A Zeb1/MtCK1 metabolic axis controls osteoclast activation and skeletal remodeling.

Osteoclasts are bone-resorbing polykaryons responsible for skeletal remodeling during health and disease. Coincident with their differentiation from myeloid precursors, osteoclasts undergo extensive transcriptional and metabolic reprogramming in order to acquire the cellular machinery necessary to demineralize bone and digest its interwoven extracellular matrix. While attempting to identify new regulatory molecules critical to bone resorption, we discovered that murine and human osteoclast differentiation is accompanied by the expression of Zeb1, a zinc-finger transcriptional repressor whose role in normal development is most frequently linked to the control of epithelial-mesenchymal programs. However, following targeting, we find that Zeb1 serves as an unexpected regulator of osteoclast energy metabolism. In vivo, Zeb1-null osteoclasts assume a hyperactivated state, markedly decreasing bone density due to excessive resorptive activity. Mechanistically, Zeb1 acts in a rheostat-like fashion to modulate murine and human osteoclast activity by transcriptionally repressing an ATP-buffering enzyme, mitochondrial creatine kinase 1 (MtCK1), thereby controlling the phosphocreatine energy shuttle and mitochondrial respiration. Together, these studies identify a novel Zeb1/MtCK1 axis that exerts metabolic control over bone resorption in vitro and in vivo.

Wound repair in sea urchin larvae involves pigment cells and blastocoelar cells.

Sea urchin larvae spend weeks to months feeding on plankton prior to metamorphosis. When handled in the laboratory they are easily injured, suggesting that in the plankton they are injured with some frequency. Fortunately, larval wounds are repaired through an efficient wound response with mesenchymal pigment cells and blastocoelar cells assisting as the epithelium closes. An injury to the epithelium leads to an immediate calcium transient that rapidly spreads around the entire larva and is necessary for activating pigment cell migration toward the wound. If calcium transport is blocked, the pigment cells fail to activate and remain in place. When activated, pigment cells initiate directed migration to the wound site from distances of at least 85 â€‹µm. Upon arrival at the wound site they participate in an innate immune response. Blastocoelar cells are recruited to the injury site as well, though the calcium transient is unnecessary for activating these cells. At the wound site, blastocoelar cells participate in several functions including remodeling the skeleton if it protrudes through the epithelium.

nkx3.2 mutant zebrafish accommodate jaw joint loss through a phenocopy of the head shapes of Paleozoic jawless fish.

The vertebrate jaw is a versatile feeding apparatus. To function, it requires a joint between the upper and lower jaws, so jaw joint defects are often highly disruptive and difficult to study. To describe the consequences of jaw joint dysfunction, we engineered two independent null alleles of a single jaw joint marker gene, nkx3.2, in zebrafish. These mutations caused zebrafish to become functionally jawless via fusion of the upper and lower jaw cartilages (ankylosis). Despite lacking jaw joints, nkx3.2 mutants survived to adulthood and accommodated this defect by: (a) having a remodeled skull with a fixed open gape, reduced snout and enlarged branchial region; and (b) performing ram feeding in the absence of jaw-generated suction. The late onset and broad extent of phenotypic changes in the mutants suggest that modifications to the skull are induced by functional agnathia, secondarily to nkx3.2 loss of function. Interestingly, nkx3.2 mutants superficially resemble ancient jawless vertebrates (anaspids and furcacaudiid thelodonts) in overall head shape. Because no homology exists in individual skull elements between these taxa, the adult nkx3.2 phenotype is not a reversal but rather a convergence due to similar functional requirements of feeding without moveable jaws. This remarkable analogy strongly suggests that jaw movements themselves dramatically influence the development of jawed vertebrate skulls. Thus, these mutants provide a unique model with which to: (a) investigate adaptive responses to perturbation in skeletal development; (b) re-evaluate evolutionarily inspired interpretations of phenocopies generated by gene knockdowns and knockouts; and (c) gain insight into feeding mechanics of the extinct agnathans.

Smoke carcinogens cause bone loss through the aryl hydrocarbon receptor and induction of Cyp1 enzymes.

Smoking is a major risk factor for osteoporosis and fracture, but the mechanism through which smoke causes bone loss remains unclear. Here, we show that the smoke toxins benzo(a)pyrene (BaP) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) interact with the aryl hydrocarbon receptor (Ahr) to induce osteoclastic bone resorption through the activation of cytochrome P450 1a/1b (Cyp1) enzymes. BaP and TCDD enhanced osteoclast formation in bone marrow cell cultures and gavage with BaP stimulated bone resorption and osteoclastogenesis in vivo. The osteoclastogenesis triggered by BaP or RANK-L was reduced in Ahr(-/-) cells, consistent with the high bone mass noted in Ahr(-/-) male mice. The receptor activator of NF-κB ligand (RANK-L) also failed to induce the expression of Cyp1 enzymes in Ahr(-/-) cells. Furthermore, the osteoclastogenesis induced by TCDD was lower in Cyp1a1/1a2(-/-) and Cyp1a1/1a2/1b1(-/-) cultures, indicating that Ahr was upstream of the Cyp enzymes. Likewise, the pharmacological inhibition of the Cyp1 enzymes with tetramethylsilane or proadifen reduced osteoclastogenesis. Finally, deletion of the Cyp1a1, Cyp1a2, and Cyp1b1 in triple knockout mice resulted in reduced bone resorption and recapitulated the high bone mass phenotype of Ahr(-/-) mice. Overall, the data identify the Ahr and Cyp1 enzymes not only in the pathophysiology of smoke-induced osteoporosis, but also as potential targets for selective modulation by new therapeutics.

miRNA-27a is essential for bone remodeling by modulating p62-mediated osteoclast signaling.

The ability to simultaneously modulate a set of genes for lineage-specific development has made miRNA an ideal master regulator for organogenesis. However, most miRNA deletions do not exhibit obvious phenotypic defects possibly due to functional redundancy. miRNAs are known to regulate skeletal lineages as the loss of their maturation enzyme Dicer impairs bone remodeling processes. Therefore, it is important to identify specific miRNA essential for bone homeostasis. We report the loss of MIR27a causing severe osteoporosis in mice. MIR27a affects osteoclast-mediated bone resorption but not osteoblast-mediated bone formation during skeletal remodeling. Gene profiling and bioinformatics further identify the specific targets of MIR27a in osteoclast cells. MIR27a exerts its effects on osteoclast differentiation through modulation of Squstm1/p62 whose mutations have been linked to Paget's disease of bone. Our findings reveal a new MIR27a-p62 axis necessary and sufficient to mediate osteoclast differentiation and highlight a therapeutic implication for osteoporosis.