Evidence for Recombinant GRP78, CALR, PDIA3 and GPI as Mediators of Genetic Instability in Human CD34+ Cells.

Abstract

Simple SummaryFactors secreted from irradiated mesenchymal stromal cells (MSC) may exert genotoxic effects in human CD34+ cells, thereby potentially contributing to the pathogenesis of hematologic disorders such as leukemias. In a proteomics approach, we recently identified four key proteins in the secretome of X-ray-irradiated MSC, among them the three chaperones GRP78, CALR, and PDIA3, and one glycolytic enzyme GPI. Here, we demonstrate that recombinant GRP78, CALR, PDIA3 and GPI induce significant levels of genetic instability in human CD34+ cells. Our data suggest that GRP78, CALR, PDIA3 and GPI released from irradiated MSC act as mediators of genetic instability in human CD34+ cells with potential implications for radiation-induced hematologic disorders.Soluble factors released from irradiated human mesenchymal stromal cells (MSC) may induce genetic instability in human CD34+ cells, potentially mediating hematologic disorders. Recently, we identified four key proteins in the secretome of X-ray-irradiated MSC, among them three endoplasmic reticulum proteins, the 78 kDa glucose-related protein (GRP78), calreticulin (CALR), and protein disulfide-isomerase A3 (PDIA3), as well as the glycolytic enzyme glucose-6-phosphate isomerase (GPI). Here, we demonstrate that exposition of CD34+ cells to recombinant GRP78, CALR, PDIA3 and GPI induces substantial genetic instability. Increased numbers of γH2AX foci (p < 0.0001), centrosome anomalies (p = 0.1000) and aberrant metaphases (p = 0.0022) were detected in CD34+ cells upon incubation with these factors. Specifically, γH2AX foci were found to be induced 4–5-fold in response to any individual of the four factors, and centrosome anomalies by 3–4 fold compared to control medium, which contained none of the recombinant proteins. Aberrant metaphases, not seen in the context of control medium, were detected to a similar extent than centrosome anomalies across the four factors. Notably, the strongest effects were observed when all four factors were collectively provided. In summary, our data suggest that specific components of the secretome from irradiated MSC act as mediators of genetic instability in CD34+ cells, thereby possibly contributing to the pathogenesis of radiation-induced hematologic disorders beyond direct radiation-evoked DNA strand breaks.