RT Journal Article SR Electronic T1 Intrabody induced cell death by targeting the T. brucei cytoskeletal protein TbBILBO1 JF bioRxiv FD Cold Spring Harbor Laboratory SP 2021.07.18.452872 DO 10.1101/2021.07.18.452872 A1 Christine E. Broster Reix A1 Miharisoa Rijatiana Ramanantsalama A1 Carmelo Di Primo A1 Laëtitia Minder A1 Mélanie Bonhivers A1 Denis Dacheux A1 Derrick R. Robinson YR 2021 UL http://biorxiv.org/content/early/2021/07/20/2021.07.18.452872.abstract AB Trypanosoma brucei belongs to a genus of protists that cause life-threatening and economically important diseases of human and animal populations in Sub-Saharan Africa. T. brucei cells are covered in surface glycoproteins some of which are used to escape the host immune system. Exo-/endocytotic trafficking of these and other molecules occurs via a single copy organelle called the flagellar pocket (FP). The FP is maintained and enclosed around the flagellum by the flagellar pocket collar (FPC). To date, the most important cytoskeletal component of the FPC is an essential, calcium-binding, polymer-forming protein called TbBILBO1. In searching for novel immune-tools to study this protein, we raised nanobodies against TbBILBO1. Nanobodies (Nb) that were selected according to their binding properties to TbBILBO1, were tested as immunofluorescence tools, and expressed as intrabodies (INb). One of them, Nb48, proved to be the most robust nanobody and intrabody. We further demonstrate that inducible, cytoplasmic expression of INb48 was lethal to these parasites, producing abnormal phenotypes resembling those of TbBILBO1 RNAi knockdown. Our results validate the feasibility of generating functional single-domain antibody derived intrabodies to target trypanosome cytoskeleton proteins.