Thus, RBP-J imposes the requirement for ITAM-mediated costimulation of RANKL. It also inhibits ITAM-mediated expression and function of the previously identified member of the key signaling complex, PLCɣ2, as well as activation of downstream calcium-CaMKK/PYK2 signaling. It does this by suppressing induction of key osteoclastogenic factors, such as NFATc1, BLIMP1 and c-FOS. RBP-J thereby limits the crosstalk between ITAM and RANK/TNFR and allows for fine-tuning of osteoblastogenesis during bone homeostasis. This is the process that appears to underlie bone health, as well as bone destruction seen in RA.
Experimental Details
In order to elucidate this pathway, the researchers used powerful, high-tech, next generation, whole transcriptome sequencing. They began their research with two mouse models of osteoporosis (Dap12-/- and Dap12-/-Fcrg-/-). Their experiments revealed that the osteoporotic bone phenotype in these mice is mediated by RBJ-1. Specifically, deletion of Rbpj in these mice substantially rescued the defects in bone remodeling, thereby reversing the osteoporotic bone phenotype. A more detailed analysis revealed that knocking out Rbpj bypassed the requirement for ITAM-mediated costimulation of RANKL-induced osteoblastogenesis.
The investigators then looked more closely at the role of PLCɣ2 in transgenic and control mice. Its relative expression and phosphorylation in the different strains suggested that it plays an important regulatory role in the pathway. In contrast, the previously identified ITAM-binding protein FcRɣ did not appear to significantly affect bone remodeling. Inhibition of PLCɣ2 activity significantly reduced the enhanced RANKL-induced osteoclastogenesis in RBP-J-deficient cells. A more detailed analysis also revealed that RBP-J regulates TGF-β signaling and, thus, suppresses the expression and activity of basal PLCɣ2 and downstream calcium-CaMKK-PYK2 signaling. Investigators additionally found that the RBP-J target gene Tgfbr1 (which codes for TGF-β receptor I) was expressed at a rate that was 2.5 times higher in RBP-J-deficient osteoclast precursors compared to controls.
The team then turned to a different mouse model of bone disease: a TNF-α-induced model of inflammatory bone resorption. They found that RBP-J deficiency allows TNF-α to induce osteoclast formation and bone resorption in DAP12-deficient mice. Moreover, RBP-J deficiency in the absence of ITAM-mediated costimulatory signaling enabled TNF-α to induce NFATc1 and BLIMP1 expression and osteoclastogenesis. Thus, RBP-J also imposes the requirement for ITAM-mediated costimulation in TNF-α-induced osteoclastogenesis.
Lara C. Pullen, PhD, is a medical writer based in the Chicago area.
Reference
- Li S, Miller CH, Giannopoulou E, Hu X, Ivashkiv LB, Zhao B. RBP-J imposes a requirement for ITAM-mediated costimulation of osteoclastogenesis. J Clin Invest. 2014 Nov 3;124(11):5057–5073.