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

Abstract

29Upon host cell infection, the obligate intracellular bacterium C. burnetii resides and multiplies 30 within the Coxiella-Containing Vacuole (CCV). The nascent CCV progresses through the 31 endosomal maturation pathway into a phagolysosome, acquiring lysosomal markers as well as 32 acidic pH and active proteases and hydrolases. Approximately 24-48 hours post infection, 33 heterotypic fusion between the CCV and host endosomes/lysosomes leads to CCV expansion 34 and subsequent bacterial replication in the mature CCV. Initial CCV acidification is required to 35 activate C. burnetii metabolism and the Type 4B Secretion System (T4BSS), which secretes 36 effector proteins required for CCV maturation. However, we recently found that the mature CCV 37 is less acidic (pH~5.2) than lysosomes (pH~4.8). Further, CCV acidification to pH~4.8 causes C. 38 burnetii lysis, suggesting C. burnetii actively regulates CCV pH. Because heterotypic fusion with 39 host endosomes/lysosomes may influence CCV pH, we investigated endosomal maturation in 40 cells infected with wildtype (WT) or T4BSS mutant (dotA) C. burnetii. We observed significantly 41 fewer LAMP1-positive lysosomes, along with less acidic "mature" endosomes (pH~5.8), in WT-42 infected cells, compared to mock or dotA-infected cells. Further, while endosomes progressively 43 acidified from the periphery (pH~5.5) to the perinuclear area (pH~4.7) in both mock and dotA-44 infected cells, endosomes did not acidify beyond pH~5.2 in WT-infected cells, indicating that the 45 C. burnetii T4BSS inhibits endosomal maturation. Finally, increasing the number of acidic 46 lysosomes by overexpressing the transcription factor EB inhibited C. burnetii growth, indicating 47 lysosomes are detrimental to C. burnetii. Overall, our data suggest that C. burnetii regulates CCV 48 pH, possibly by reducing the number of host lysosomes available for heterotypic fusion. 49 50 51 3 52 Author summary 53 54 The obligate intracellular bacterium Coxiella burnetii causes human Q fever, which manifests as 55 a flu-like illness but can develop into a life-threatening and difficult to treat endocarditis. C. burnetii, 56in contrast to many other intracellular bacteria, thrives within a lysosome-like vacuole in host cells. 57However, we previously found that the C. burnetii vacuole is not as acidic as lysosomes and 58 increased acidification kills the bacteria, suggesting that C. burnetii regulates the pH of its vacuole. 59Here, we discovered that C. burnetii blocks endosomal maturation and acidification during hostresulting in fewer lysosomes in the host cell. Moreover, increasing lysosomes in the 61 host cells blocked C. burnetii growth. Together, our study suggests that C. burnetii regulates 62 vacuole acidity and blocks endosomal acidification in order to produce a permissive intracellular 63 niche. 64 65 79 infection, phagosome expansion, presumably through fusion with the endocytic pathway, gives 80 rise to a large acidic phagolysosomal-like compartment termed the Coxiella-containing vacuole 81 (CCV) [...

Current perspectives on the occurrence of Q fever: highlighting the need for systematic surveillance for a neglected zoonotic disease in Indian subcontinent

“…This marks the vegetative growth leading to the differences in morphological appearances. Meanwhile, the acidic pH maintained within the phagolysosome activates the metabolic enzymes of the pathogen (Samanta et al ., 2019). The pathogen could produce superoxide dismutase that protects from host cells generated superoxide anion and hydrogen peroxide for its survival in phagolysosome (Brennan et al ., 2015).…”