Rapid in vivo analysis of mutant forms of the LAT adaptor using Pax5-Lat double-deficient pro-B cells
✍ Scribed by Laurence Ardouin; Antonius G. Rolink; Anne-Marie Mura; Julien Gommeaux; Fritz Melchers; Meinrad Busslinger; Marie Malissen; Bernard Malissen
- Publisher
- John Wiley and Sons
- Year
- 2005
- Tongue
- English
- Weight
- 472 KB
- Volume
- 35
- Category
- Article
- ISSN
- 0014-2980
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✦ Synopsis
Abstract
Following injection into recombinase‐activating gene‐deficient (Rag1^–/–^) mice, pro‐B cells lacking the Pax5 transcription factor (Pax5^–/–^) develop into most major hematopoietic lineages, with the notable exception of B cells. We assessed whether Pax5^–/–^ pro‐B cells that were also rendered deficient for the linker for activation of T cells (LAT), an adaptor essential for T cell receptor signaling, can be used for the rapid in vivo analysis of mutant forms of LAT. We showed that Pax5^–/–^ Lat^–/–^ pro‐B cell lines can be infected with recombinant retroviruses expressing a LAT cDNA and sorted for the expression of LAT. When injected into Rag1^–/–^ mice, they restore normal intrathymic T cell development and give rise to functional peripheral T cells. Considering that the handling of Pax5^–/–^ pro‐B cell lines is easier than that of bone marrow hematopoietic precursors, we used them for the rapid functional analysis of a novel Lat allelic series. When compared to knock‐in and transgenic approaches, a major advantage of our Pax5^–/–^ pro‐B cell‐based experimental approach consists in the production of mice bearing a given mutation within 2–3 months. Therefore, it constitutes a powerful first‐line screen for mutations worth fastidious knock‐in approaches.