Proline-rich tyrosine kinase 2 regulates osteoprogenitor cells and bone formation, and offers an anabolic treatment approach for osteoporosis

Bone is accrued and maintained primarily through the coupled actions of bone-forming osteoblasts and bone-resorbing osteoclasts. Cumulative in vitro studies indicated that proline-rich tyrosine kinase 2 (PYK2) is a positive mediator of osteoclast function and activity. However, our investigation of PYK2−/− mice did not reveal evidence supporting an essential function for PYK2 in osteoclasts either in vivo or in culture. We find that PYK2−/− mice have high bone mass resulting from an unexpected increase in bone formation. Consistent with the in vivo findings, mouse bone marrow cultures show that PYK2 deficiency enhances differentiation and activity of osteoprogenitor cells, as does expressing a PYK2-specific short hairpin RNA or dominantly interfering proteins in human mesenchymal stem cells. Furthermore, the daily administration of a small-molecule PYK2 inhibitor increases bone formation and protects against bone loss in ovariectomized rats, an established preclinical model of postmenopausal osteoporosis. In summary, we find that PYK2 regulates the differentiation of early osteoprogenitor cells across species and that inhibitors of the PYK2 have potential as a bone anabolic approach for the treatment of osteoporosis

Unusual characteristics of amino-terminal and hydrophobic domains in nuclear-encoded thylakoid signal peptides

Thylakoid transfer signals carry information specifying translocation by either a Sec- or ΔpH-dependent protein translocator in the chloroplast thylakoid membrane, yet all resemble classical signal peptides in overall structural terms. Comparison of known transfer signals reveals two differences: (a) signals for the ΔpH-driven system invariably contain a critical twin-argrinine (Arg-Arg) motif prior to the hydrophobic (H) domain, whereas known Sec-dependent signals contain lysine, and (b) the H-domains of Sec-dependent signals are generally longer. Previous work has shown that a twin-Arg motif before the H-domain is critical for targeting by the ΔpH-dependent pathway; in this report we show that the charge characteristics of this region are not important for sorting by the Sec pathway. Twin-Lys, twin-Am or single Arg are all acceptable to the Sec system, although single Lys/Arg is preferred. The single Lys in pre-plastocyanin can even be replaced by an uncharged residue without apparent effect. We have also generated a pre-plastocyanin mutant containing an H-domain which, in terms of hydropathy profile, is identical to that of a ΔpH-dependent protein. This mutant is also transported efficiently by the Sec system, demonstrating that hydrophobicity per se is not a key sorting determinant. However, the characteristics of the H-domain may be important in avoiding a different form of mis-targeting: to the endoplasmic reticulum. Thylakoid signal peptides have undergone substantial structural changes during the evolution of the chloroplast from endosymbiotic cyanobacterium: plastid-encoded and cyanobacterial signals contain H-domains that are highly hydrophobic and enriched in Leu and aromatic residues, whereas nuclear-encoded counterparts are Ala-rich and far less hydrophobic. We speculate that this trend may reflect a need to avoid mistargeting through recognition by cytosolic signal recognition particle, which preferentially interacts with more hydrophobic signal peptides.</p