Background/Purpose: Bruton’s Tyrosine Kinase (Btk) regulates innate immune responses downstream of multiple Toll-like receptors (TLRs), including TLR7. Given the role of TLR7 in driving type I interferon and inflammatory cytokine production in the autoimmune condition systemic lupus erythrematosus (SLE), pharmacological inhibition of the tyrosine kinase Btk has been proposed as a potential therapeutic. Given this interest it is essential that the role of Btk downstream of TLR7 activation be thoroughly explored.

Methods: Primary WT and Btk deficient murine macrophage were used to carry out a 2D proteomic study to identify differences downstream of TLR7 stimulation. Immunoprecipitation studies were carried out in macrophages and the human monocytic cell line THP-1. Confocal analysis was used to investigate localisation of proteins as well as uptake of apoptotic cells by primary macrophages.

Results: Our data describes a novel role for Btk as a regulator of the molecular chaperone calreticulin (CRT). Studies in primary macrophages demonstrate that CRT undergoes TLR7-induced tyrosine phosphorylation in a Btk-dependent manner. In addition Btk can directly interact with and phosphorylate CRT. Given the key role CRT in apoptotic cell uptake, we observe that in the absence of Btk, CRT fails to localise to the cell membrane or interact with the transmembrane receptor CD91, also known to be required for apoptotic cell uptake. Also, Btk-deficient macrophages show an impaired uptake of human apoptotic neutrophils in a phagocytosis assay as well as reduced staining for CRT within the phagocytic cup. In a preliminary study, pharmacological inhibition of Btk in monocytes derived from healthy controls demonstrated a reduction in apoptotic cell uptake following Btk inhibition, an effect that was exacerbated in monocytes derived from SLE patients.  

Conclusion: Overall our data has revealed a novel regulatory role for the tyrosine kinase Btk with regard to CRT. Loss of Btk results in an alteration in the cellular trafficking of CRT downstream of TLR7 stimulation in myeloid cells as well as a reduction in their ability clear apoptotic cells. Given the interest in the use of pharmacological inhibitors of Btk for the treatment of SLE, our data indicates that further investigation into the effect of Btk inhibition with regard to myeloid cell function is warranted.  

This work was funded by Science Foundation Ireland under grant no. 08/IN.1/B2091

ACR Meeting Abstracts - https://acrabstracts.org/abstract/bruton%e2%80%b2s-tyrosine-kinase-and-calreticulin-a-novel-interaction-with-implications-for-inflammatory-and-autoimmune-disease/