The membrane protein claudin-3 (CLDN3) is critical for the formation and maintenance of tight junction and its high expression has been implicated in dictating malignant progression in various cancers. However, the post-translational modification of CLDN3 and its biological function remains poorly understood. Here, we report that CLDN3 is positively correlated with ovarian cancer progression both in vitro and in vivo. Of interest, CLDN3 undergoes S-palmitoylation on three juxtamembrane cysteine residues, which contribute to the accurate plasma membrane localization and protein stability of CLDN3. Moreover, the deprivation of S-palmitoylation in CLDN3 significantly abolishes its tumorigenic promotion effect in ovarian cancer cells. By utilizing the co-immunoprecipitation assay, we further identify ZDHHC12 as a CLDN3-targating palmitoyltransferase from 23 ZDHHC family proteins. Furthermore, the knockdown of ZDHHC12 also significantly inhibits CLDN3 accurate membrane localization, protein stability and ovarian cancer cells tumorigenesis. Thus, our work reveals S-palmitoylation as a novel regulatory mechanism that modulates CLDN3 function, which implies that targeting ZDHHC12-mediated CLDN3 S-palmitoylation might be a potential strategy for ovarian cancer therapy.
The metabotropic glutamate receptor 2 (mGlu2) is a transmembrane-spanning class C G protein-coupled receptor that is an attractive therapeutic target for multiple psychiatric and neurological disorders. A key challenge has been deciphering the contribution of mGlu2 relative to other closely related mGlu receptors in mediating different physiological responses, which could be achieved through the utilization of subtype selective pharmacological tools. In this respect, allosteric modulators that recognize ligand-binding sites distinct from the endogenous neurotransmitter glutamate offer the promise of higher receptor-subtype selectivity. We hypothesized that mGlu2-selective positive allosteric modulators could be derivatized to generate bifunctional pharmacological tools. Here we developed clickable photoaffinity probes for mGlu2 based on two different positive allosteric modulator scaffolds that retained similar pharmacological activity to parent compounds. We demonstrate successful probe-dependent incorporation of a commercially available clickable fluorophore using bioorthogonal conjugation. Importantly, we also show the limitations of using these probes to assess in situ fluorescence of mGlu2 in intact cells where significant nonspecific membrane binding is evident.
