Pannexin 1 mutation found in melanoma tumor reduces phosphorylation, glycosylation, and trafficking of the channel-forming protein.

Pannexin 1 (PANX1) is a glycoprotein that forms large pore channels capable of passing ions and metabolites such as ATP for cellular communication. PANX1 has been implicated in many diseases including breast cancer and melanoma, where inhibition or deletion of PANX1 reduced the tumorigenic and metastatic properties of the cancer cells. We interrogated the effect of single amino acid changes in various PANX1 domains using naturally occurring variants reported in cancer patient tumors. We found that a previously reported variant (Q5H) is present in cancer cells, but was not different from the wild type (Q5) in glycosylation, trafficking, or channel function and did not affect cellular properties. We discovered that the Q5H variant is in fact the highly conserved ancestral allele of PANX1 with 89% of humans carrying at least one Q5H allele. Another mutated form Y150F, found in a melanoma patient tumor, prevented phosphorylation at Y150 as well as complex N-glycosylation while increasing intracellular localization. Sarcoma (SRC) is the predicted kinase to phosphorylate the Y150 residue, and its phosphorylation is not likely to be constitutive, but rather dynamically regulated. The Y150 phosphorylation site is the first one reported to play a role in regulating posttranslational modifications and trafficking of PANX1, with potential consequences on its large-pore channel structure and function in melanoma cells.

Targeting mRNA stability arrests inflammatory bone loss.

Many proinflammatory cytokines contain adenylate-uridylate-rich elements (AREs) within the 3'-untranslated region (UTR) that confer rapid mRNA destabilization. During the inflammatory response, cytokine mRNA are stabilized via complex interactions with RNA-binding proteins controlled by phosphorylation via multiple signaling pathways including the mitogen-activated protein kinases (MAPKs). In the absence of inflammation, a key cytokine-regulating RNA-binding protein, tristetraprolin (TTP), shuttles mRNA transcripts to degradation machinery in order to maintain low levels of inflammatory cytokines. Using this general model of mRNA decay, over expression of TTP was evaluated in an experimental model of inflammatory bone loss to determine whether altering cytokine mRNA stability has an impact in pathological bone resorption. Using adenoviral-delivered TTP, significant reductions of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and prostaglandin (PG)E(2) were observed in vitro through a mechanism consistent with targeting mRNA stability. In vivo analysis indicates a significant protective effect from inflammation-induced bone loss and inflammatory infiltrate in animals overexpressing TTP compared with reporter controls. These findings provide experimental evidence that mRNA stability is a valid therapeutic target in inflammatory bone loss.