bone in uxs1 mutants. Proteoglycans were dramatically reduced and defectively localized in uxs1 mutants. Although col2a1a transcripts over-accumulated in uxs1 mutants, diminished quantities of Col2a1 protein suggested a role for proteoglycans in collagen secretion or localization. Expression of col10a1, indian hedgehog, and patched was disrupted in mutants, reflecting improper chondrocyte/perichondrium signaling. Up-regulation of sox9a, sox9b, and runx2b in mutants suggested a molecular mechanism consistent with a role for proteoglycans in regulating skeletal cell fate. Together, our data reveal time-dependent changes to gene expression in uxs1 mutants that support a signaling role for proteoglycans during at least two distinct phases of skeletal development. These investigations are the first to examine the effect of mutation on the structure and function of Uxs1 protein in any vertebrate embryos, and reveal that Uxs1 activity is essential for the production and organization of skeletal extracellular matrix, with consequent effects on cartilage, perichondral, and bone morphogenesis.
AB - UDP-xylose synthase (Uxs1) is strongly conserved from bacteria to humans, but because no mutation has been studied in any animal, we do not understand its roles in development. Furthermore, no crystal structure has been published. Uxs1 synthesizes UDP-xylose, which initiates glycosaminoglycan attachment to a protein core during proteoglycan formation. Crystal structure and biochemical analyses revealed that an R233H substitution mutation in zebrafish uxs1 alters an arginine buried in the dimer interface, thereby destabilizing and, as enzyme assays show, inactivating the enzyme. Homozygous uxs1 mutants lack Alcian blue-positive, proteoglycan-rich extracellular matrix in cartilages of the neurocranium, pharyngeal arches, and pectoral girdle. Transcripts for uxs1 localize to skeletal domains at hatching. GFP-labeled neural crest cells revealed defective organization and morphogenesis of chondrocytes, perichondrium, and bone in uxs1 mutants. Proteoglycans were dramatically reduced and defectively localized in uxs1 mutants. Although col2a1a transcripts over-accumulated in uxs1 mutants, diminished quantities of Col2a1 protein suggested a role for proteoglycans in collagen secretion or localization. Expression of col10a1, indian hedgehog, and patched was disrupted in mutants, reflecting improper chondrocyte/perichondrium signaling. Up-regulation of sox9a, sox9b, and runx2b in mutants suggested a molecular mechanism consistent with a role for proteoglycans in regulating skeletal cell fate. Together, our data reveal time-dependent changes to gene expression in uxs1 mutants that support a signaling role for proteoglycans during at least two distinct phases of skeletal development. These investigations are the first to examine the effect of mutation on the structure and function of Uxs1 protein in any vertebrate embryos, and reveal that Uxs1 activity is essential for the production and organization of skeletal extracellular matrix, with consequent effects on cartilage, perichondral, and bone morphogenesis.
KW - Cartilage
KW - Chondroitin sulfate
KW - Extracellular matrix
KW - FGF signaling
KW - Glycosaminoglycans
KW - Hedgehog signaling
KW - Heparan sulfate
KW - Neural crest
KW - Perichondrium
KW - Proteoglycans
KW - Skeletogenesis
KW - UDP-xylose synthase
KW - Zebrafish
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U2 - 10.1016/j.ydbio.2010.02.035
DO - 10.1016/j.ydbio.2010.02.035
M3 - Article
C2 - 20226781
AN - SCOPUS:77952298562
SN - 0012-1606
VL - 341
SP - 400
EP - 415
JO - Developmental Biology
JF - Developmental Biology
IS - 2
ER -