Ablation of Y1 receptor impairs osteoclast bone-resorbing activity.

Y1 receptor (Y1R)-signalling pathway plays a pivotal role in the regulation of bone metabolism. The lack of Y1R-signalling stimulates bone mass accretion that has been mainly attributed to Y1R disruption from bone-forming cells. Still, the involvement of Y1R-signalling in the control of bone-resorbing cells remained to be explored. Therefore, in this study we assessed the role of Y1R deficiency in osteoclast formation and resorption activity. Here we demonstrate that Y1R germline deletion (Y1R(-/-)) led to increased formation of highly multinucleated (n > 8) osteoclasts and enhanced surface area, possibly due to monocyte chemoattractant protein-1 (MCP-1) overexpression regulated by RANKL-signalling. Interestingly, functional studies revealed that these giant Y1R(-/-) multinucleated cells produce poorly demineralized eroded pits, which were associated to reduce expression of osteoclast matrix degradation markers, such as tartrate-resistant acid phosphatase-5b (TRAcP5b), matrix metalloproteinase-9 (MMP-9) and cathepsin-K (CTSK). Tridimensional (3D) morphologic analyses of resorption pits, using an in-house developed quantitative computational tool (BonePit), showed that Y1R(-/-) resorption pits displayed a marked reduction in surface area, volume and depth. Together, these data demonstrates that the lack of Y1Rs stimulates the formation of larger multinucleated osteoclasts in vitro with reduced bone-resorbing activity, unveiling a novel therapeutic option for osteoclastic bone diseases based on Y1R-signalling ablation.

DNAJB4 molecular chaperone distinguishes WT from mutant E-cadherin, determining their fate in vitro and in vivo.

E-cadherin (Ecad) is a well-known invasion suppressor and its loss of expression is common in invasive carcinomas. Germline Ecad mutations are the only known genetic cause of hereditary diffuse gastric cancer (HDGC), demonstrating the causative role of Ecad impairment in gastric cancer. HDGC-associated Ecad missense mutations can lead to folding defects and premature proteasome-dependent endoplasmic reticulum-associated degradation (ERAD), but the molecular determinants for this fate were unidentified. Using a Drosophila-based genetic screen, we found that Drosophila DnaJ-1 interacts with wild type (WT) and mutant human Ecad in vivo. DnaJ (Hsp40) homolog, subfamily B, member 4 (DNAJB4), the human homolog of DnaJ-1, influences Ecad localization and stability even in the absence of Ecad endogenous promoter, suggesting a post-transcriptional level of regulation. Increased expression of DNAJB4 leads to stabilization of WT Ecad in the plasma membrane, while it induces premature degradation of unfolded HDGC mutants in the proteasome. The interaction between DNAJB4 and Ecad is direct, and is increased in the context of the unfolded mutant E757K, especially when proteasome degradation is inhibited, suggesting that DNAJB4 is a molecular mediator of ERAD. Post-translational regulation of native Ecad by DNAJB4 molecular chaperone is sufficient to influence cell adhesion in vitro. Using a chick embryo chorioallantoic membrane assay with gastric cancer derived cells, we demonstrate that DNAJB4 stimulates the anti-invasive function of WT Ecad in vivo. Additionally, the expression of DNAJB4 and Ecad is concomitantly decreased in human gastric carcinomas. Altogether, we demonstrate that DNAJB4 is a sensor of Ecad structural features that might contribute to gastric cancer progression.