RT Journal Article SR Electronic T1 Single-cell multi-omic analysis of thymocyte development reveals NFAT as a driver of CD4/CD8 lineage commitment JF bioRxiv FD Cold Spring Harbor Laboratory SP 2021.07.12.452119 DO 10.1101/2021.07.12.452119 A1 Zoƫ Steier A1 Laura L. McIntyre A1 Lydia K. Lutes A1 Tse-Shun Huang A1 Ellen A. Robey A1 Nir Yosef A1 Aaron Streets YR 2021 UL http://biorxiv.org/content/early/2021/07/13/2021.07.12.452119.abstract AB CD4 and CD8 T cells play critical roles in the mammalian immune system. While their development within the thymus from the CD4+CD8+ stage has been widely studied as a model of lineage commitment, the underlying mechanism remains unclear. To deconstruct this process, we apply CITE-seq, measuring the transcriptome and over 100 surface proteins in thymocytes from wild-type and lineage-restricted mice. We jointly analyze the paired measurements to build a comprehensive timeline of RNA and protein expression in each lineage, supporting a sequential model of lineage determination in which both lineages go through an initial phase of CD4 lineage audition, which is followed by divergence and specification of CD8 lineage cells. We identify early differences implicating T cell receptor signaling via calcineurin-NFAT in driving CD4 lineage commitment. Pharmacological inhibition validates the requirement of calcineurin- NFAT for CD4, but not CD8, lineage development, providing insight into the CD4/CD8 commitment mechanism.Competing Interest StatementT.H. is an employee of BioLegend Inc. The other authors declare no competing interests.