Person:

Bettigole, Sarah Elizabeth

The transcription factor XBP1 has been linked to the development of highly secretory tissues such as plasma cells and Paneth cells, yet its function in granulocyte maturation has remained unknown. Here we discovered an unexpectedly selective and absolute requirement for XBP1 in eosinophil differentiation without an effect on the survival of basophils or neutrophils. Progenitors of myeloid cells and eosinophils selectively activated the endoribonuclease IRE1α and spliced Xbp1 mRNA without inducing parallel endoplasmic reticulum (ER) stress signaling pathways. Without XBP1, nascent eosinophils exhibited massive defects in the post-translational maturation of key granule proteins required for survival, and these unresolvable structural defects fed back to suppress critical aspects of the transcriptional developmental program. Hence, we present evidence that granulocyte subsets can be distinguished by their differential reliance on secretory-pathway homeostasis.

The endoplasmic reticulum (ER) is a critical regulator of cellular homeostasis, primarily responsible for handling calcium storage and signaling, lipid synthesis, and the proper glycosylation and folding of nascent transmembrane and secreted proteins. Numerous stimuli such as dysregulated oxidative stress, depletion of calcium stores, hypo- and hyperglycemia, hypoxia, inefficient cellular degradation pathways, and inflammation can disrupt the protein folding capacity of the ER, leading to a condition known as ER stress. The canonical unfolded protein response (UPR) is a three-pronged signaling axis charged with ameliorating ER stress, and is particularly important for the development of highly secretory tissues such as plasma cells, pancreatic acinar cells, and Paneth cells. Here we examine the requirement for the ER stress response transcription factor XBP1 in hematopoiesis, focusing specifically on granulocyte development. In this study we found that XBP1 is selectively required for the development of the eosinophil lineage, but not for other granulocyte lineages. Targeted hematopoietic ablation of XBP1 or its upstream activator IRE1α using Vav1-Cre conditional knockout mice resulted in complete loss of mature eosinophils and dramatic decrease in Lin-, Sca1-, CD34+, c-Kitlo, IL-5Rα+ eosinophil progenitors without altering neighboring hematopoietic lineages such as basophils and neutrophils. Myeloid and eosinophil progenitors selectively activated XBP1 without induction of parallel canonical ER stress signaling pathways. Through the use of mixed bone marrow chimeras, in vitro bone marrow-derived eosinophil cultures, and eosinophil-selective Cre mouse models, we demonstrated that XBP1 is required after eosinophil lineage commitment in a cell-intrinsic manner to sustain cell viability. RNA-sequencing and bioinformatic analyses of hematopoietic progenitors at different stages of eosinophil differentiation revealed that Xbp1 deficiency reduced the adaptive protein folding capacity of the ER. Upon eosinophil commitment, this vulnerability led to massive defects in post-translational maturation of key granule proteins required for survival, and these unresolvable structural defects fed back to suppress critical aspects of the transcriptional developmental program.
Taken together, we present the first evidence that granulocyte subsets can be distinguished by their differential sensitivities to perturbations in XBP1-mediated secretory pathway functions. Furthermore, this work implicates the IRE1α/XBP1 signaling axis as a potential therapeutic target for eosinophil-mediated diseases.