A Xenopus Laevis Research Resource for Immunobiology

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The goal of this renewal application is to maintain and further develop the world's most comprehensive research resource specializing in the use of the amphibian Xenopus laevis as a multi-faceted experimental platform for biomedical and immunological research and for the benefit of the whole scientific community. Interests and medical relevance of X. laevis are due to the remarkable similarity of its immune system with that of human, the accessibility to experimentation at all developmental stages, as well as the availability of large genetic and genomic resources, invaluable MHC-defined inbred strains, clones and transgenic (Tg) lines of frogs, as well as tools such as cell lines, monoclonal antibodies, MHC tetramers, tagged recombinant reagents and batteries of validated PCR primers for immune-relevant genes. These animals and reagents that are not commercially available need to be preserved, enriched, and made available to the scientific community. As in previous proposals, two major main aims are proposed: (1) Preserving and promoting the X. laevis research resource for immunobiology by keeping on producing, managing and distributing animal stocks and reagents to laboratories in the US and abroad. We will maintain the quality, productivity and welfare of our inbred and Tg animals. We will assist, train and inform scientists, students and educators interested in using X. laevis for research. We will foster the information, accessibility, networking and public awareness of the resource using our web site, and by interacting with other Xenopus resources nationally and internationally. (2) Developing new methodologies and generating new experimental animals and reagents by leveraging X. laevis to advance research in 3 main areas: (1) Genetically modified X. laevis lines of frogs: to improve genome annotation, generate new CRISPR/Cas9-mediated inbred Tg knockout (KO) lines and lines with traceable immune cell types by knocking-in long single stranded donor templates (lssDNA). (2) X. laevis cell culture systems and reagents: to produce transfected and deficient cell lines, recombinant tagged immune reagents and MHC tetramers as well as to develop VSV-lentiviral transducing vectors for in vitro and in vivo studies in Xenopus. (3) X. laevis experimental platforms for infectious disease and developmental immunotoxicology: to develop cost-effective and reliable experimental platforms to study infection and pathogenicity of emerging human non-tuberculosis mycobacterium (NTM) pathogens (e.g., M. abscessus); and assess overlooked risk for immunity and human health of water pollutants with endocrine disruption activity. In addition to maintaining a research resource that is crucial for the Xenopus scientific community, this project will develop new approaches and technologies broadly applicable for gathering innovative insights into tissue and organ physiology, immunology and developmental biology. This will contribute to the efforts of the scientific community assisted by the NIH to establish Xenopus as a relevant model for biomedical research.