Immunological Activity Difference between Native Calreticulin Monomers and Oligomers

Abstract

We have recently demonstrated that the greatly increased immunological activities of recombinant murine calreticulin (rCRT) are largely attributed to its self-oligomerization. Although native CRT (nCRT) can also oligomerize under stress conditions in vitro, whether this phenomenon could occur inside cells and the immunological activity difference between nCRT monomers and oligomers remained unclear. In this study, we illustrated the formation of CRT oligomers in tranfectant cells under “heat & low pH” (42°C/pH 6.5) condition. The mixture of nCRT oligomers and monomers (OnCRT) was obtained after 3 hr treatment of murine monomeric nCRT (MnCRT) under similar condition (42°C/pH 5.0) in vitro. The OnCRT thus obtained was better recognized by 2 monoclonal Abs from mice that had been immunized with oligomeric rCRT. Unlike MnCRT, OnCRT was able to elicit CRT-specific IgG production in mice. OnCRT also stimulated bone-marrow derived dendritic cells (BMDCs) to secrete significantly higher levels of TNF-α, IL-6 and IL-12p40 than did MnCRT in vitro. We postulate that oligomerization of soluble CRT may occur under certain pathophysiological conditions (e.g. ultrahyperpyrexia) and the resultant oligomers may exhibit exaggerated immunostimulating activities, thereby affiliating the inflammatory responses in vivo.

Calreticulin exploits TGF‐β for extracellular matrix induction engineering a tissue regenerative process

“…Other receptors including scavenger receptor-1 Sca-1 in the context of CRT-induced cytokine release by macrophages and endocytic mechanisms for signaling have been shown. 36,90,91 Similar to HFFs and HDFs, eCRT induction of ECM proteins, but not α5 integrin in mouse embryo fibroblasts (MEFs) is mediated by TGF-β signaling. The TGF-β-mediated fibrotic response involves regulation by iCRT 5 as CRT null mice and lung fibroblasts from idiopathic pulmonary fibrosis with siRNA CRT knockdown compared to wild-type and non-transfected cells, showed defects in both collagen and fibronectin protein production in response to TGF-β with Smad2/3 activation remaining intact.…”

Calreticulin: Challenges Posed by the Intrinsically Disordered Nature of Calreticulin to the Study of Its Function

“…Detailed structural analysis of CALR is complicated not only by the presence of long disordered regions, but also by the tendency of this protein to dimerize and self-oligomerize, a property that was shown to be important for the chaperone activity of CALR, since self-oligomerization noticeably increased CALR binding to peptides and denatured proteins (Jørgensen et al, 2003 ; Rizvi et al, 2004 ; Boelt et al, 2016 ) and also promoted its immunological activities (Huang et al, 2013 ; He et al, 2014 ). Obviously, any experimental technique that is used for the analysis of structural properties and conformational behavior of wholly disordered proteins can by applied for the investigation of isolated domains of CALR.…”

The Biophysical Interaction of the Danger-Associated Molecular Pattern (DAMP) Calreticulin with the Pattern-Associated Molecular Pattern (PAMP) Lipopolysaccharide

“…Chaperone proteins are highly active and important to processes involving the function of innate (e.g., phagocytosis, cytokine release) and adaptive immune response (e.g., peptide loading complex and antigen presentation). However, by binding to LPS and the same array of signaling receptors, for example, Hsp60, Hsp70, as well as CRT, has confounded a straightforward dissection of separate roles for chaperones in immune regulation from LPS [63,64,65,66,67,68,69,70] [39]. For example, HSPs have been shown to bind to many classes of scavenger receptors and LRP1 (CD91), and TLRs and to signal for different functions [71].…”