Abstract
S-Acylation is a major post-translational modification affecting several cellular processes. It is particularly important for neuronal functions. This modification is catalyzed by a family of transmembrane S-acyltransferases that contain a conserved zinc finger DHHC (zDHHC) domain. Typically, eukaryote genomes encode for 7-24 distinct zDHHC enzymes, with two members also harboring an ankyrin repeat (AR) domain at their cytosolic N termini. The AR domain of zDHHC enzymes is predicted to engage in numerous interactions and facilitates both substrate recruitment and S-acylation-independent functions; however, the sequence/structural features recognized by this module remain unknown. The two mammalian AR-containing S-acyltransferases are the Golgi-localized zDHHC17 and zDHHC13, also known as Huntingtin-interacting proteins 14 and 14-like, respectively; they are highly expressed in brain, and their loss in mice leads to neuropathological deficits that are reminiscent of Huntington's disease. Here, we report that zDHHC17 and zDHHC13 recognize, via their AR domain, evolutionary conserved and closely related sequences of a [VIAP][VIT]XXQP consensus in SNAP25, SNAP23, cysteine string protein, Huntingtin, cytoplasmic linker protein 3, and microtubule-associated protein 6. This novel AR-binding sequence motif is found in regions predicted to be unstructured and is present in a number of zDHHC17 substrates and zDHHC17/13-interacting S-acylated proteins. This is the first study to identify a motif recognized by AR-containing zDHHCs.
Highlights
S-Acylation, a protein lipidation process that is essential for neuronal functions, is catalyzed by zinc finger DHHC (zDHHC) S-acyltransferases
Regions of Sequence Homology within CSP␣ and SNAP25b Are Involved in Binding to the ankyrin repeat (AR) Domain of zDHHC17/13— We have previously shown that SNAP25b and CSP␣, being S-acylated by many Golgi zDHHC enzymes [26, 27], are recruited by the AR domains of zDHHC17 and zDHHC13 [21]
To identify the regions of SNAP25b and CSP␣ that bind to the AR domain of zDHHC17/13, we created a series of truncated and point mutants of these proteins and assessed their binding to zDHHC17 and zDHHC13 (Fig. 1, A and B), using the mating-based Split Ubiquitin System (SUS) in yeast [30], which we have previously evaluated for assessment of zDHHC substrate specificity [21]
Summary
S-Acylation, a protein lipidation process that is essential for neuronal functions, is catalyzed by zDHHC S-acyltransferases. We report that zDHHC17 and zDHHC13 recognize, via their AR domain, evolutionary conserved and closely related sequences of a [VIAP][VIT]XXQP consensus in SNAP25, SNAP23, cysteine string protein, Huntingtin, cytoplasmic linker protein 3, and microtubule-associated protein 6 This novel AR-binding sequence motif is found in regions predicted to be unstructured and is present in a number of zDHHC17 substrates and zDHHC17/13-interacting S-acylated proteins. This is the first study to identify a motif recognized by AR-containing zDHHCs. Protein S-acylation ( known as palmitoylation) is a prominent post-translational modification in eukaryotes involved in the regulation of protein trafficking, localization, stability, and function; this process is catalyzed by a family of transmembrane S-acyltransferases (zDHHCs) that share a conserved catalytic. We report that zDHHC17 and zDHHC13 recognize a novel sequence motif in a number of proteins previously found to interact with zDHHC17
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