IL-7 promotes the formation of DNA double strand breaks and DNA repair in murine pro-B cells

Alessia Lamolinara^1,2Chiara Di Lisio³Julie A Hixon⁴Pasquale Simeone^2,3Antonella De Cola^5,6Maria D Falco⁷Thomas J Meyer⁸Alessio Ferrone³Domenico Genovesi⁷Paola Lanuti^2,3Wenqing Li⁴Vincenzo De Laurenzi^2,9Manuela Iezzi^1,2Francesca Bianca Aiello^2,4,9*Scott Kenneth Durum⁴

• ¹Department of Neurosciences, Imaging and Clinical Sciences, University of Studies G. d'Annunzio Chieti and Pescara, Chieti, Italy
• ²Center for Advanced Studies and Technology (CAST), University of Studies G. d'Annunzio Chieti and Pescara, Chieti, Italy
• ³Department of Medicine and Aging Sciences, University of Studies G. d'Annunzio Chieti and Pescara, Chieti, Italy
• ⁴Cytokines and Immunity Section, Cancer Innovation Laboratory (CIL), National Institutes of Health (NIH), National Cancer Institute Frederick, Frederick, United States
• ⁵Department of Oncology, Early Cancer Institute, University of Cambridge, Cambridge, United Kingdom
• ⁶CRUK Children's Brain Tumour Centre of Excellence, University of Cambridge, Cambridge, United Kingdom
• ⁷Department of Radiation Oncology, “S.S. Annunziata” Hospital, University of Studies G. d'Annunzio Chieti and Pescara, Chieti, Italy
• ⁸CCR Collaborative Bioinformatics Resource (CCBR), National Institutes of Health (NIH), National Cancer Institute Frederick, Frederick, United States
• ⁹Department of Innovative Technologies in Medicine and Dentistry, University of Studies G. d'Annunzio Chieti and Pescara, Chieti, Italy

In pro-B cells, VDJ recombination at the immunoglobulin heavy chain locus is impaired. B cell progenitor recombination implies the formation of DNA double strand breaks (DSBs) by the RAG recombinase, which are subsequently repaired by specific mechanisms. We cultured primary murine pro-B cells with IL-7 to evaluate H2AX histone phosphorylation, a well-established marker of DSB formation (γ-H2AX foci) and the expression of proteins involved in DNA repair. Our results indicated that IL-7 upregulated the expression of several molecules involved in homologous recombination, the most accurate DSB repair mechanism. Quantitative analyses of γ-H2AX foci revealed that IL-7 significantly increased DSB formation in a time-dependent manner. Furthermore, γ-H2AX expression was altered in RAG2-deficient pro-B cells and absent in RAG1-deficient pro-B cells treated with IL-7, demonstrating the requirement of both RAG1 and RAG2 recombinase subunits. CD43 expression inversely correlates with the degree of cell differentiation and its level is often evaluated to assess the B lymphoid developmental stage. We observed that IL-7 upregulated CD43 expression and the percentage of large CD43/γ-H2AX double-positive cells, suggesting an effect on less differentiated, immature cells. Notably, we also found that IL-7 increased radiation-induced DSBs, while simultaneously supporting cell survival. This study uncovers novel effects of IL-7 on B cell differentiation, DSB formation, and DNA repair. It is well established that IL-7 promotes the proliferation and survival of acute lymphoblastic leukemia (ALL) cells. Our data suggest that drugs targeting IL-7 could improve ALL therapeutic protocols.