Affiliation:
1. Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri, USA
Abstract
ABSTRACT
Apicomplexan parasites actively invade host cells using a mechanism predicted to be powered by a parasite actin-dependent myosin motor. In the model apicomplexan
Toxoplasma gondii
, inducible knockout of the actin gene,
ACT1
, was recently demonstrated to limit but not completely abolish invasion. This observation has led to the provocative suggestion that
T. gondii
possesses alternative, ACT1-independent invasion pathways. Here, we dissected the residual invasive ability of Δ
act1
parasites. Surprisingly, we were able to detect residual ACT1 protein in inducible Δ
act1
parasites as long as 5 days after
ACT1
deletion. We further found that the longer Δ
act1
parasites were propagated after
ACT1
deletion, the more severe an invasion defect was observed. Both findings are consistent with the quantity of residual ACT1 retained in Δ
act1
parasites being responsible for their invasive ability. Furthermore, invasion by the Δ
act1
parasites was also sensitive to the actin polymerization inhibitor cytochalasin D. Finally, there was no clear defect in attachment to host cells or moving junction formation by Δ
act1
parasites. However, Δ
act1
parasites often exhibited delayed entry into host cells, suggesting a defect specific to the penetration stage of invasion. Overall, our results support a model where residual ACT1 protein retained in inducible Δ
act1
parasites facilitates their limited invasive ability and confirm that parasite actin is essential for efficient penetration into host cells during invasion.
IMPORTANCE
The prevailing model for apicomplexan invasion has recently been suggested to require major revision, based on studies where core components of the invasion machinery were genetically disrupted using a Cre-Lox-based inducible knockout system. For the myosin component of the motor thought to power invasion, an alternative parasite myosin was recently demonstrated to functionally compensate for loss of the primary myosin involved in invasion. Here, we highlight a second mechanism that can account for the surprising ability of parasites to invade after genetic disruption of core invasion machinery. Specifically, residual actin protein present in inducible knockout parasites appears able to support their limited invasion of host cells. Our results have important implications for the interpretation of the apicomplexan invasion model and also highlight significant considerations when analyzing the phenotypes of inducible knockout parasites generated using Cre-Lox technology.
Publisher
American Society for Microbiology
Cited by
41 articles.
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