Bone resorption by osteoclasts is essential for bone homeostasis. The kinase Src promotes osteoclast activity and is activated in osteoclasts by the receptor-type tyrosine phosphatase PTPROt. In other contexts, however, PTPROt can inhibit Src activity. Through in vivo and in vitro experiments, we show that PTPROt is bifunctional and can dephosphorylate Src both at its inhibitory residue Tyr⁵²⁷ and its activating residue Tyr⁴¹⁶. Whereas wild-type and PTPROt knockout mice exhibited similar bone masses, mice in which a putative C-terminal phosphorylation site, Tyr³⁹⁹, in endogenous PTPROt was replaced with phenylalanine had increased bone mass and reduced osteoclast activity. Osteoclasts from the knock-in mice also showed reduced Src activity. Experiments in cultured cells and in osteoclasts derived from both mouse strains demonstrated that the absence of phosphorylation at Tyr³⁹⁹ caused PTPROt to dephosphorylate Src at the activating site pTyr⁴¹⁶. In contrast, phosphorylation of PTPROt at Tyr³⁹⁹ enabled PTPROt to recruit Src through Grb2 and to dephosphorylate Src at the inhibitory site Tyr⁵²⁷, thus stimulating Src activity. We conclude that reversible phosphorylation of PTPROt at Tyr³⁹⁹ is a molecular switch that selects between its opposing activities toward Src and maintains a coherent signaling output, and that blocking this phosphorylation event can induce physiological effects in vivo. Because most receptor-type tyrosine phosphatases contain potential phosphorylation sites at their C termini, we propose that preventing phosphorylation at these sites or its consequences may offer an alternative to inhibiting their catalytic activity to achieve therapeutic benefit.

破骨细胞的骨吸收对于骨稳态是必不可少的。激酶Src促进破骨细胞活性，并在破骨细胞中被受体型酪氨酸磷酸酶PTPROt激活。但是，在其他情况下，PTPROt可以抑制Src活性。通过体内和体外实验，我们显示PTPROt是双功能的，并且可以在其抑制残基Tyr ⁵²⁷和其活化残基Tyr ^416上使Src脱磷酸化。野生型和PTPROt基因敲除小鼠显示相似的骨量，而推定的C末端磷酸化位点Tyr ^399的小鼠，在内源性PTPROt替换为苯丙氨酸会增加骨量并降低破骨细胞活性。来自敲入小鼠的破骨细胞也显示出降低的Src活性。在两种小鼠品系的培养细胞和破骨细胞中进行的实验表明，Tyr ³⁹⁹的磷酸化缺失导致PTPROt在激活位点pTyr ⁴¹⁶上使Src脱磷酸化。相反，PTPROt在Tyr ^399的磷酸化使PTPROt能够通过Grb2募集Src，并使Src在抑制位点Tyr ⁵²⁷上磷酸化，从而刺激Src活性。我们得出的结论是，Tyr ³⁹⁹处的PTPROt可逆磷酸化是一个分子开关，可以在其对Src的相反活性之间进行选择，并维持一致的信号输出，并且阻断这种磷酸化事件可以在体内诱导生理作用。因为大多数受体型酪氨酸磷酸酶在其C末端均含有潜在的磷酸化位点，所以我们建议防止这些位点的磷酸化或其后果可能是抑制其催化活性以获得治疗益处的另一种选择。