title: Diverse mechanisms of metaeffector activity in an intracellular bacterial pathogen, Legionella pneumophila.
abstract: Pathogens deliver complex arsenals of translocated effector proteins to host cells during infection, but the extent to which these proteins are regulated once inside the eukaryotic cell remains poorly defined. Among all bacterial pathogens, Legionella pneumophila maintains the largest known set of translocated substrates, delivering over 300 proteins to the host cell via its Type IVB, Icm/Dot translocation system. Backed by a few notable examples of effector-effector regulation in L. pneumophila, we sought to define the extent of this phenomenon through a systematic analysis of effector-effector functional interaction. We used Saccharomyces cerevisiae, an established proxy for the eukaryotic host, to query > 108,000 pairwise genetic interactions between two compatible expression libraries of ~330 L. pneumophila-translocated substrates. While capturing all known examples of effector-effector suppression, we identify fourteen novel translocated substrates that suppress the activity of other bacterial effectors and one pair with synergistic activities. In at least nine instances, this regulation is direct-a hallmark of an emerging class of proteins called metaeffectors, or "effectors of effectors". Through detailed structural and functional analysis, we show that metaeffector activity derives from a diverse range of mechanisms, shapes evolution, and can be used to reveal important aspects of each cognate effector's function. Metaeffectors, along with other, indirect, forms of effector-effector modulation, may be a common feature of many intracellular pathogens-with unrealized potential to inform our understanding of how pathogens regulate their interactions with the host cell.
authors.affiliation: Department of Biochemistry, University of Toronto, Toronto, ON, Canada.
authors.forename: Malene L
authors.initials: ML
authors.lastname: Urbanus
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, ON, Canada.
authors.forename: Andrew T
authors.identifier: http://orcid.org/0000-0002-1194-3712
authors.initials: AT
authors.lastname: Quaile
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, ON, Canada.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Stogios
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, ON, Canada.
authors.forename: Mariya
authors.initials: M
authors.lastname: Morar
authors.affiliation: Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada.
authors.forename: Chitong
authors.initials: C
authors.lastname: Rao
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, ON, Canada.
authors.forename: Rosa
authors.initials: R
authors.lastname: Di Leo
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, ON, Canada.
authors.forename: Elena
authors.initials: E
authors.lastname: Evdokimova
authors.affiliation: Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON, Canada.
authors.forename: Mandy
authors.initials: M
authors.lastname: Lam
authors.affiliation: Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada.
authors.forename: Christina
authors.initials: C
authors.lastname: Oatway
authors.affiliation: Biosciences Division, Argonne National Laboratory, Structural Biology Center, Lemont, IL, USA.|Midwest Center for Structural Genomics, Lemont IL, USA.
authors.forename: Marianne E
authors.initials: ME
authors.lastname: Cuff
authors.affiliation: Biosciences Division, Argonne National Laboratory, Structural Biology Center, Lemont, IL, USA.|Midwest Center for Structural Genomics, Lemont IL, USA.
authors.forename: Jerzy
authors.initials: J
authors.lastname: Osipiuk
authors.affiliation: Biosciences Division, Argonne National Laboratory, Structural Biology Center, Lemont, IL, USA.|Midwest Center for Structural Genomics, Lemont IL, USA.
authors.forename: Karolina
authors.initials: K
authors.lastname: Michalska
authors.affiliation: Biosciences Division, Argonne National Laboratory, Structural Biology Center, Lemont, IL, USA.|Midwest Center for Structural Genomics, Lemont IL, USA.
authors.forename: Boguslaw P
authors.initials: BP
authors.lastname: Nocek
authors.affiliation: Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada.|Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON, Canada.
authors.forename: Mikko
authors.identifier: http://orcid.org/0000-0003-3811-1761
authors.initials: M
authors.lastname: Taipale
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, ON, Canada alexei.savchenko@utoronto.ca alex.ensminger@utoronto.ca.|Midwest Center for Structural Genomics, Lemont IL, USA.
authors.forename: Alexei
authors.initials: A
authors.lastname: Savchenko
authors.affiliation: Department of Biochemistry, University of Toronto, Toronto, ON, Canada alexei.savchenko@utoronto.ca alex.ensminger@utoronto.ca.|Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada.|Public Health Ontario, Toronto, ON, Canada.
authors.forename: Alexander W
authors.identifier: http://orcid.org/0000-0003-0824-3704
authors.initials: AW
authors.lastname: Ensminger
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D054884:Host-Pathogen Interactions; D016952:Legionella pneumophila / Q000378:metabolism / Q000472:pathogenicity*; D008954:Models, Biological; D060066:Protein Interaction Maps; D012441:Saccharomyces cerevisiae / Q000254:growth & development*; D049490:Systems Biology / Q000379:methods
chemical_list: D001426:Bacterial Proteins
keywords: 
  Legionella
  ; effector; genetic interaction; metaeffector; structure‐function