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The role of calreticulin in cell adhesion

University of British Columbia

Acute lymphoblastic leukemia (ALL) is the most commonly diagnosed childhood cancer in Canada. Even though childhood ALL has a high overall survival rate, therapeutic options remain limited for those experiencing relapse; therefore, understanding the causes for treatment failure is vital. Adhesion of leukemic cells to bone marrow and the extracellular matrix provides chemotherapy protection to ALL, leading to drug resistance. In this thesis, I investigated the involvement of an endoplasmic reticulum (ER) chaperone, calreticulin (CRT), in integrin mediated cell adhesion. I show that integrin-mediated adhesion of Jurkat T-lymphoblasts is disrupted in CRT-/- cells, and CRT re-expression restores the wildtype phenotype. To determine the requirement of cytosolic CRT for integrin-mediated cell adhesion, I used ERp57-/- cells, previously characterized to express no CRT in the extra-ER compartment. Interestingly, ERp57-/- cells have decreased adhesion compared to wildtype cells, indicating the involvement of cytosolic CRT in integrin-mediated cell adhesion. To evaluate the requirement of LRP1 as a surface receptor for CRT during immunogenic cell death (ICD), I utilized CRISPR-Cas9 technology to generate LRP1-/- lymphoblasts. I show that drug-treated stimulation of surface CRT presentation is lost in LRP1-/- cells, a result consistent with LRP1 as a cis-acting receptor for CRT during ICD. I utilized CRT-/- HEK cells expressing various CRT constructs that are localized in different cellular compartments. All constructs successfully rescued the adhesion defect of CRT-/- cells. Furthermore, I found that mutant variants of CRT that lack the C-terminal KDEL ER retention motif were proteolytically truncated, and that the N-terminal truncated fragment can be detected on the cell surface.

Text

2019-09-05

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/71633

Cell and Developmental Biology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/