β2M Signals Monocytes Through Non-Canonical TGFβ Receptor Signal Transduction.
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2021
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Circulation Research
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Abstract
Rationale: Circulating monocytes can have pro-inflammatory or pro-reparative phenotypes. The
endogenous signaling molecules and pathways that regulate monocyte polarization in vivo are poorly
understood. We have shown that platelet derived beta-2 microglobulin (β2M) and transforming growth
factor beta (TGFβ) have opposing effects on monocytes by inducing inflammatory and reparative
phenotypes respectively, but each bind and signal through the same receptor. We now define the signaling
pathways involved.
Objective: To determine the molecular mechanisms and signal transduction pathways by which β2M and
TGFβ regulate monocyte responses both in vitro and in vivo.
Methods and Results: Wild-type (WT) and platelet specific β2M knockout (Plt-β2M-/-) mice were treated
intravenously with either β2M or TGFβ to increase plasma concentrations to those in cardiovascular
diseases. Elevated plasma β2M increased pro-inflammatory monocytes, while increased plasma TGFβ
increased pro-reparative monocytes. TGFβ receptor (TGFβR) inhibition blunted monocyte responses to
both β2M and TGFβ in vivo. Using imaging flow cytometry, we found that β2M decreased monocyte
SMAD2/3 nuclear localization, while TGFβ promoted SMAD nuclear translocation, but decreased noncanonical/
inflammatory (JNK and NFκB nuclear localization). This was confirmed in vitro using both
imaging flow cytometry and immunoblots. β2M, but not TGFβ, promoted ubiquitination of SMAD3 and
SMAD4, that inhibited their nuclear trafficking. Inhibition of ubiquitin ligase activity blocked noncanonical
SMAD-independent monocyte signaling and skewed monocytes towards a pro-reparative
monocyte response.
Conclusions: Our findings indicate that elevated plasma β2M and TGFβ dichotomously polarize
monocytes. Furthermore, these immune molecules share a common receptor, but induce SMAD-dependent
canonical signaling (TGFβ) versus non-canonical SMAD-independent signaling (β2M) in a ubiquitin ligase
dependent manner. This work has broad implications as β2M is increased in several inflammatory
conditions, while TGFβ is increased in fibrotic diseases.
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Keywords
Cell signaling, Cytokine, Inflammation, Platelet, Monocyte