Parathyroid hormone (PTH) is an essential regulator of calcium homeostasis and became the prototypic osteoanabolic hormone for treating osteoporosis. On the other hand, hyperparathyroidism, with its consequent bone breakdown, is a major problem in and of itself and in chronic renal failure. The hormone acts through its G-protein-coupled receptor on the osteoblast to elicit enhanced bone resorption by the osteoclast. To do this, the osteoblast produces receptor activator of nuclear factor kappa-β ligand (RANKL) in response to PTH. RANKL is widely considered a principal mediator of PTH-induced bone catabolism and has been associated with bone loss in hyperparathyroidism. Our work in the last cycle of this grant revealed that PTH activates cAMP/protein kinase A (PKA) to inhibit salt-inducible kinases (SIKs) and requires protein phosphatases (PPs) to stimulate Rankl expression in osteoblasts. Knockdown of salt-inducible kinases (SIKs) 2 and 3 and cAMP- regulated transcription coactivators 2 and 3 (CRTC2 and 3) indicate that all four are part of this pathway. Moreover, inhibition of serine-threonine protein phosphatases decreased both PTH-induced Rankl expression and the stimulation by PTH(1-34) of CRTC2 and 3 translocation into the nucleus. Upon entry into the nucleus, these CRTCs associate with unknown basic leucine zipper domain (bZip) transcription factor(s), activating Rankl transcription through binding to its distal PTH-responsive cAMP-response elements (CREs). From these data of cells in culture and preliminary data in vivo, we have developed the central hypothesis that SIK/PP regulation of CRTC2/3 is essential to their function in the skeleton, and PTH controls this regulation. The long- term goals of this work are to delineate the signaling and transcriptional regulatory mechanisms conveying PTH action in bone. Consequently, the specific aims to test our hypothesis of this resubmitted competing renewal proposal focus on the SIK and PP regulation of CRTC2/3 function and action on Rankl, and will, 1) determine the mechanism of PTH regulation of Rankl transcription in osteoblasts by a. elucidating the regulatory mechanisms involved in CRTC2/3 nuclear translocation in osteoblasts, b. identifying the bZip transcription factor(s) responsible for CRTC2/3-induced Rankl transcription, 2) determine the role of CRTC2/3 in bone development and PTH’s anabolic and catabolic effects by a. determining the site and role of osteoblast CRTC2/3 in bone development, b. ascertaining the effects of deletion of CRTC2/3 on PTH’s anabolic and catabolic actions in bones of adult mice. The results of this work will make major contributions to our knowledge of how PTH exerts its nuclear effects on skeletal function through the SIK/PP/CRTC/bZip pathway. In so doing, the data will also provide new perspectives into treatment of disorders of calcium metabolism and other severe diseases functioning through the PTHR1 pathway such as Jansen’s metaphyseal chondrodysplasia and ...