Effective RNAi-mediated β2-microglobulin loss of function by transgenesis in Xenopus laevis

To impair MHC class I (class I) function in vivo in the amphibian Xenopus, we developed an effective reverse genetic loss of function approach by combining I-SceI meganuclease-mediated transgenesis with RNAi technology. We generated transgenic outbred X. laevis and isogenetic laevis/gilli cloned lines with stably silenced expression of beta 2-microglobulin (b2m) critical for class I function. Transgenic F1 frogs exhibited decreased surface class I expression on erythrocytes and lymphocytes, decreased frequency of peripheral CD8 T cells and impaired CD8 T cell-mediated skin allograft rejection. Additionally, b2m knockdown increased susceptibility to viral infection of F-0 transgenic larvae. This loss of function strategy offers new avenues for studying ontogeny of immunity and other developmental processes in Xenopus. (C) 2013. Published by The Company of Biologists Ltd.