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Thrombin-Par1 signaling axis disrupts COP9 signalosome subunit 3-mediated ABCA1 stabilization in inducing foam cell

Monoranjan Boro¹, Suresh Govatati¹, Raj Kumar¹

¹Department of Physiology, University of Tennessee Health Science Center, Memphis, TN, 38163, USA.

Cell Death and Differentiation

|September 24, 2020

Abstract:

ATP-binding cassette transporters A1 (ABCA1) and G1 (ABCG1) play a vital role in promoting cholesterol efflux. Although, the dysregulation of these transporters was attributed as one of the mechanisms of atherogenesis, what renders their dysfunction is not well explored. Previously, we have reported that thrombin without having any effect on ABCG1 levels depletes ABCA1 levels affecting cholesterol efflux. In this study, we explored the mechanisms underlying thrombin-induced depletion of ABCA1 levels both in macrophages and smooth muscle cells. Under normal physiological conditions, COP9 signalosome subunit 3 (CSN3) was found to exist in complex with ABCA1 and in the presence of proatherogenic stimulants such as thrombin, ABCA1 was phosphorylated and dissociated from CSN3, leading to its degradation. Forced expression of CSN3 inhibited thrombin-induced ABCA1 ubiquitination and degradation, restored cholesterol efflux and suppressed foam cell formation. In Western diet (WD)-fed ApoE^-/- mice, CSN3 was also disassociated from ABCA1 otherwise remained as a complex in Chow diet (CD)-fed ApoE^-/- mice. Interestingly, depletion of CSN3 levels in WD-fed ApoE^-/- mice significantly lowered ABCA1 levels, inhibited cholesterol efflux and intensified foam cell formation exacerbating the lipid laden atherosclerotic plaque formation. Mechanistic studies have revealed the involvement of Par1-Gα₁₂-Pyk2-Gab1-PKCθ signaling in triggering phosphorylation of ABCA1 and its disassociation from CSN3 curtailing cholesterol efflux and amplifying foam cell formation. In addition, although both CSN3 and ABCA1 were found to be colocalized in human non-lesion coronary arteries, their levels were decreased as well as dissociated from each other in advanced atherosclerotic lesions. Together, these observations reveal for the first time an anti-atherogenic role of CSN3 and hence, designing therapeutic drugs protecting its interactions with ABCA1 could be beneficial against atherosclerosis.

Insights

COP9 signalosome subunit 3 (CSN3) protects ATP-binding cassette transporter A1 (ABCA1) from degradation, preventing cholesterol buildup and foam cell formation. Maintaining CSN3-ABCA1 interaction is key to preventing atherosclerosis.

Area of Science:

Cardiovascular Biology
Molecular Medicine
Cellular Biochemistry

Background:

ATP-binding cassette transporters ABCA1 and ABCG1 are crucial for cholesterol efflux, and their dysregulation contributes to atherosclerosis.
Thrombin depletes ABCA1 levels, impairing cholesterol efflux, but the underlying mechanisms remain unclear.

Purpose of the Study:

To investigate the mechanisms behind thrombin-induced ABCA1 depletion in macrophages and smooth muscle cells.
To elucidate the role of COP9 signalosome subunit 3 (CSN3) in regulating ABCA1 stability and function.
To explore the therapeutic potential of targeting the CSN3-ABCA1 interaction in atherosclerosis.

Main Methods:

Investigated ABCA1 and CSN3 interactions in cell cultures and ApoE-/- mouse models under different dietary conditions.
Utilized Western blotting, immunoprecipitation, and immunofluorescence to assess protein levels, phosphorylation, ubiquitination, and localization.
Analyzed the impact of CSN3 modulation on cholesterol efflux, foam cell formation, and atherosclerotic plaque development.
Elucidated signaling pathways involved in ABCA1 regulation using pharmacological inhibitors and genetic manipulation.

Main Results:

CSN3 forms a complex with ABCA1 under normal conditions; thrombin induces ABCA1 phosphorylation and dissociation from CSN3, leading to ABCA1 degradation.
Overexpression of CSN3 prevents thrombin-induced ABCA1 ubiquitination and degradation, restoring cholesterol efflux and reducing foam cell formation.
In Western diet-fed ApoE-/- mice, CSN3 dissociates from ABCA1, leading to reduced ABCA1 levels, impaired cholesterol efflux, and exacerbated atherosclerosis.
The Par1-Gα12-Pyk2-Gab1-PKCθ signaling pathway mediates thrombin-induced ABCA1 phosphorylation and dissociation from CSN3.
CSN3 and ABCA1 levels are decreased and dissociated in advanced human atherosclerotic lesions.

Conclusions:

CSN3 plays a critical anti-atherogenic role by stabilizing ABCA1 and promoting cholesterol efflux.
Thrombin-induced ABCA1 degradation, mediated by CSN3 dissociation, contributes to atherosclerosis development.
Targeting the CSN3-ABCA1 interaction presents a potential therapeutic strategy for atherosclerosis.

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