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TransLeish
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Resumo(s)
For the protozoan parasite Leishmania, completion of its life cycle requires sequential adaptation of cellular physiology and nutrient scavenging mechanisms to the different environments of a sand fly alimentary tract and the acidic mammalian host cell phagolysosome. Transmembrane transporters are the gatekeepers of intracellular environments, controlling the flux of solutes and ions across membranes. To discover which transporters are vital for survival as intracellular amastigote forms, we carried out a systematic loss-of-function screen of the L. mexicana transportome. A total of 312 protein components of small molecule carriers, ion channels and pumps were identified and targeted in a CRISPR-Cas9 gene deletion screen in the promastigote form, yielding 188 viable null mutants. Forty transporter deletions caused significant loss of fitness in macrophage and mouse infections. A striking example is the Vacuolar H+ ATPase (V-ATPase), which, unexpectedly, was dispensable for promastigote growth in vitro but essential for survival of the disease-causing amastigotes.
Descrição
Funding Information: EG was supported by a Royal Society University Research Fellowship (UF160661). AAW was the recipient of a Marie Sk\u0142odowska-Curie Individual Fellowship (trans-LEISHion-EU FP7, No. 798736). TB was supported by MRC PhD studentship (15/16_MSD_836338; https://mrc.ukri.org/ ), EMBO Postdoctoral Fellowship (ALTF 727-2021) and Marie Sk\u0142odowska-Curie Actions Postdoctoral Fellowship (101064428 \u2013 LeishMOM). RJW is supported by a Wellcome Trust Henry Dale Fellowship (211075/Z/18/Z). The James and Lillian Martin Centre for Stem Cell Research (SAC) is supported by James Martin 21st Century Research Foundation. This work was supported by a UKRI Medical Research Council grant (MR/V000446/1; This UK funded award is part of the EDCTP2 programme supported by the European Union), the Wellcome Trust (221944/A/20/Z, 200807/Z/16/Z, 104627/Z/14/Z) and the Wellcome Centre for Integrative Parasitology (WCIP) core Wellcome Centre Award (104111/Z/14/Z) and a project grant from the Swiss National Science Foundation. We like to thank Amanda Williams (University of Oxford), Julie Galbraith and Csilla Balazs (University of Glasgow, Polyomics facility), Pamela Nicholson and Daniela Steiner (NGS Facility, University of Bern) for help with Illumina sequencing, William James (James & Lillian Martin Centre, Sir William Dunn School of Pathology, University of Oxford) for supporting the work with iPSC-derived macrophages, Caroline Ricce Espada for generating the Ros3 knockout cell line, Keith Gull (Sir William Dunn School of Pathology, University of Oxford) for access to equipment and all past and current members of EG lab for helpful discussions. Publisher Copyright: © The Author(s) 2024.
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General Chemistry General Biochemistry,Genetics and Molecular Biology General Physics and Astronomy