Abstract
Serum amyloid A protein (SAA) is an acute-phase reactant, known to mediate pro-inflammatory cellular responses. This study reports that CLA-1 (CD36 and LIMPII Analogous-1; human orthologue of the Scavenger Receptor Class B Type I (SR-BI)) mediates SAA uptake and downstream SAA signaling. Flow cytometry experiments revealed more than a 5-fold increase of Alexa-488 SAA uptake in HeLa cells stably transfected with CLA-1. Alexa 488-HDL uptake directly correlated with SAA uptake when determined in several CLA-1 stably transfected HeLa cell clones expressing various levels of CLA-1. SAA directly binds to CLA-1 as determined by cross-linking and colocalization of anti-CLA-1 antibody with SAA. SAA was co-internalized with transferrin to the endocytic recycling compartment pointing to a potential site of SAA metabolism. Alexa-488 SAA uptake in the CLA-1-overexpressing HeLa cells, as well as in THP-1 monocyte cell line, can be efficiently blocked by unlabeled SAA, high density lipoprotein, and other CLA-1 ligands. At the same time, markedly enhanced levels of phosphorylation of the mitogen-activated protein kinases (MAPKs), ERK1/2, and p38, were observed in cells stably transfected with CLA-1 cells following SAA stimulation when compared with mock transfected cells. The levels of the SAA-induced interleukin-8 (IL-8) secretion by CLA-1-overexpressing cells also significantly exceeded (5- to 10-fold) those detected for control cells. Synthetic amphipathic peptides possessing a structural alpha-helical motif inhibited SAA-induced activation of both MAPKs and IL-8 secretion in THP-1 cells. The results of this study demonstrate for the first time that CLA-1 functions as an endocytic SAA receptor and is involved in SAA-mediated cell signaling events associated with the immune-related and inflammatory effects of SAA.
Highlights
Serum amyloid A protein (SAA) is an acute-phase reactant, known to mediate pro-inflammatory cellular responses
This study reports that CLA-1 (CD36 and LIMPII Analogous-1; human orthologue of the Scavenger Receptor Class B Type I (SR-BI)) mediates SAA uptake and downstream SAA signaling
In this study, using the CLA-1-overexpressing HeLa cell model and human monocyte cell line, we demonstrate that lipid-poor SAA directly binds to CLA-1 and that CLA-1 ligands efficiently compete with SAA for CLA-1 binding
Summary
Vol 280, No 9, Issue of March 4, pp. 8031–8040, 2005 Printed in U.S.A. Serum Amyloid A Binding to CLA-1 (CD36 and LIMPII Analogous-1) Mediates Serum Amyloid A Protein-induced Activation of ERK1/2 and p38 Mitogen-activated Protein Kinases*. The results of this study demonstrate for the first time that CLA-1 functions as an endocytic SAA receptor and is involved in SAA-mediated cell signaling events associated with the immune-related and inflammatory effects of SAA. Because SAA is known to be an amphipathic protein, having two amphipathic ␣-helical regions corresponding to the 1–18 N-terminal and 72– 86 C-terminal sequences [14], we evaluated whether CLA-1 is a potential SAA receptor involved in binding, internalization, and pro-inflammatory signaling. The report of Grewal et al [42] provided additional evidence for SR-BI signaling by describing SR-BI as a primary candidate for HDL-initiated signaling through activating Ras in a protein kinase C-independent manner These multiple observations have led us to investigate the role of CLA-1 as a potential SAA-signaling receptor, contributing to the SAA-induced proinflammatory response. SAA is found to activate p44/p42 and p38 kinases, two members of the mitogen-activated protein kinase (MAPK) superfamily in both HeLa and THP-1 cells in a CLA-1dependent manner
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