Replication Protein A Presents Canonical Functions and Is Also Involved in the Differentiation Capacity of Trypanosoma cruzi

Replication protein a (rpa),the major single stranded dna binding protein in eukaryotes,is composed of three subunits and is a fundamental player in dna metabolism,participating in replication,transcription,repair,and the dna damage response. in human pathogenic trypanosomatids,only limited studies have been performed on rpa-1 from leishmania. here,we performed in silico,in vitro and in vivo analysis of trypanosoma cruzi rpa-1 and rpa-2 subunits. although computational analysis suggests similarities in dna binding and ob-fold structures of rpa from t. cruzi compared with mammalian and fungi rpa,the predicted tridimensional structures of t. cruzi rpa-1 and rpa-2 indicated that these molecules present a more flexible tertiary structure,suggesting that t. cruzi rpa could be involved in additional responses. here,we demonstrate experimentally that the t. cruzi rpa complex interacts with dna via rpa-1 and is directly related to canonical functions,such as dna replication and dna damage response. accordingly,a reduction of tcrpa-2 expression by generating heterozygous knockout cells impaired cell growth,slowing down s-phase progression. moreover,heterozygous knockout cells presented a better efficiency in differentiation from epimastigote to metacyclic trypomastigote forms and metacyclic trypomastigote infection. taken together,these findings indicate the involvement of tcrpa in the metacyclogenesis process and suggest that a delay in cell cycle progression could be linked with differentiation in t. cruzi. © 2016 pavani et al.