Interaction between autophagic vesicles and the Coxiella-containing vacuole requires CLTC (clathrin heavy chain)

Abstract

Coxiella burnetii is an intracellular bacterial pathogen which causes Q fever, a human infection with the ability to cause chronic disease with potentially life-threatening outcomes. In humans, Coxiella infects alveolar macrophages where it replicates to high numbers in a unique, pathogen-directed lysosome-derived vacuole. This compartment, termed the Coxiella-containing vacuole (CCV), has a low internal pH and contains markers both of lysosomes and autophagosomes. The CCV membrane is also enriched with CLTC (clathrin heavy chain) and this contributes to the success of the CCV. Here, we describe a role for CLTC, a scaffolding protein of clathrin-coated vesicles, in facilitating the fusion of autophagosomes with the CCV. During gene silencing of CLTC, CCVs are unable to fuse with each other, a phenotype also seen when silencing genes involved in macroautophagy/autophagy. MAP1LC3B/LC3B, which is normally observed inside the CCV, is excluded from CCVs in the absence of CLTC. Additionally, this study demonstrates that autophagosome fusion contributes to CCV size as cell starvation and subsequent autophagy induction leads to further CCV expansion. This is CLTC dependent, as the absence of CLTC renders autophagosomes no longer able to contribute to the expansion of the CCV. This investigation provides a functional link between CLTC and autophagy in the context of Coxiella infection and highlights the CCV as an important tool to explore the interactions between these vesicular trafficking pathways.

Coxiella burnetii Type 4B Secretion System-dependent manipulation of endolysosomal maturation is required for bacterial growth

“…During autophagy, a double-membrane phagophore surrounds and sequesters the cargo in autophagosomes, which then fuse with lysosomes to form autolysosomes [48]. The CCV interacts with autophagosomes as early as 18 hours post infection, acquiring the autophagic proteins LC3 and p62 (sequestosome-1) [7,8,49,50]. Additionally, inducing autophagy by LC3 overexpression improves C. burnetii growth in Chinese Hamster Ovary (CHO) cells [51].…”

Taming the Triskelion: Bacterial Manipulation of Clathrin

“…Clathrin is not associated with lysosomes under steady-state conditions; however, CHC does colocalize with LAMP1positive lysosomes upon autophagy induction (90). This work sparked our recent research which examined the role of CHC in the context of autophagy during C. burnetii infection (236). We demonstrated that CHC is required for fusion of autophagosomes with the CCV.…”

From neglected to dissected: How technological advances are leading the way to the study ofCoxiella burnetiipathogenesis

“…The Newton laboratory has further characterised the role of this effector protein in the development of CCVs and intracellular replication, showing that Cig57 subverts clathrin‐mediated traffic by interacting with FCHO2, an accessory protein of clathrin‐coated pits (Latomanski et al, ) (Figure ). More recently, CTLC (clathrin heavy chain) has been observed at CCVs, where it plays an essential role in vacuole expansion (Latomanski and Newton, ). Importantly, CTLC recruitment to the CCV is related to autophagy, and conversely, the fusion of autophagosomes with CCVs is dependent on CTLC (Latomanski and Newton, ).…”

“…More recently, CTLC (clathrin heavy chain) has been observed at CCVs, where it plays an essential role in vacuole expansion (Latomanski and Newton, ). Importantly, CTLC recruitment to the CCV is related to autophagy, and conversely, the fusion of autophagosomes with CCVs is dependent on CTLC (Latomanski and Newton, ). The Roy visual screen also highlighted a multivacuolar phenotype associated with transposon insertions in the gene CBU0021 ( cig2 ), identical to that previously observed following the knockdown of autophagy‐related genes (McDonough et al, ), suggesting a functional link between a bacterial effector protein and the manipulation of a specific host cell function (Figure O).…”

“…More recently, CTLC (clathrin heavy chain) has been observed at CCVs, where it plays an essential role in vacuole expansion (Latomanski and Newton, 2018). Importantly, CTLC recruitment to the CCV is related to autophagy, and conversely, the fusion of autophagosomes with CCVs is dependent on CTLC (Latomanski and Newton, 2018).…”