In the heart, DHHC5 is a key player in the phenomenon of massive palmitoylation-dependent endocytosis (MEND) 14. DHHC5 has few confirmed substrates, but it is known to palmitoylate the plasma membrane protein flotillin-2 (Flot2), which is enriched in cholesterol-rich microdomains 13. It plays a role in the growth and invasion of certain types of non-small-cell lung cancer, and DHHC5 knockdown reduces the growth and invasion of these types of tumours 12. It is expressed in all tissues in mammals, but it is highly enriched in the heart and the brain. While the majority of PATs are localised to the Golgi, DHHC5 is localised to the plasma membrane 3, and in neurons, it cycles from the synaptic membrane to dendritic shafts in response to neuronal activity 11. Interestingly, Golga7b was also identified as a putative palmitoylated protein in both humans and mice 9, 10, although confirmation of this palmitoylation and the identity of the cognate PAT is lacking. This interaction has yet to be confirmed, and it is unknown whether Golga7b could regulate DHHC5 in a manner analogous to how DHHC9 is regulated by GCP16. Golga7b is a member of the Erf4 protein family which is closely related to GCP16 (Golga7) with 61% sequence identity. In a large-scale affinity purification–mass spectrometry study, Golga7b was identified as a putative interactor of DHHC5 8. The mechanism by which Erf4 acts on Erf2 was subsequently determined to involve stabilisation of the intermediate state where the acyl chain is transferred to the PAT before the final transfer of the acyl chain to the substrate 7. Subsequently, it was discovered that in mammalian cells, DHHC9 requires the protein GCP16, also known as Golga7, to form a complex capable of palmitoylating Ras 6. This was first demonstrated in yeast, where the palmitoylation of Ras, which is essential for its signalling and function 4, requires a complex of two proteins localised to the endoplasmic reticulum, Erf2 and Erf4 5. However, it has been shown that in some cases, additional proteins are required for PAT activity. They are also referred to as DHHC proteins as their characteristic active site motif contains the sequence Asp-His-His-Cys, which is required for palmitoylation of substrates. There are 23 PATs encoded in the human genome, all of which are integral membrane proteins 2, with a range of cellular locations and tissue expression patterns 3. For example, the regulation of the family of enzymes which attach the fatty acid onto proteins, palmitoyl acyltransferases (PATs), is still poorly understood. While palmitoylation was discovered several decades ago as a modification of viral proteins 1, the progress in our understanding of palmitoylation has been slower than that for other modifications such as phosphorylation. S-acylation (palmitoylation) is unique among post-translational modifications as it is the only fully reversible modification that involves the attachment of a lipid to a protein, in this case to the side chain of cysteine residues via a thioester bond. This work uncovers a novel mechanism of DHHC5 regulation by Golga7b and demonstrates a role for the DHHC5/Golga7b complex in the regulation of cell adhesion. Loss of DHHC5/Golga7b causes functional impairments in cell adhesion, suggesting these proteins have a wider role in cell adhesion beyond desmosome assembly. We show that desmoglein-2 and plakophilin-3 are substrates of DHHC5 and that DHHC5 and Golga7b are required for localisation of desmoglein-2 to the plasma membrane and for desmosomal patterning. Proteomic analysis of the composition of DHHC5/Golga7b-associated protein complexes reveals a striking enrichment in adhesion proteins, particularly components of desmosomes.

Palmitoylation of Golga7b prevents clathrin-mediated endocytosis of DHHC5 and stabilises it at the plasma membrane. Here, we demonstrate that DHHC5 binds to and palmitoylates a novel accessory protein Golga7b. DHHC5 is a PAT that is mainly localised at the plasma membrane with roles in synaptic plasticity, massive endocytosis and cancer cell growth/invasion. S-acylation (palmitoylation) is the only fully reversible lipid modification of proteins however, little is known about how protein S-acyltransferases (PATs) that mediate it are regulated.

Sds drop dead aerosol marc