Expression of the pro-apoptotic proteins Bak and Bax during in-vitro infection of murine cells with Toxoplasma gondii Tachyzoites

Parasite survival depends greatly on how well they can exploit their host. Toxplasma gondii is no exception to this and its exploitation game starts very early in its host-invasion process. More specifically, right after the parasite infects the cell. T. gondii can not only manipulate the arrangement of certain organeles, but it is also can regulate the cell apoptotic mechanisms by delaying them. With this, the parasite can comfortably replicate within the cell and leave it on its own terms. The understanding of how T. gondii delays apoptosis has not been completly elucidated. The objective of this study is to evaluate the expression of Bak and Bax: two pro-apoptotic proteins from the BCL-2 family in murine neurons. These proteins normally work together to produce permeabilization of the mitochondrial outer membrane (MOMP) For this, we used two different T. gondii strains: RH (Type I) and Pru (Type II). We separately infected confluent neuroblastoma-derived cell (cell line Na 42/13) monolayers with each parasite strain and 2 infection groups: MOI=1 and MOI =2 for 3, 24, and 24 hours (h). A negative control of non-infected na 42/13 monolayers was added to the study. Afterwards, we purified RNA from the cells with a commercial kit and generated a complementary DNA strain through a reverese transcriptase-regulared process. Afterwards, we amplify the mRNA region of both BAK and Bax using quantitative PCR.At 3h post infection (hpi), downregulation of Bax was already shown in all infection groups. The major difference was shown at 24 hpi with Bax expression levels significantly low in both RH strain infection groups. Cells infected with T. gondii Pru strain show Bax expression levels mildly higher than the negative control at a MOI =1 and mildly lower than the negative control at a MOI = 2. Bak levels were always higher than Bax at 3hpi and 24hpi. At 48hpi all expression levels of both Bak and Bax were lower than the negative control.

It is common to use Bak/Bax as a unit in the apoptotic reaction. However, in the case of neurons, it is considered that Bax is the main excecutor of MOMP (toghether with other apoptotic proteins). In this study we demostrate that Toxoplasma specifically targets Bax downregulation, particularly at 24 hpi and more importantly, such singling out do not seem to be the same in all strains. How well T. gondii can play the apoptotic game would need to be further investigated.