The GTPase TcRjl of the human pathogen Trypanosoma cruzi is involved in the cell growth and differentiation

Abstract

The protozoan parasite Trypanosoma cruzi, the etiological agent of Chagas Disease, undergoes through a complex life cycle where rounds of cell division and differentiation occur initially in the gut of triatominae vectors and, after transmission, inside of infected cells in vertebrate hosts. Members of the Ras superfamily of GTPases are molecular switches which play pivotal regulatory functions in cell growth and differentiation. We have previously described a novel GTPase in T. cruzi, TcRjl, which belongs to the RJL family of Ras-related GTP binding proteins. Here we show that most of TcRjl protein is found bound to GTP nucleotides and may be locked in this stage. In addition, we show that TcRjl is located close to the kinetoplast, in a region corresponding possibly to flagellar pocket of the parasite and the expression of a dominant-negative TcRjl construct (TcRjlS37N) displays a significative growth phenotype in reduced serum medium. Remarkably, overexpression of TcRjl inhibits differentiation of epimastigotes to trypomastigote forms and promotes the accumulation of intermediate differentiation stages. Our data suggest that TcRjl might play a role in the control of the parasite growth and differentiation.

Functional studies of TcRjl, a novel GTPase of Trypanosoma cruzi , reveals phenotypes related with MAPK activation during parasite differentiation and after heterologous expression in Drosophila model system

“…1B and C). Thus, the in vivo data corroborated ours previous results that revealed a remarkable phenotype consisting of the accumulation of intermediate forms of the parasites overexpressing TcRjl and a strong reduction of differentiated metacyclic trypomastigotes, during in vitro metacyclogenesis [18]. Attempts to generate TcRjl negative mutants were unsuccessful, probably due to the vital role of TcRjl in parasite survival.…”

“…Trypanosoma cruzi epimastigotes, Dm28-GFP or Dm28-GFP-TcRjl lineages, were maintained at 28 C in LIT medium (Liver Infusion Tryptose) [25] supplemented with 10% heat-inactivated fetal calf serum (FCS, Gibco) and 0.025 mg/mL hemin (SigmaeAldrich), supplemented with 300 mg/mL of G418 (SigmaeAldrich) [18].…”

“…The Rjl genes are present in many protist lineages, encoding a core GTPase domain, without predictable membrane anchoring signal, while Rbj gene products are found in the choanoflagellata/metazoan lineage as chimaeras of a conventional Rjl protein fused to a "J" domain [16,17]. We have previously shown the involvement of TcRjl protein in cell growth and in vitro metacyclogenesis abrogating differentiation of metacyclic trypomastigotes [18].…”

The roles of evolutionarily conserved functional modules in cilia-related trafficking

“…For example, although not directly linked to trafficking, the DnaJ-domain-containing small GTPase RJL displays a phylogenetic profile strongly suggestive of being motile-cilium-associated 59 . Indeed, the Trypanosoma orthologue localizes near the ciliary base 60 .…”

Golgi UDP-GlcNAc:Polypeptide O -α- N -Acetyl- d -Glucosaminyltransferase 2 (TcOGNT2) Regulates Trypomastigote Production and Function in Trypanosoma cruzi

“…S5), whose inhibition can also inhibit metacyclogenesis. The inhibition of metacyclogenesis by TcOGNT2 overexpression was far stronger than in the case of cruzipain inhibition by chagasin overexpression (75), and it did not involve intermediary forms or death as observed with overexpression of the Ras-related GTP-binding protein TcRjl (83). The reduced Epimastigotes derived from TCT forms were transferred to TAU-P and incubated at 28°C to induce metacyclogenesis.…”