Identification of human alpha-enolase as an interaction partner of a Brucella abortus VirB substrate

MARCHESINI, MARÍA INÉS; SUSANA MORRONE SEIJO; FRANCISCO GUAIMAS; COMERCI, DIEGO JOSÉ

Berlin

Congreso; Brucellosis 2014 International Research Conference.; 2014

Brucella abortus, the causative agent of bovine brucellosis, invades and replicates within cells inside a membrane-bound compartment known as the Brucella containing vacuole (BCV). After trafficking along the endocytic and secretory pathways, BCVs mature into endoplasmic reticulum (ER)-derived compartments permissive for bacterial replication. Brucella Type IV Secretion System (VirB) is a major virulence factor essential for the biogenesis of the replicative organelle. VirB system translocates effector proteins thought to modulate host cell signaling pathways to favor intracellular survival and replication. Recently, many VirB substrates have been identified. Uncovering the targets and functions of these translocated effectors is essential to understand the role of VirB in pathogenesis. BPE123 (YP_418361.1) is a B. abortus VirB-translocated effector protein recently identified by our group. It is a hypothetical protein whose function remains unknown. In an attempt to identify host cell proteins interacting with BPE123, a pull-down assay was performed and human alpha-enolasa (ENO1) was identified by LC/MS-MS as a potential interaction partner of BPE123. These results were confirmed by immunoprecipitation assays. Microscopy studies further confirmed BPE123-ENO1 interaction: ENO1 relocalization was detected upon ectopic expression of BPE123 in HeLa cells, where both proteins localized to the ER. Furthermore, during macrophage infection we observed recruitment of ENO1 to the vicinities of BPE123 positive VCBs, indicating that interaction with translocated BPE123 might also be occurring during the intracellular phase of B. abortus. Taken together, these preliminary results suggest a direct interaction between BPE123 and ENO1, a multifunctional protein that has already been shown to be required for Brucella replication in host cells. Further experiments are underway to determine how BPE123-ENO1 interaction modulates the outcome of the infection process.