Palmitoyl Acyltransferases Control the Membrane Localization of DNAJC5 to Regulate Unconventional Protein Secretion

Area of Science:

• Molecular Biology
• Cell Biology
• Biochemistry

Background:

• Unconventional protein secretion (UcPS) allows cells to secrete proteins lacking signal sequences.
• Misfolding-associated protein secretion (MAPS) is a UcPS pathway involving HSP70 and DNAJC5.
• DNAJC5 is a co-chaperone associated with HSP70 in MAPS.

Purpose of the Study:

• To investigate the role of DNAJC5 post-translational modifications in MAPS.
• To identify the enzymes responsible for DNAJC5 modification and their impact on secretion.
• To determine the specific regions of DNAJC5 critical for its localization and secretion.

Main Methods:

• Human cell culture and transfection.
• Analysis of protein palmitoylation using DHHC enzymes.
• Mutagenesis studies to identify functional domains of DNAJC5.
• Confocal microscopy to assess subcellular localization (Golgi association).
• Measurement of protein secretion levels.

Main Results:

• DNAJC5 is palmitoylated by multiple DHHC palmitoyl acyltransferases in human cells.
• Overexpression of DHHC11 leads to Golgi enrichment of DNAJC5 and increased secretion.
• A minimal DNAJC5 module (DC95) containing the cysteine string domain is sufficient for palmitoylation, Golgi translocation, and secretion.
• Specific residues within or flanking the cysteine string domain are essential for these processes.

Conclusions:

• Palmitoylation is a key regulatory mechanism for DNAJC5 function in MAPS.
• The palmitoylation sites and adjacent sequences dictate DNAJC5's subcellular localization and its role in UcPS.
• This study elucidates a novel regulatory mechanism controlling the unconventional secretion of misfolded proteins.