Alternative Splicing of PRG4 in the Temporomandibular Joints of Baboons

Temporomandibular joint disorders (TMJD) affect a significant percentage of the population yet the molecular events underlying the progression of TMJDs are poorly understood. It has been proposed that a defect in the lubricating system of the TMJ is the precipitating event leading to the development of degenerative joint disease. Proteoglycan 4 (PRG4) is a secreted, mucin-like glycoprotein believed to provide boundary lubrication to articulating joints. At least four splice variants of PRG4 have been identified in other tissues however no studies to date have examined the presence of PRG4 splice variants in the TMJ. Objectives: The purpose of the proposed research is to better understand the molecular mechanisms of the lubrication system of the joint in an effort to identify causative factors responsible for common TMJ disorders. Methods: Total RNA was isolated from baboon TMJ cartilage, articular disc and synovial tissues and cDNAs synthesized. Quantitative real-time PCR (qRT-PCR) was performed to determine the expression level of PRG4 in baboon TMJ cells. Standard PCR was performed using primers designed to amplify the spliced regions and baboon cartilage, disc, and synovial cDNA templates to establish the presence of alternatively spliced PRG4 transcripts. Fluorescent dye-terminator sequencing was carried out to confirm the identity of each amplicon. Results: qRT-PCR analysis verified the presence of PRG4 transcripts in tissues of the TMJ. Standard PCR results indicated differential alternative splicing of PRG4 among the various cDNAs tested with at least three variants detected. DNA sequencing confirmed the identities of three alternatively spliced forms of PRG4. Conclusion: These studies show for the first time the presence of PRG4 splice variants in the TMJ of baboons. We anticipate that better understanding of the functions of this molecule will provide important insights into early molecular mechanisms that may be involved in common degenerative TMJ diseases.