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Principal Investigator: NICHOLAS P CIANCIOTTO
Organization: NORTHWESTERN UNIVERSITY AT CHICAGO
Fiscal Year: 2024
Award: $189,810
Funding agency: National Institute of Allergy and Infectious Diseases
PROJECT SUMMARY
Gram- Stenotrophomonas maltophilia (Sm) is an emergent, multi-drug resistant pathogen. Sm infection occurs
throughout the body but is most often manifest as pneumonia. Sm is notable in cystic fibrosis (CF) patients,
often heightening risk of lung exacerbations, and its significance is increased further by recent reports of Sm
infecting COVID-19 patients. Despite this clinical importance, knowledge of Sm is still relatively limited.
Recently, we found that Sm encodes a type IV secretion system (T4SS) that promotes, in a contact-dependent
manner, apoptosis of macrophages, while blunting apoptosis in lung epithelial cells. Yet, perhaps even more
significant was our finding that the Sm T4SS mediates, in contact-dependent fashion, killing of E. coli (Ec) and
multiple clinical isolates of Pseudomonas aeruginosa (Pa), including those from CF patients. Thus, we posited
that the antibacterial effect of Sm T4SS impacts human infection, as Sm and Pa are often together in water
systems in hospitals and in various types of infections, especially in the (CF) lungs. Based on the analysis of
mutants and their complements, we identified two putative effectors (TfcA and TfcB) as being required for the
bactericidal activity of the Sm T4SS. A mutant lacking both these proteins was as impaired as a mutant lacking
the T4SS apparatus, indicating that TfcA and TfcB account for (nearly all) the killing effects seen. Using both a
2-hybrid assay that measures binding to the T4SS coupling protein and an interbacterial protein translocation
assay, we determined that TfcA and TfcB are bona fide substrates of the T4SS, a result confirmed by testing
mutants lacking both the T4SS apparatus and the individual effectors. Delivery of cloned TfcA (alone) into the
periplasm resulted in the killing of target bacteria, indicating that this effector is both necessary and sufficient
for bactericidal activity. Bioinformatics suggested that TfcA is representative of a large but uncharacterized
branch of microbial lipases, whereas TfcB appears to signify a large but ill-defined group of lysozyme-like
enzymes. In contrast to the vast data re T4SSs impacting mammalian hosts and the many studies on the
antibacterial role of type VI secretion systems, knowledge of the antibacterial role of T4SSs is very minimal.
Indeed, Sm T4SS is thus far the only antibacterial T4SS reported for a human pathogen, and TfcA and TfcB its
only documented bactericidal effectors. Thus, we propose to i) define the enzyme activities encoded by TfcA
and TfcB, ii) discern if those activities promote killing of Pa and Ec, and iii) explore if Sm T4SS, TfcA, and TfcB
also kill other co-inhabitants of the CF lung, ranging from other emerging Gram- bacteria to classic Gram+
pathogens to fungi. Besides improving our knowledge of Sm, the data obtained will have broad implications for
the roles of other T4SSs and possible new links between T4SS effectors and anti-microbial therapies.
Terms: <3-D structure><3-dimensional structure><3D structure><A fumigatus><A xylosoxidans><A. fumigatus><A. xylosoxidans><Achromobacter xylosoxidans><Anti-Bacterial Agents><Antibiotic Agents><Antibiotic Drugs><Antibiotics><Apoptosis><Apoptosis Pathway><Aspergillus fumigatus><Assay><B cenocepacia><B cepacia><B. cenocepacia><B. cepacia><Bacteria><Binding><Bio-Informatics><Bioassay><Bioinformatics><Biological Assay><Burkholderia cenocepacia><Burkholderia cepacia><C-terminal><CF lung disease><CF patients><COVID infected patient><COVID patient><COVID positive patient><COVID-19 infected patient><COVID-19 patient><COVID-19 positive patient><COVID19 patient><COVID19 positive patient><Cell Body><Cell Death><Cells><Cessation of life><Clinical><Consensus Sequence><Coupling><Data><Death><Development><E coli><E. coli><Enzyme Gene><Enzymes><Epithelial Cells><Escherichia coli><Genes><H influenzae><H. influenzae><Haemophilus influenzae><Hospitals><Human><Hybrids><Hydrogen Oxide><Immunity><Impairment><Individual><Infection><Knowledge><LE-Enzyme><Link><Lipase><Lipids><Lung><Lung Respiratory System><Lysozyme><MDR organism><MDR pathogen><Macrophage><Measures><Mediating><Medical><Membrane><Microbe><Miscellaneous Antibiotic><Modern Man><Molecular Interaction><Mucopeptide Amidohydrolase><Muramidase><Murein><Murein Hydrolase><Mφ><N-Acetylmuramide Glycanhydrolase><N-Acetylmuramoyl-L-alanine Amidase><NIH><National Institutes of Health><P aeruginosa><P cepacia><P. aeruginosa><P. cepacia><Pathogenesis><Peptidoglycan><Peptidoglycan Hydrolase><Peptidoglycan N-acetylmuramoylhydrolase><Peptidoglycan amidohydrolase><Periplasmic Space><Pneumonia><Programmed Cell Death><Protein Secretion><Protein translocation><Proteins><Pseudomonas aeruginosa><Pseudomonas cepacia><Pseudomonas maltophilia><Pseudomonas pyocyanea><Pulmonary Cystic Fibrosis><R-Series Research Projects><R01 Mechanism><R01 Program><Ralstonia><Reporting><Research Grants><Research Project Grants><Research Projects><Resistance><Risk><Role><S aureus><S maltophilia><S. aureus><S. maltophilia><SARS-CoV-2 infected patient><SARS-CoV-2 patient><SARS-CoV-2 positive patient><Staph aureus><Staphylococcus aureus><Stenotrophomonas maltophilia><System><T4SS><Testing><Transmembrane Protein Transport><Triacylglycerol Hydrolase><Triacylglycerol Lipase><Triacylglycerol acylhydrolase><Tributyrinase><Triglyceridase><Triglyceride Lipase><Triolean Hydrolase><Type IV Secretion System><Type IV Secretion System Pathway><United States National Institutes of Health><Water><Work><Xanthomonas maltophilia><anti-bacterial><anti-microbial><antimicrobial><bactericidal><bactericide><coronavirus disease 2019 infected patient><coronavirus disease 2019 patient><coronavirus disease 2019 positive patient><coronavirus disease infected patient><coronavirus disease patient><coronavirus disease positive patient><coronavirus disease-19 patient><coronavirus patient><cystic fibrosis lung><cystic fibrosis lung disease><cystic fibrosis patients><developmental><disease control><disorder control><emerging human pathogen><enzyme activity><fungus><human pathogen><improved><individuals with CF><individuals with cystic fibrosis><membrane structure><microbial><microbial interaction><microorganism interaction><multi-drug resistant organism><multi-drug resistant pathogen><multidrug resistant organism><multidrug resistant pathogen><multiple drug resistant organism><multiple drug resistant pathogen><mutant><necrocytosis><novel><pathogen><patient infected with COVID><patient infected with COVID-19><patient infected with SARS-CoV-2><patient infected with coronavirus disease><patient infected with coronavirus disease 2019><patient infected with severe acute respiratory syndrome coronavirus 2><patient with COVID><patient with COVID-19><patient with COVID19><patient with SARS-CoV-2><patient with coronavirus disease><patient with coronavirus disease 2019><patient with severe acute respiratory distress syndrome coronavirus 2><patients with CF><patients with cystic fibrosis><periplasm><pulmonary><resistant><severe acute respiratory syndrome coronavirus 2 infected patient><severe acute respiratory syndrome coronavirus 2 patient><severe acute respiratory syndrome coronavirus 2 positive patient><social role><three dimensional structure><tool><tributyrase><type 4 secretion system>