Acquisition of Host Cytosolic Protein byToxoplasma gondiiBradyzoites

Abstract

Toxoplasma gondii is a protozoan parasite that persists in the central nervous system as intracellular chronic stage bradyzoites that are encapsulated by a thick cyst wall. While the cyst wall separates bradyzoites from the host cytosol, it has been posited that small solutes can traverse the cyst wall to sustain bradyzoites. Recently it was found that host cytosolic macromolecules can cross the parasitophorous vacuole and are ingested and digested by actively replicating acute stage tachyzoites. However, the extent to which bradyzoites have an active ingestion pathway remained unknown. To interrogate this, we modified previously published protocols that look at tachyzoite acquisition and digestion of host proteins by measuring parasite accumulation of a host-expressed reporter protein after impairment of an endolysosomal protease (Cathepsin Protease L, CPL). Using two cystogenic parasite strains (ME49 and Pru), we demonstrate that T. gondii bradyzoites can ingest host-derived cytosolic mCherry. Bradyzoites acquire host mCherry within 4 hours of invasion and post-cyst wall formation. This study provides direct evidence that host macromolecules can be internalized by T. gondii bradyzoites across the cyst wall in infected cells.

Toxoplasma TgATG9 is critical for autophagy and long-term persistence in tissue cysts

“…We propose two possibilities, that bradyzoites uptake host proteins that are trafficked to the VAC for degradation via the endolysosomal system or that, in the absence of TgATG9, bradyzoites have an alternative mechanism of autophagy. The first hypothesis is supported by our recent work, which demonstrated that bradyzoites can ingest and degrade host-derived cytosolic proteins (18). In the second possibility, as discussed further below, we observed the formation of vesicles with two or more membranes in ∆atg9 bradyzoites by TEM microscopy and while we are currently unable to determine the nature of these vesicles, it remains possible that they represent alternative autophagosome-like structures that do not stain well with the cationic amphiphilic dye CytoID.…”

“…However, this also raises the question as to how T. gondii tissue cysts containing bradyzoites are able to persist within the host with potentially limited access to host material for sustenance. Although recent work reported that T. gondii bradyzoites can ingest and digest protein from the cytosol of infected cells in vitro (18), the extent to which this process helps to sustain bradyzoites remains unknown.…”

Emerging Mechanisms of Endocytosis in Toxoplasma gondii

“…Finally, although host cytosol ingestion has been recently reported to also occur in chronic stage Toxoplasma gondii bradyzoites [ 124 ], the precise role of this pathway in bradyzoite biology remains to be determined. Parasites lacking CPL can differentiate into cysts normally, but eventually accumulate undigested autophagosomes and die [ 125 ].…”