The Dishevelled gene was first identified in Drosophila mutants with disoriented hair and bristle polarity [1-3]. The Dsh gene (Dsh/Dvl, in Drosophila and vertebrates respectively) gained popularity when it was discovered that it plays a key role in segment polarity during early embryonic development in Drosophila [4]. Subsequently, the vertebrate homolog of Dishevelled genes were identified in Xenopus (Xdsh), mice (Dvl1, Dvl2, Dvl3), and in humans (DVL1, DVL2, DVL3) [5-10]. Dishevelled functions as a principal component of Wnt signaling pathway and governs several cellular processes including cell proliferation, survival, migration, differentiation, polarity and stem cell renewal. This review will revisit seminal discoveries and also summarize recent advances in characterizing the role of Dishevelled in both normal and pathophysiological settings.

The CYP19A1 gene encodes aromatase, an enzyme that converts androgens into estrogens and consequently directly contributes to both the depletion of androgens and the synthesis of estrogens in several organs. Aromatase is critical for diverse biological processes such as proliferation, regulation of fat metabolism and hormone signaling. Additionally, it is also overexpressed in diverse cancers and drives hormone-dependent tumor progression and increases 17-β-estradiol (E2) within tumors and the tumor microenvironment. Although the inhibition of E2 production via aromatase inhibitors represents a major therapeutic paradigm in clinical oncology, fundamental questions regarding how cancer cells gain the capacity to overexpress aromatase remain unanswered. Multiple tissue-specific CYP19A1 promoters are known to be aberrantly active in tumors, yet how this occurs is unclear. Here, for the first time, we report that Dishevelled (DVL) proteins, which are key mediators of Wnt signaling, regulate aromatase expression in multiple breast cancer cell lines. We also report that DVL enters the nucleus and localizes to at least two different CYP19A1 promoters (pII and I.4) previously reported to drive overexpression in breast tumors and to a very distal CYP19A1 placental promoter (I.1) that remains poorly characterized. We go on to demonstrate that DVL-1 and DVL-3 loss of function leads to differential changes in various aromatase transcripts and in E2 production. The report, herein, uncovers a new regulator of CYP19A1 transcription and for the first time demonstrates that DVL, a critical mediator of WNT signaling, contributes to aberrant breast cancer-associated estrogen production.

Dishevelled (DVL) proteins are central mediators of the Wnt signalling pathway and are versatile regulators of several cellular processes, yet little is known about their post-translational regulation. Acetylation is a reversible post-translational modification (PTM) which regulates the function of several non-histone proteins involved in tumorigenesis. Since we previously demonstrated that lysine deacetylase, SIRT-1, regulates DVL protein levels and its function, we reasoned that DVL could potentially be a substrate for SIRT-1 mediated deacetylation. To further examine the potential role of multiple families of lysine deacetylases in the post-translational regulation of DVL, we screened for novel acetylation sites using liquid chromatography mass-spectrometry (LC-MS/MS) analysis. Herein, we report 12 DVL-1 lysine residues that show differential acetylation in response to changes in oxygen tension and deacetylase inhibition in triple-negative breast cancer (TNBC). PTMs are well documented to influence protein activity, and cellular localization. We also identify that acetylation of two key lysine residues, K69 and K285, present on the DIX and PDZ domains respectively, promote nuclear over cytoplasmic localization of DVL-1, and influences its promoter binding and regulation of genes implicated in cancer. Collectively, these findings for the first time, uncover acetylation as a novel layer of regulation of DVL-1 proteins.

Key points• The novel signalling molecule hydrogen sulphide (H 2 S) regulates diverse cell physiological processes in several organs and systems including airway smooth muscle contractility.• We explored the mechanisms of H 2 S-induced smooth muscle relaxation in small intrapulmonary airways using lung slices and imaging approaches.• We found that exogenous and endogenous H 2 S inhibited intracellular Ca 2+ release specifically through the inositol-1,4,5-trisphosphate (InsP 3 ) receptor in smooth muscle cells and reversibly inhibited acetylcholine-induced intracellular Ca 2+ oscillations, thus leading to airway dilatation.• The effects of H 2 S on InsP 3 -induced Ca 2+ release and airway contraction were mimicked by the reducing agent dithiothreitol and inhibited by the oxidizing agent diamide, suggesting that H 2 S acts as a thiol-reducing agent to reduce Ca 2+ release through InsP 3 receptors and to evoke relaxation.• Our results suggest that endogenously produced H 2 S is a novel modulator of InsP 3 -mediated Ca 2+ signalling in airway smooth muscle and thus promotes bronchodilatation.Abstract Hydrogen sulphide (H 2 S) is a signalling molecule that appears to regulate diverse cell physiological process in several organs and systems including vascular and airway smooth muscle cell (SMC) contraction. Decreases in endogenous H 2 S synthesis have been associated with the development of cardiovascular diseases and asthma. Here we investigated the mechanism of airway SMC relaxation induced by H 2 S in small intrapulmonary airways using mouse lung slices and confocal and phase-contrast video microscopy. Exogenous H 2 S donor Na 2 S (100 μM) reversibly inhibited Ca 2+ release and airway contraction evoked by inositol-1,4,5-trisphosphate (InsP 3 ) uncaging in airway SMCs. Similarly, InsP 3 -evoked Ca 2+ release and contraction was inhibited by endogenous H 2 S precursor L-cysteine (10 mM) but not by L-serine (10 mM) or either amino acid in the presence of DL-propargylglycine (PPG). Consistent with the inhibition of Ca 2+ release through InsP 3 receptors (InsP 3 Rs), Na 2 S reversibly inhibited acetylcholine (ACh)-induced Ca 2+ oscillations in airway SMCs. In addition, Na 2 S, the H 2 S donor GYY-4137, and L-cysteine caused relaxation of airways pre-contracted with either ACh or 5-hydroxytryptamine (5-HT). Na 2 S-induced airway relaxation was resistant to a guanylyl cyclase inhibitor (ODQ) and a protein kinase G inhibitor (Rp-8-pCPT-cGMPS). The effects of H 2 S on InsP 3 -evoked Ca 2+ release and contraction as well as on the relaxation of agonist-contracted airways were mimicked by the thiol-reducing agent dithiothreitol (DTT, 10 mM) and inhibited by the oxidizing agent diamide (30 μM). These studies indicate that H 2 S causes airway SMC relaxation by inhibiting Ca 2+ release through InsP 3 Rs and consequent reduction of agonist-induced Ca 2+ oscillations in SMCs. The results suggest a novel role for endogenously produced H 2 S that involves the modulation of