Palmitoylation of Desmoglein 2 Is a Regulator of Assembly Dynamics and Protein Turnover

Abstract

Desmosomes are prominent adhesive junctions present between many epithelial cells as well as cardiomyocytes. The mechanisms controlling desmosome assembly and remodeling in epithelial and cardiac tissue are poorly understood. We recently identified protein palmitoylation as a mechanism regulating desmosome dynamics. In this study, we have focused on the palmitoylation of the desmosomal cadherin desmoglein-2 (Dsg2) and characterized the role that palmitoylation of Dsg2 plays in its localization and stability in cultured cells. We identified two cysteine residues in the juxtamembrane (intracellular anchor) domain of Dsg2 that, when mutated, eliminate its palmitoylation. These cysteine residues are conserved in all four desmoglein family members. Although mutant Dsg2 localizes to endogenous desmosomes, there is a significant delay in its incorporation into junctions, and the mutant is also present in a cytoplasmic pool. Triton X-100 solubility assays demonstrate that mutant Dsg2 is more soluble than wild-type protein. Interestingly, trafficking of the mutant Dsg2 to the cell surface was delayed, and a pool of the non-palmitoylated Dsg2 co-localized with lysosomal markers. Taken together, these data suggest that palmitoylation of Dsg2 regulates protein transport to the plasma membrane. Modulation of the palmitoylation status of desmosomal cadherins can affect desmosome dynamics.

S-acylated Golga7b stabilises DHHC5 at the plasma membrane to regulate desmosome assembly and cell adhesion

“…Our unbiased AP-MS experiments uncovered a new role for DHHC5 in cell adhesion. Palmitoylation regulates several proteins involved in cell:cell and cell:matrix adhesion (Little et al, 1998;Roberts et al, 2014;Lievens et al, 2016;Roberts et al, 2016;Aramsangtienchai et al, 2017) and for many of them, palmitoylation regulates their localisation. Among cell adhesion complexes, desmosomes are notable for the number of proteins in the complex that are palmitoylated, with at least 6 members of the complex palmitoylated in A431 cells (Roberts et al, 2014), and in this work, we have shown that DHHC5 is the PAT responsible for the palmitoylation of the key complex members desmoglein-2 and plakophilin-3.…”

“…Among these is the important desmosomal cadherin desmoglein-2 (DSG2, Figure 4d). DSG2 is a known palmitoylated protein (Roberts et al, 2016), where its palmitoylation regulates the localisation and trafficking of the protein to the plasma membrane. Because of this, we decided to investigate whether DHHC5 is the PAT responsible for DSG2 palmitoylation, so we performed an ABE assay on DHHC5 depleted HeLa cells and control siRNA treated cells (Figure 6a).…”

“…Palmitoylation of plakophilin-3 is essential for both its inclusion into desmosomes and is required for the efficient assembly of desmosomes. One of the two desmosomal cadherins, desmoglein-2, is palmitoylated on a pair of cysteines (Roberts et al, 2016). Mutation of the palmitoylation sites of desmoglein-2 does not prevent incorporation of desmoglein-2 into desmosomes but does cause a trafficking defect with a portion of desmoglein-2 localised to lysosomes.…”

Proteomic analysis of the desmosome identifies novel components required for skin integrity

“…Considering that SH2/SH3 adaptor proteins are often involved in tyrosine kinase signaling, the interaction between these networks and desmosomes deserves increased attention. In addition to these novel pathways, our BioID analysis also highlights possible regulators of desmosome stability, such as a single palmitoyltransferase and thioesterase that are excellent candidates to mediate the palmitoylation of desmosomal proteins (49,50) .…”

Mass‐Tag Labeling Using Acyl‐PEG Exchange for the Determination of Endogenous Protein S‐Fatty Acylation

“…We speculate this might be due to differences in transfer to the western blot membrane, or PEG-mediated differences in exposure of the protein to antibodies or HRP. While the signal differences challenge the accuracy of the S-fatty acylation ratios, the APE still provides a reliable method to determine the qualitative measurement of S-fatty acylation states (Roberts et al, 2016), as well as a semi-quantitative comparative assay between different conditions (Nathan P. Westcott, In Press; Yokoi et al, 2016). …”