Dynamics of CLIMP-63 S-acylation control ER morphology

Abstract

The complex architecture of the endoplasmic reticulum (ER) comprises distinct dynamic features, many at the nanoscale, that enable the coexistence of the nuclear envelope, regions of dense sheets and a branched tubular network that spans the cytoplasm. A key player in the formation of ER sheets is cytoskeleton-linking membrane protein 63 (CLIMP-63). The mechanisms by which CLIMP-63 coordinates ER structure remain elusive. Here, we address the impact of S-acylation, a reversible post-translational lipid modification, on CLIMP-63 cellular distribution and function. Combining native mass-spectrometry, with kinetic analysis of acylation and deacylation, and data-driven mathematical modelling, we obtain in-depth understanding of the CLIMP-63 life cycle. In the ER, it assembles into trimeric units. These occasionally exit the ER to reach the plasma membrane. However, the majority undergoes S-acylation by ZDHHC6 in the ER where they further assemble into highly stable super-complexes. Using super-resolution microscopy and focused ion beam electron microscopy, we show that CLIMP-63 acylation-deacylation controls the abundance and fenestration of ER sheets. Overall, this study uncovers a dynamic lipid post-translational regulation of ER architecture.

S‐acylation: an orchestrator of the life cycle and function of membrane proteins

“…While different FA chain lengths, from C14 to C22, may be transferred to the target [9,43], the most frequently transferred FA is palmitate (16-carbon acyl chain) [44]; hence, S-acylation is often referred to as S-palmitoylation. Each ZDHHC enzyme displays a specific localization pattern throughout the endomembrane system, between the endoplasmic reticulum (ER) (e.g., ZDHHC4/6/7/9), the Golgi (e.g., ZDHHC3/4/8/2/20), [22,26,29] and the endocytic system, including the plasma membrane, endosomes, and vesicular structures (e.g., ZDHHC2/5/20) [22,26,45]. For some ZDHHCs, the subcellular distribution can be controlled by defined cytosolic motifs [46,47] but may vary between cell types or species [23,48,49].…”

“…Both IP3R and CLIMP-63 interact with mitochondrial porin voltagedependent anion-selective channels (VDACs) at MCSs and coordinate ER-mitochondrial calcium exchanges [101] in an S-acylation dependent manner [100,102]. Interestingly, calnexin, CLIMP-63, IP3R, and TMX4 are all modified by the same acyltransferase, namely ZDHHC6 [22,52,103,104], which could act as a master regulator of some ER-mitochondrial contacts.…”

“…2. These include modulation of protein conformation [12][13][14][15][16]; capacity to transduce signals [17][18][19]; ability to interact with other proteins [18,20,21]; regulation of their trafficking and localization [12,[22][23][24][25]; physicochemical properties of their transmembrane domains (TMDs) and their association with surrounding lipids [26][27][28]; and cross-talk with other PTMs [29][30][31]. By modifying such a vast number of proteins and affecting their function through a broad diversity of means, S-acylation is involved in various diseases, including neurodegenerative diseases, cancer, and inflammatory diseases (reviewed in [10,19,32]).…”

The Multifaceted Role of Human Dickkopf-3 (DKK-3) in Development, Immune Modulation and Cancer

“…Quite recently, Kimura et al [ 31 ] discovered that DKK-3, as well as the other DKKs, physically bind to CKAP4 (cytoskeleton-associated protein 4, also known as P63, CLIMP-63 or ERGIC-63). CKAP4 is a type II transmembrane protein that is primarily expressed in the endoplasmic reticulum (ER) and regulates its architecture [ 32 ]. Yet, a minor fraction of CKAP4 (less than 10% of the total) is transported to the plasma membrane of specific cell types, including pulmonary, vascular smooth muscle, and bladder epithelial cells [ 33 ].…”

Refining S-acylation: Structure, regulation, dynamics, and therapeutic implications

“…The tree was constructed using the neighbor-joining method and was visualized using iTOL ( Letunic and Bork, 2011 ). ZDHHCs are color-coded based on their reported subcellular localization(s) ( Abrami et al, 2017 , 2021 ; Ernst et al, 2018 ; Sandoz et al, 2023 ), and examples of substrates for each enzyme are shown in black (for a complete list of reported S-acylated proteins visit https://www.swisspalm.org/ [ Blanc et al, 2015 ]).…”