Palmitoylation is a reversible post-translational addition of a 16-carbon lipid chain involved in trafficking and compartmentalizing target proteins. It is important for many cellular functions, including signaling via membrane-localized estrogen receptors (ERs). Within the nervous system, palmitoylation of ER is necessary for membrane surface localization and mediation of downstream signaling through the activation of metabotropic glutamate receptors (mGluRs). Substitution of the single palmitoylation site on ER prevents its physical association with the integral membrane protein caveolin-1 (CAV1), required for the formation of the ER/mGluR signaling complex. Interestingly, siRNA knockdown of either of two palmitoyl acyltransferases, zinc finger DHHC type–containing 7 (DHHC7) or DHHC21, also eliminates this signaling mechanism. Because ER has only one palmitoylation site, we hypothesized that one of these DHHCs palmitoylates CAV1. We investigated this possibility by using an acyl–biotin exchange assay in HEK293 cells in conjunction with DHHC overexpression and found that DHHC7 increases CAV1 palmitoylation. Substitution of the palmitoylation sites on CAV1 eliminated this effect but did not disrupt the ability of the DHHC enzyme to associate with CAV1. In contrast, siRNA-mediated knockdown of DHHC7 alone was not sufficient to decrease CAV1 palmitoylation but rather required simultaneous knockdown of DHHC21. These findings provide additional information about the overall influence of palmitoylation on the membrane-initiated estrogen signaling pathway and highlight the importance of considering the influence of palmitoylation on other CAV1-dependent processes.
Bibliographical noteFunding Information:
This work was supported by National Institutes of Health Grants R01DA035008 and R01DA041808 (to P. G. M.), R01NS040701 (to M. L. D.), and R01MH109471 (to J. M.). K. R. T. E. was supported by National Institutes of Health Grant T32DA007234 (to P. G. M.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.