Zfp281 and Zfp148 control CD4 T cell thymic development and T2 functions.

How CD4 lineage gene expression is initiated in differentiating thymocytes remains poorly understood. Here, we show that the paralog transcription factors Zfp281 and Zfp148 control both this process and cytokine expression by T helper cell type 2 (T2) effector cells. Genetic, single-cell, and spatial transcriptomic analyses showed that these factors promote the intrathymic CD4 T cell differentiation of class II major histocompatibility complex (MHC II)-restricted thymocytes, including expression of the CD4 lineage-committing factor Thpok.

Generation of Retrogenic Mice to Investigate T Cell Development.

T cells develop in the thymus from bone marrow precursors, and genetic manipulation is an indispensable tool to explore their development in vivo. Retroviral transduction of T cell precursors in the bone marrow can be used to specifically eliminate or enforce gene expression. Here, we describe a fast and efficient method to ectopically express a gene in T cell precursors through retroviral transduction and transplant into recipient mice, which will enable laboratories to evaluate gene function in T cell development in vivo.

Generation of Bone Marrow Chimeras.

Bone marrow chimeras are widely used in immunological studies, to dissect the contributions of hematopoietic and non-hematopoietic cells in immune cell development or functions, to quantify the impact of a given mutation, or in preclinical studies for hematopoietic stem cell transplantation. Here we describe a set of procedures for the generation of bone marrow chimeras.

Purification of Thymocyte and T Cell Subsets.

Many analytical or cell culture procedures require homogeneous starting cell populations that cannot be obtained directly from organ dissection. Here, we describe two enrichment procedures to achieve this goal and discuss their respective advantages in specific experimental contexts.