The tetrabasic KKKK147-150 motif determines intracrine regulatory effects of PTHrP 1-173 on chondrocyte PPi metabolism and matrix synthesis

Expression of PTHrP is a major regulator of growth cartilage development an d also becomes robust in osteoarthritic cartilage. We further defined how P THrP 1-173, which we observed to be the preferentially expressed PTHrP isof orm in normal and osteoarthritic cartilage, functions in chondrocyte. We tr ansfected both immortalized human juvenile costal chondrocytes (TC28 cells) and rabbit articular chondrocytes with wild-type PTHrP 1-173 and mutants o f putative PTHrP 1-173 endoproteolytic processing sites. Wild-type PTHrP 1- 173 inhibited collagen synthesis and decreased extracellular PH (which crit ically regulates hydroxyapatite deposition) by 50-80% in both chondrocytic cell types. In contrast, PTHrP 1-173 mutated at the PTHrP 147-150 motif KKK K (but not the other site-directed mutants) and increased both extracellula r PPi and collagen synthesis by >50%. Synthetic PTHrP 140-173 mutated at am ino acids 147-150 and also increased extracellular PR, and wild-type 140-17 3 decreased extracellular PPi in permeabilized cells. The 147-150 KKKK doma in of PTHrP 1-173 acted, in part, by regulating nuclear localization of PTH rP. We conclude that the tetrabasic 147-150 motif functions to determine ho w PTHrP 1-173 regulates collagen synthesis and levels of extracellular PPi by an intracrine mechanism in chondrocytes, and it may prove useful as a th erapeutic target for regulation of mineralization.