Reconceptualizing the chlamydial inclusion as a pathogen-specified parasitic organelle: an expanded role for Inc proteins

Abstract

Chlamydia is an obligate intracellular pathogen that develops in the host cell in a vacuole termed the chlamydial inclusion. The prevailing concept of the chlamydial inclusion is of a parasitophorous vacuole. Here, the inclusion is the recipient of one-way host-pathogen interactions thus draining nutrients from the cell and negatively impacting it. While Chlamydia orchestrates some aspects of cell function, recent data indicate host cells remain healthy up until, and even after, chlamydial egress. Thus, while Chlamydia relies on the host cell for necessary metabolites, the overall function of the host cell, during chlamydial growth and development, is not grossly disturbed. This is consistent with the obligate intracellular organism's interest to maintain viability of its host. To this end, Chlamydia expresses inclusion membrane proteins, Incs, which serve as molecular markers for the inclusion membrane. Incs also contribute to the physical structure of the inclusion membrane and facilitate host-pathogen interactions across it. Given the function of Incs and the dynamic interactions that occur at the inclusion membrane, we propose that the inclusion behaves similarly to an organelle-albeit one that benefits the pathogen. We present the hypothesis that the chlamydial inclusion acts as a pathogen-specified parasitic organelle. This representation integrates the inclusion within existing subcellular trafficking pathways to divert a subset of host-derived metabolites thus maintaining host cell homeostasis. We review the known interactions of the chlamydial inclusion with the host cell and discuss the role of Inc proteins in the context of this model and how this perspective can impact the study of these proteins. Lessons learnt from the chlamydial pathogen-specified parasitic organelle can be applied to other intracellular pathogens. This will increase our understanding of how intracellular pathogens engage the host cell to establish their unique developmental niches.

Keywords: Chlamydia; inc protein; inclusion membrane; parasitophorous vacuole; pathogen specified organelle.

Figure 1

Model of the function of the chlamydial inclusion. In (A), there are two representations of the chlamydial inclusion: the classical paradigm of the chlamydial inclusion as a parasitophorous vacuole and our concept of the chlamydial inclusion as a pathogen-specified parasitic organelle, which is more consistent with recent data. In (B), the composition of the inclusion membrane is represented with Inc proteins serving as scaffolds to organize the membrane by creating microdomains to support host-chlamydial interactions and organize the inner leaflet of the chlamydial inclusion membrane. In our model, microdomains are collections of discrete subsets of Inc proteins as described in Mital et al. (2010) and Alzhanov et al. (2009).

Figure 2

Hypothetical model for temporal changes in the inclusion membrane with proposed functions. Boxed and circled proteins are known host-cell derived proteins that localize to the inclusion. Unboxed proteins or letters represent Incs. The only interaction to be validated by multiple biochemical assays demonstrating in-cell interactions is IncD and CERT (Agaisse and Derre, 2014).

Figure 3

Inducible expression of IncA_TM-APEX2. HeLa cells were inoculated with C. trachomatis serovar L2 transformed with pASK-IncA_TM_APEX2-mKate2::L2 to allow for inducible expression of the N-terminal region of IncA encoding the transmembrane domains (TM) fused to APEX2. The plasmid backbone (pASK-GFP/mKate2-L2 plasmid) for this study was generously provided by P. Scott Hefty (Department of Molecular Biosciences, University of Kansas) (Wickstrum et al., 2013). 6 h after infection, IncA_TM-APEX2 expression was induced by treating cultures with 10 ng/ml anhydrotetracycline for an additional 18 h. Monolayers were then treated with or without biotin-phenol, processed for immunofluorescence and visualized with a 60X objective on an Olympus Fluoview 1000 Laser Scanning Confocal Microscope. White arrows depict IncA_TM-APEX2 (red) localized to the IM or fibers extending from the inclusion. In the presence of biotin-phenol the construct is able to biotinylate (green, detected with streptavidin-488) the IM. Scalebar = 5 μm.