Novel Small Leucine-Rich Protein Chondroadherin-like (CHADL) is Expressed in Cartilage and Modulates Chondrocyte Differentiation

The constitution and biophysical properties of extracellular matrices can dramatically influence cellular phenotype during development, homeostasis, or pathogenesis. These effects can be signaled through a differentially regulated assembly of collagen fibrils, orchestrated by a family of collagenassociated Small Leucine-Rich Proteins, SLRPs. In this report, we describe the tissue-specific expression and function of a previously uncharacterized SLRP – Chondroadherin-like (CHADL). We have developed antibodies against CHADL and, by immunohistochemistry, detected CHADL expression mainly in skeletal tissues, particularly in fetal cartilage and in pericellular space of adult chondrocytes. In situ hybridizations and immunoblots on tissue lysates confirmed this tissue-specific expression pattern. Recombinant CHADL bound collagen in cell culture, and inhibited in vitro collagen fibrillogenesis. After Chadl shRNA knockdown chondrogenic ATDC5 cells increased their differentiation, indicated by increased transcript levels of Sox9, Ihh, Col2a1, and Col10a1. The knockdown increased collagen II and aggrecan deposition in the cell layers. Microarray analysis of the knockdown samples suggested collagen receptorrelated changes, although other upstream effects could not be excluded. Together, our data indicate that the novel SLRP CHADL is expressed in cartilaginous tissues, influences collagen fibrillogenesis, and modulates chondrocyte differentiation. CHADL appears to have a negative regulatory role, possibly ensuring the formation of a stable extracellular matrix. To gain a comprehensive understanding of connective tissue biology it is crucial to study proteins that regulate tissueand function-specific collagen fibril assembly. Over the past three decades, several homologous Small Leucine-Rich Proteins (SLRPs1) have been identified and functionally evaluated; many of these proteins are extracellularly associated with collagen and can influence collagen fibrillogenesis in vitro (1) – for example, decorin, biglycan, fibromodulin, and