Genome-scale RNA interference profiling of Trypanosoma brucei cell cycle progression defects

• DOI: 10.1038/s41467-022-33109-y
• Publication Date: 2022-09-10T04:03:03Z
• Authors: Catarina A. Marques; Melanie Ridgway; Michele Tinti; Andrew Cassidy; David Horn
• Keywords: 0301 basic medicine; 570; sleeping sickness; Trypanosoma cruzi; Science; Trypanosoma brucei brucei; 610; Mitosis; Article; 03 medical and health sciences; checkpoint; name=General; /dk/atira/pure/subjectarea/asjc/1000; name=General Biochemistry,Genetics and Molecular Biology; Humans; Trypanosoma brucei; mitosis; Leishmania; 2. Zero hunger; /dk/atira/pure/subjectarea/asjc/1300/1300; Genome; Q; Cell Cycle; name=General Physics and Astronomy; RNA Interference; /dk/atira/pure/subjectarea/asjc/3100/3100; kinetoplastid; name=General Chemistry; RIT-seq; /dk/atira/pure/subjectarea/asjc/1600/1600

AbstractTrypanosomatids, which include major pathogens of humans and livestock, are flagellated protozoa for which cell cycle controls and the underlying mechanisms are not completely understood. Here, we describe a genome-wide RNA-interference library screen for cell cycle defects inTrypanosoma brucei. We induced massive parallel knockdown, sorted the perturbed population using high-throughput flow cytometry, deep-sequenced RNAi-targets from each stage and digitally reconstructed cell cycle profiles at a genomic scale; also enabling data visualisation using an online tool (https://tryp-cycle.pages.dev/). Analysis of several hundred genes that impact cell cycle progression reveals >100 flagellar component knockdowns linked to genome endoreduplication, evidence for metabolic control of the G1-S transition, surface antigen regulatory mRNA-binding protein knockdowns linked to G2M accumulation, and a putative nucleoredoxin required for both mitochondrial genome segregation and for mitosis. The outputs provide comprehensive functional genomic evidence for the known and novel machineries, pathways and regulators that coordinate trypanosome cell cycle progression.