Integrated multi-modal analysis reveals tumor and immune characteristics that distinguish Epstein-Barr virus-positive and -negative B cell post-transplant lymphoproliferative disorders [scRNA-seq]

The highly prevalent oncogenic Epstein-Barr virus (EBV) can drive tumorigenesis with disrupted host immunity, causing malignancies including post-transplant lymphoproliferative disorders (PTLD). PTLD can also arise in the absence of EBV but the biological differences underlying EBV(+) and EBV(-) B cell PTLD and the associated host-EBV-tumor interactions remain poorly understood. Here, we reveal the core differences between EBV(+) and EBV(-) PTLD, characterized by increased expression of genes related to immune processes or DNA interactions respectively, and the augmented ability of EBV(+) PTLD B cells to modulate the tumor microenvironment through elaboration of monocyte-attracting cytokines/chemokines. We create a reference resource of proteins distinguishing EBV(+) B lymphoma cells from EBV(-) B lymphoma including the immunomodulatory molecules CD300a and CD24, respectively. Moreover, we show that CD300a is essential for maximal survival of EBV(+) PTLD B lymphoma cells. Our comprehensive multi-modal analyses uncover the biological underpinnings of PTLD and offer opportunities for precision therapies. Overall design: CD300A knockout (KO) cell lines were generated from three EBV(+) B PTLD patient-derived spontaneous lymphoblastoid cell lines (AB5, JB7, VB5) via CRISPR/Cas9 gene editing, using nucleofection to introduce ribonucleoprotein complexes composed of Cas9-2NLS and CD300A sgRNA. Corresponding mock controls for each cell line were also generated via nucleofection with Cas9-2NLS in the absence of sgRNA. No single-cell cloning was performed for CD300A KO cell lines. The resulting bulk cell populations were analysed using scRNAseq.