Proteomic Identification of Coxiella burnetii Effector Proteins Targeted to the Host Cell Mitochondria During Infection

Autor: Chen Ai Khoo, Nichollas E. Scott, Laura F. Fielden, Catherine S Palmer, Diana Stojanovski, Hayley J. Newton
Jazyk: angličtina
Rok vydání: 2020
Předmět:
TIM
translocase of the inner mitochondrial membrane
Proteomics
Bacterial effector protein
Proteome
THP-1 Cells
Mitochondrion
medicine.disease_cause
Biochemistry
Intracellular bacteria
Analytical Chemistry
Organelle purification
Protein targeting
Proteomics screen
T4SS
Type IV Secretion System
0303 health sciences
IMS
intermembrane space
biology
Mce
Mitochondrial Coxiella effector protein
Effector
AGC
automatic gain control
030302 biochemistry & molecular biology
LFQ
label-free quantitation
Cell biology
Mitochondria
T4SS
Coxiella burnetii
TOM
translocase of the outer mitochondrial membrane
PK
Proteinase K
Q Fever
IMM
inner mitochondrial membrane
THP-1 cells
human monocyte derived macrophage cell line
TCA
trichloroacetic acid
03 medical and health sciences
Bacterial Proteins
medicine
protein targeting
Humans
Molecular Biology
030304 developmental biology
Host-pathogen interactions
Intracellular parasite
Research
biology.organism_classification
Host cell mitochondrion
HEK293 Cells
PMSF
phenylmethylsulfonyl fluoride
Label-free quantitative proteomics
HeLa Cells
Zdroj: Molecular & Cellular Proteomics : MCP
ISSN: 1535-9484
1535-9476
Popis: Modulation of the host cell is integral to the survival and replication of microbial pathogens. Several intracellular bacterial pathogens deliver bacterial proteins, termed “effector proteins” into the host cell during infection by sophisticated protein translocation systems, which manipulate cellular processes and functions. The functional contribution of individual effectors is poorly characterized, particularly in intracellular bacterial pathogens with large effector protein repertoires. Technical caveats have limited the capacity to study these proteins during a native infection, with many effector proteins having only been demonstrated to be translocated during over-expression of tagged versions. Here, we developed a novel strategy to examine effector proteins in the context of infection. We coupled a broad, unbiased proteomics-based screen with organelle purification to study the host–pathogen interactions occurring between the host cell mitochondrion and the Gram-negative, Q fever pathogen Coxiella burnetii. We identify four novel mitochondrially-targeted C. burnetii effector proteins, renamed Mitochondrial Coxiella effector protein (Mce) B to E. Examination of the subcellular localization of ectopically expressed proteins confirmed their mitochondrial localization, demonstrating the robustness of our approach. Subsequent biochemical analysis and affinity enrichment proteomics of one of these effector proteins, MceC, revealed the protein localizes to the inner membrane and can interact with components of the mitochondrial quality control machinery. Our study adapts high-sensitivity proteomics to study intracellular host–pathogen interactions, providing a robust strategy to examine the subcellular localization of effector proteins during native infection. This approach could be applied to a range of pathogens and host cell compartments to provide a rich map of effector dynamics throughout infection.
Graphical Abstract
Highlights • Mitochondrial purification and proteomics to study host–pathogen interactions. • Quantitative proteomics reveals Coxiella effector proteins at mitochondria. • Insights into effector protein targeting during native infection. • Further characterization of MceC reveals localization and interaction.
In Brief A broad, unbiased proteomics-based screen with organelle purification to study the host–pathogen interactions occurring between the host cell mitochondrion and the Q fever pathogen Coxiella burnetii. This reveals a subset of Coxiella effector proteins at mitochondria during infection. Our study adapts high-sensitivity proteomics, providing a robust strategy to examine the subcellular localization of effector proteins during native infection.
Databáze: OpenAIRE
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