Xylosyltransferase 1 is a key regulator of early chondrocyte maturation and mature bone formation.

摘要

The long bones of the vertebrate body are built by the initial formation of a cartilage template that is later replaced by mineralized bone. The proliferation and maturation of the skeletal precursor cells (chondrocytes) within the cartilage template and their replacement by bone is a highly coordinated process which, if misregulated, can lead to a number of defects including dwarfism and other skeletal deformities. This is exemplified by the fact that abnormal skeletal structure and development is one of the most common types of human birth defects. Skeletal development requires the interaction between different chondrocyte maturation states within the growth plate and the extracellular matrix, including transcription factors, signaling molecules, and extracellular matrix proteins. Though much is known about these regulatory mechanisms, many of the factors that initiate and regulate chondrocyte maturation have not been described. We identified a recessive dwarf mouse mutant (pug ) from an N-ethyl-N-nitrosourea (ENU) mutagenesis screen. pug mutant skeletal elements are patterned normally during development, but display a ∼20% length reduction compared to wild-type embryos. We show that the pug mutation does not lead to changes in chondrocyte proliferation but instead promotes premature maturation and early ossification, which ultimately leads to disproportionate dwarfism in adulthood. This skeletal dysplasia phenotype appears to be the only defect in both the embryo and the adult, revealing a specificity for the pug mutation in regulating skeletal formation and maintenance. Using sequence capture and high-throughput sequencing, we identified a missense mutation in the Xylosyltransferase 1 (Xylt1) gene in pug mutants. Consistent with the phenotype observed, Xylt1 is exclusively expressed within the developing skeletal elements during development, with no significant expression observed in other tissues during embryonic development. Xylosyltransferases catalyze the initial step in glycosaminoglycan (GAG) chain addition to proteoglycan core proteins, and these modifications are essential for normal proteoglycan function and normal extracellular matrix structure and function. We show that the pug mutation disrupts both Xylt1 enzymatic activity and subcellular localization, leading to a reduction in GAG chains in pug mutants. This reduction in GAG chains results in altered Indian Hedgehog (Ihh) signaling within the growth plate, but these signaling changes occur after the first morphological differences can be observed within pug skeletal elements. The lack of changes in the major signaling pathways prior to chondrocyte maturation points to a potential role for the extracellular matrix in regulating chondrocyte maturation initiation in the absence of signaling. As one of only two xylosyltransferases in vertebrates, further characterization of the pug mutant in conjunction with mutations in Xylt2 will uncover the requirements for processed proteoglycans in development and tissue homeostasis. Our analysis has revealed that the pug mutant serves as a novel model for mammalian dwarfism and identifies a key role for proteoglycan modifications in the initiation of chondrocyte maturation.