Ubiquitination is associated with lysosomal degradation of cell surface-resident ATP-binding cassette transporter A1 (ABCA1) through the endosomal sorting complex required for transport (ESCRT) pathway

Abstract

ATP-binding cassette transporter A1 (ABCA1) plays an essential role in the biogenesis of high-density lipoprotein in liver and in the prevention of foam cell formation in macrophages by mediating the efflux of cellular cholesterol and phospholipids to apolipoprotein A-I (apoA-I). Our current study investigated the mechanism of degradation of cell surfaceresident ABCA1, focusing on ubiquitination. A coimmunoprecipitation study indicated the presence of ubiquitinated ABCA1 in the plasma membrane of the human hepatoma cell line, HuH-7, of cells from mouse liver, and of macrophages differentiated from the human acute monocytic leukemia cell line, THP-1 (THP-1 macrophages). In HuH-7 cells, degradation of cell surface-resident ABCA1 was inhibited by the overexpression of a dominant-negative form of ubiquitin. Moreover, the disruption of the endosomal sorting complex required for transport (ESCRT) pathway, a dominant mechanism for ubiquitinationmediated lysosomal degradation, by the knockdown of hepatocyte growth factor-regulated tyrosine kinase substrate (HRS), significantly delayed the degradation of cell surface-resident ABCA1. This was accompanied by an increase in ABCA1 expression as well as in apoA-I-mediated [ 3 H]-cholesterol efflux function. The effect of HRS knockdown was also observed after calpain inhibitor treatment, which is reported to retard ABCA1 degradation. The induction of ABCA1 by HRS knockdown was confirmed in THP-1 macrophages. Conclusion: Together with the fact that lysosomal inhibitor treatments increased ABCA1 expression in HuH-7 and THP-1 macrophages, these results suggest that ubiquitination mediates the lysosomal degradation of cell surface-resident ABCA1 through the ESCRT pathway, and thereby controls the expression and cholesterol efflux function of ABCA1. This mechanism seems to mediate ABCA1 degradation independently of the calpain-involving pathway. The modulation of ABCA1 ubiquitination could thus be a potential new therapeutic target for antiatherogenic drugs. (HEPATOLOGY 2011;54:631-643)

Genetic and Pharmacologic Inhibition of the Neutrophil Elastase Inhibits Experimental Atherosclerosis

“…More importantly, our data further demonstrate that ABCA1 protein degradation, but not its transcription and translation, was altered by NE gene inactivation or exogenous HNE treatment. Currently, 2 major signal pathways have been implicated in ABCA1 protein degradation: PEST sequence‐mediated calpain proteolysis55 and the proteasome‐lysosome system 56, 57. We found no evidence to suggest that the calpain pathway is the underlying mechanism of NE‐promoted proteolysis of ABCA1, since NE‐mediated ABCA1 protein degradation was not affected by calpain‐specific inhibition.…”

Unexpected requirement for a binding partner of the syntaxin family in phagocytosis by murine testicular Sertoli cells

“…(i) SCs transfected with Ctrl siRNA or Txlna siRNA were pretreated with PD168393 inhibitor (10 μM) for 6 h, followed by the binding and phagocytic analysis as described in (h) has serine phosphorylation sites that negatively regulate ABCA1 expression. 43 Here, we show that upon activation, EGFR interacts with ABCA1, promoting its serine phosphorylation and ubiquitination (Figures 6c and e-g). This effect occurred in cells expressing WT EGFR upon EGF binding as well as in cells with activating mutant EGFR (Figures 6d and g).…”

Inhibition of ABCA1 protein degradation promotes HDL cholesterol efflux capacity and RCT and reduces atherosclerosis in mice

“…Thus, via effects on both ABCA1-expressing cells and circulating HDL function, the attenuation of ABCA1 degradation by IMM-H007 results in greater capacity for HDL to mediate RCT, and this likely underlies the atheroprotection that the intervention affords. The ABCA1 protein undergoes degradation via both calpain-mediated and ubiquitin-mediated processes ( 18,32 ). The former is dependent on the protease calpain to hydrolyze the ABCA1 protein, and the PEST-enriched motif is required ( 30 ), and ubiquitin mediates ABCA1 protein proteolysis by both lysosomal and nonlysosomal degradation pathways ( 18 ).…”

“…The ABCA1 protein undergoes degradation via both calpain-mediated and ubiquitin-mediated processes ( 18,32 ). The former is dependent on the protease calpain to hydrolyze the ABCA1 protein, and the PEST-enriched motif is required ( 30 ), and ubiquitin mediates ABCA1 protein proteolysis by both lysosomal and nonlysosomal degradation pathways ( 18 ). We determined that IMM-H007 decreases intracellular calpain activity and it also decreases Ca 2+ -dependent ABCA1 turnover, indicating that the agent of ABCA1 protein degradation affects HDL function has been unknown.…”

“…To investigate ABCA1 abundance on the plasma membrane and the degradation of the cell surface-resident protein, biotinylation was carried out on THP-1 macrophages as previously described ( 13,17,18 ). Briefl y, cells were incubated at 4°C for 60 min with PBS containing EZ-Link sulfo-NHS-SS-biotin (Pierce).…”