SMARCC1 Controls the Oncogenic NOTCH1 Transcriptional Program in T-Cell Acute Lymphoblastic Leukemia

Introduction: Mutations in NOTCH1 are highly prevalent in T-ALL and aberrant NOTCH1 signaling is required for maintenance of the leukemogenic state in T-cell progenitors. However, little is known about which interaction partners are functionally required for oncogenic NOTCH1 signaling. We therefore aimed to identify previously unappreciated molecular dependencies required for NOTCH1-driven T-ALL cells.

Methods: We performed an in vitro miR-E-based loss-of-function RNAi screen employing an shRNA-library targeting 340 genes of the NOTCH1-interactome in a murine model of human intracellular NOTCH1 (hICN1)-induced T-ALL (Tx17 cell line). To identify NOTCH1-specific depletors, the screen was also performed in two murine Notch1-negative cell lines, the pro-B-cell line BaF/3 and the T-cell lymphoma cell line EL4.

Results: We identified Smarcc1 as the top NOTCH1-specific candidate in our screen. Smarcc1 knockdown in Tx17 cells resulted in rapid depletion of shRNA-expressing cells and induction of apoptosis.These results were confirmed in the NOTCH1-dependent human T-ALL cell line JURKAT and by probing public data from the depmap portal, revealing a strong SMARCC1-dependency in the two most NOTCH1-dependent cell lines tested, JURKAT and PF-382. Pulldown of human SMARCC1 in Tx17 cells and proteomic analysis revealed binding of all known SWI/SNF complex members and Notch1. Retransplantation of Tx17 cells harboring Smarcc1 or neutral shRNAs and in vivo shRNA induction led to a significantly decreased spleen infiltration of Tx17 cells transduced with the Smarcc1 shRNA compared to a neutral shRNA. RNAseq analysis of Smarcc1 depleted cells indicated a differentiation switch from DP to CD4 SP cells, implicating Smarcc1 as a mediator of NOTCH1-induced differentiation blockade.

Conclusion: By functionally interrogating the molecular networks directly involved in NOTCH1 signaling in a murine model of hICN1-induced T-ALL, we identified Smarcc1 as a major contributor to oncogenic NOTCH1 signaling. Knockdown of Smarcc1 in NOTCH1-driven T-ALL cell lines resulted in apoptosis and rapid depletion of shRNA-expressing cells. Mechanistically, Smarcc1 binds NOTCH1 and regulates a NOTCH1-dependent transcriptional program. In ongoing investigations including CUT&RUN analysis, we are currently further probing the precise mechanism through which Smarcc1 maintains aberrant NOTCH1 signaling in T-ALL.

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