DOI: 10.5061/dryad.kj666
title: Data from: Diverse mechanisms of metaeffector activity in an intracellular bacterial pathogen, Legionella pneumophila

description: 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 &gt; 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.
description: Query strain identities
 and corresponding screening set.In order to obtain an
 IDTS-wide genetic interaction map, we modified the automated, high-density
 replica plating approaches previously developed for analyzing S.
 cerevisiae double mutants through Synthetic Genetic Array (SGA) analysis
 (Tong AH, Evangelista M, Parsons AB, Xu H, Bader GD, Page N, Robinson M,
 Raghibizadeh S, Hogue CW, Bussey H, Andrews B, Tyers M, Boone C (2001)
 Systematic genetic analysis with ordered arrays of yeast deletion mutants.
 Science 294: 2364-2368). Instead of looking at double mutants, however, we
 used yeast genetics to systematically assess the effects of Legionella
 effector co-expression on yeast growth. Query strains that express one
 effector were mated to an array of ~330 effectors in groups of ~10 queries
 at a time ("Analysis Set"). The arrays were then imaged using a
 high-resolution camera and the spot sizes were quantified using SGAtools
 (http://sgatools.ccbr.utoronto.ca/) (Wagih O, Usaj M, Baryshnikova A,
 VanderSluis B, Kuzmin E, Costanzo M, Myers CL, Andrews BJ, Boone CM, Parts
 L (2013) SGAtools: One-stop analysis and visualization of array-based
 genetic interaction screens. Nucleic acids research 41: W591-596). Outlier
 spot sizes flagged by the Jackknife filter (JK) in SGAtools were removed
 and the average and standard deviation of the remaining values were
 calculated and normalized to the average empty vector control. This
 spreadsheet lists all query strains and links them to one or more specific
 analysis set.Array_set_annotation.xlsSGA output for analysis sets 1-10.Query strains that express one effector were mated to an array of ~330 effectors in groups of ~10 queries at a time ("Analysis Set"). The arrays were then imaged using a high-resolution camera and the spot sizes were quantified using SGAtools (http://sgatools.ccbr.utoronto.ca/). Outlier spot sizes flagged by the Jackknife filter (JK) in SGAtools were removed and the average and standard deviation of the remaining values were calculated and normalized to the average empty vector control. This .zip archive includes spreadsheets that encompass the raw SGAtools data output from the paper for analysis sets 1-10.UrbanusMSB_SGA.zipUrbanus_SGA_1-10.zipSGA output for analysis sets 11-20.Query strains that express one effector were mated to an array of ~330 effectors in groups of ~10 queries at a time ("Analysis Set"). The arrays were then imaged using a high-resolution camera and the spot sizes were quantified using SGAtools (http://sgatools.ccbr.utoronto.ca/). Outlier spot sizes flagged by the Jackknife filter (JK) in SGAtools were removed and the average and standard deviation of the remaining values were calculated and normalized to the average empty vector control. This .zip archive includes spreadsheets that encompass the raw SGAtools data output from the paper for analysis sets 11-20.Urbanus_SGA_11-20.zipSGA output for analysis sets 21-30.Query strains that express one effector were mated to an array of ~330 effectors in groups of ~10 queries at a time ("Analysis Set"). The arrays were then imaged using a high-resolution camera and the spot sizes were quantified using SGAtools (http://sgatools.ccbr.utoronto.ca/). Outlier spot sizes flagged by the Jackknife filter (JK) in SGAtools were removed and the average and standard deviation of the remaining values were calculated and normalized to the average empty vector control. This .zip archive includes spreadsheets that encompass the raw SGAtools data output from the paper for analysis sets 21-30.Urbanus_SGA_21-30.zipSGA output for analysis sets 31-37.Query strains that express one effector were mated to an array of ~330 effectors in groups of ~10 queries at a time ("Analysis Set"). The arrays were then imaged using a high-resolution camera and the spot sizes were quantified using SGAtools (http://sgatools.ccbr.utoronto.ca/). Outlier spot sizes flagged by the Jackknife filter (JK) in SGAtools were removed and the average and standard deviation of the remaining values were calculated and normalized to the average empty vector control. This .zip archive includes spreadsheets that encompass the raw SGAtools data output from the paper for analysis sets 31-37.Urbanus_SGA_31-37.zip