The trypanosome Variant Surface Glycoprotein mRNA is stabilized by an essential unconventional RNA-binding protein

• Published in: bioRxiv
• Main Authors: Larissa Melo Do Nascimento, Egler, Franziska, Arnold, Katharina, Papavisiliou, Nina, Clayton, Christine, Erben, Esteban
• Format: Paper
• Language: English
• Published: Cold Spring Harbor Cold Spring Harbor Laboratory Press 08.10.2020
• Subjects: 3' Untranslated regions; Adaptive immunity; African trypanosomiasis; Cell cycle; Cell division; DNA-directed RNA polymerase; Gene expression; Glycoproteins; Initiation complex; Livestock; mRNA processing; mRNA stability; Parasites; Pathogenicity; Poly(A); Polyadenylation; Protein purification; Proteins; RNA polymerase; RNA-binding protein; Transcription; Translation initiation; Ubiquitination; Variant surface glycoprotein
• DOI: 10.1101/2020.10.08.331769

Salivarian trypanosomes cause human sleeping sickness and economically important livestock diseases. The “bloodstream forms”, which replicate extracellularly in the blood and tissue fluids of mammals, are coated by a monolayer of Variant Surface Glycoprotein (VSG). Switching of the expressed VSG gene is central to parasite pathogenicity because it enables the parasites to evade adaptive immunity via antigenic variation. Adequate levels of VSG expression - 10% of total protein and 7% of mRNA - are attained through very active RNA polymerase I transcription, efficient mRNA processing (trans splicing of a capped leader and polyadenylation), and high mRNA stability. We here show how VSG mRNA stability is maintained. Purification of the VSG mRNA with associated proteins specifically selected CFB2, an F-box mRNA-binding protein that lacks known RNA-binding domains. CFB2 binds to a stabilizing complex (MKT1-PBP1-XAC1-LSM12) that recruits poly(A) binding protein and a specialized cap-binding translation initiation complex, EIF4E6-EIF4G5. The interaction of CFB2 with MKT1 is essential for CFB2’s expression-promoting activity, while the F-box auto-regulates CFB2 abundance via interaction with SKP1, a component of the ubiquitination machinery. The results of reporter experiments indicate that CFB2 acts via conserved sequences in the VSG mRNA 3’-untranslated region. Depletion of CFB2 leads to highly specific loss of VSG mRNA. VSG expression is essential not only for antigenic variation but also for trypanosome cell division. Correspondingly, depletion of CFB2 causes cell cycle arrest, dramatic morphological abnormalities and trypanosome death. Competing Interest Statement The authors have declared no competing interest.