Cargo-Specific Role for Retriever Subunit VPS26C in Hepatocyte Lipoprotein Receptor Recycling to Control Postprandial Triglyceride-Rich Lipoproteins

Abstract

BACKGROUND: The coiled-coil domain containing 22/coiled-coil domain containing 93/copper metabolism MURR1 domains (CCC) complex is required for the transport of low-density lipoprotein receptor (LDLR) and LRP1 (LDLR-related protein 1) from endosomes to the cell surface of hepatocytes. Impaired functioning of hepatocytic CCC causes hypercholesterolemia in mice, dogs, and humans. Retriever, a protein complex consisting of subunits VPS26C, VPS35L, and VPS29, is associated with CCC, but its role in endosomal lipoprotein receptor transport is unclear. We here investigated the contribution of retriever to hepatocytic lipoprotein receptor recycling and plasma lipids regulation. METHODS: Using somatic CRISPR/Cas9 gene editing, we generated liver-specific VPS35L or VPS26C-deficient mice. We determined total and surface levels of LDLR and LRP1 and plasma lipids. In addition, we studied the protein levels and composition of CCC and retriever. RESULTS: Hepatocyte VPS35L deficiency reduced VPS26C levels but had minimal impact on CCC composition. VPS35L deletion decreased hepatocytic surface expression of LDLR and LRP1, accompanied by a 21% increase in plasma cholesterol levels. Hepatic VPS26C ablation affected neither levels of VPS35L and CCC subunits, nor plasma lipid concentrations. However, VPS26C deficiency increased hepatic LDLR protein levels by 2-fold, probably compensating for reduced LRP1 functioning, as we showed in VPS26C-deficient hepatoma cells. Upon PCSK9 (proprotein convertase subtilisin/kexin type 9)-mediated LDLR elimination, VPS26C ablation delayed postprandial triglyceride clearance and increased plasma TG levels by 26%. CONCLUSIONS: Our study suggests that VPS35L is shared between retriever and CCC to facilitate LDLR and LRP1 transport from endosomes to the cell surface. Conversely, retriever subunit VPS26C selectively transports LRP1, but not LDLR, and thereby may control hepatic uptake of postprandial TG-rich lipoprotein remnants.

Structural organization of the retriever–CCC endosomal recycling complex

“…Retriever and CCC play crucial roles in cellular and organismal physiology, as evident by myriad phenotypes identified thus far when these systems are disturbed, ranging from developmental alterations in humans and mouse models [36][37][38][39][40][41] , to changes in copper 25,[42][43][44] and lipid metabolism 24,45,46 , and alterations in immune signaling [47][48][49] and function [50][51][52][53] . However, despite two decades of work in this area, the structural mechanism underlying the assembly of these complexes remained elusive.…”

Mechanism and regulation of cargo entry into the Commander recycling pathway

“…Retriever is an essential cargo sorting complex and is a stable heterotrimer of VPS26C, VPS35L and VPS29, that together with the dodecameric CCDC22, CCDC93, COMMD (CCC) complex and DENND10, forms the 16subunit Commander super-assembly (Phillips-Krawczak et al, 2015;Mallam and Marcotte, 2017;McNally et al, 2017;Boesch et al, 2023;Healy et al, 2023;Laulumaa et al, 2023). Defects in Commander assembly and function are associated with metabolic disorders including hypercholesterolemia (Bartuzi et al, 2016;Fedoseienko et al, 2018;Rimbert et al, 2020;Vos et al, 2023), viral infection (Daniloski et al, 2021;Zhu et al, 2021), and lead to Ritscher-Schinzel syndrome, a multi-system developmental disorder characterized by abnormal craniofacial features, cerebellar hypoplasia, and stunted cardiovascular development (Kolanczyk et al, 2015;Kato et al, 2020;Otsuji et al, 2023).…”

“…Retriever is an essential cargo sorting complex and is a stable heterotrimer of VPS26C, VPS35L and VPS29, that together with the dodecameric CCDC22, CCDC93, COMMD (CCC) complex and DENND10, forms the 16-subunit Commander super-assembly (Phillips-Krawczak et al, 2015; Mallam and Marcotte, 2017; McNally et al, 2017; Boesch et al, 2023; Healy et al, 2023; Laulumaa et al, 2023). Defects in Commander assembly and function are associated with metabolic disorders including hypercholesterolemia (Bartuzi et al, 2016; Fedoseienko et al, 2018; Rimbert et al, 2020; Vos et al, 2023), viral infection (Daniloski et al, 2021; Zhu et al, 2021), and lead to Ritscher-Schinzel syndrome, a multi-system developmental disorder characterized by abnormal craniofacial features, cerebellar hypoplasia, and stunted cardiovascular development (Kolanczyk et al, 2015; Kato et al, 2020; Otsuji et al, 2023). In the Commander trafficking pathway sequence-dependent integral protein recognition is principally mediated by the cargo adaptor sorting nexin-17 (SNX17) (McNally et al, 2017), the FERM domain of which binds to a ØxNxx[Y/F] sorting motif presented in the cytoplasmic facing domains of integral proteins (where Ø is a hydrophobic residue and x is any residue) (van Kerkhof et al, 2005; Bottcher et al, 2012; Steinberg et al, 2012; Ghai et al, 2013).…”

Monocytic Differentiation of Human Acute Myeloid Leukemia Cells: A Proteomic and Phosphoproteomic Comparison of FAB-M4/M5 Patients with and without Nucleophosmin 1 Mutations

“…• Clathrin. Another small network included the clathrin interactor 1 (CLINT1), clathrin light chain beta (CLTB), and aftiphilin (AFTPH) (Table 2 and S3) [37,38,[51][52][53][54][55][56][57][58][59][60][61][62][63][64]. Clathrinmediated endocytosis is important for cellular nutrition as well as receptor-mediated endocytosis, membrane recycling, and control of signal pathways [26,30].…”