We have performed microarray analysis to identify PTHrP target genes in chondrocytes. ATDC5 cells were cultured as micromasses to induce chondrocyte differentiation. At day 8 of culture the cells had a pre-hypertrophic appearance. This time point was chosen for isolation of RNA at 0 h, 1 h, 2 h, and 4 h after a challenge with 10^-7 M PTHrP. Samples were subjected to a cDNA microarray using competition hybridization.

A list of 12 genes (P < 10^-3), of which the expression regulation by PTHrP was confirmed by qPCR analysis, was generated. This included 7 upregulated and 5 downregulated genes. Three genes were known to be involved in PTHrP regulation and 6 were previously found in growth plate chondrocytes. Most of the genes (10 out of 12) were implicated in signal transduction and regulation. PTHrP also induced the expression of the upregulated genes in KS483 osteoblasts, suggesting involvement in a more generalized response to PTHrP. The vast majority of the upregulated genes (6 out of 7) contained CREB and/or AP-1 transcription factor binding sites in their promoter regions. Remarkably, a number of PTHrP regulated genes contained Stat transcription factor binding sites in their promoters. In transient transfection assays we show that PTHrP is able to positively regulate the activity of Stat3- and negatively regulate the activity of Stat5-specific promoter-reporter constructs in ATDC5 and UMR106 cells. In combination with the expression regulation of genes involved in Jak/Stat signaling, this data are suggestive for a previously not recognized interaction between PTHrP and Jak/Stat signaling.