High-throughput chemical genomic screens produce informative datasets, providing valuable insights into unknown gene function on a genome-wide level. However, there is currently no comprehensive analytic package publicly available. We developed and benchmarked ChemGAPP to bridge this gap. ChemGAPP allows integration of various steps in a streamlined and user-friendly format, including rigorous quality control measures to curate screening data. ChemGAPP provides three sub-packages for different chemical-genomic screens: ChemGAPP Big for handling large-scale high-throughput screens; ChemGAPP Small, designed for small-scale screen analysis and ChemGAPP GI for genetic interaction screen analysis. ChemGAPP is available at https://github.com/HannahMDoherty/ChemGAPP.

Motivation High-throughput chemical genomic screens produce informative datasets, providing valuable insights into unknown gene function on a genome-wide level. However, there is currently no comprehensive analytic package publicly available. We developed ChemGAPP to bridge this gap. ChemGAPP integrates various steps in a streamlined and user-friendly format, including rigorous quality control measures to curate screening data. Results ChemGAPP provides three sub-packages for different chemical-genomic screens: ChemGAPP Big for large-scale screens; ChemGAPP Small, for small-scale screens and ChemGAPP GI for genetic interaction screens. ChemGAPP Big, tested against the E. coli KEIO collection, revealed reliable fitness scores which displayed biologically relevant phenotypes. ChemGAPP Small, demonstrated significant changes in phenotype in a small-scale screen. ChemGAPP GI was benchmarked against three sets of genes with known epistasis types and successfully reproduced each interaction type. Availability ChemGAPP is available at https://github.com/HannahMDoherty/ChemGAPP, as a standalone Python package as well as Streamlit applications. Supplementary information Supplementary data are available at Bioinformatics online.

Bacterial amidases are essential to split the shared envelope of adjunct daughter cells to allow cell separation. Their activity needs to be precisely controlled to prevent cell lysis. In Escherichia coli, amidase activity is controlled by three regulatory proteins NlpD, EnvC and ActS. However, recent studies linked the outer membrane lipoprotein DolP (formerly YraP) as a potential upstream regulator of NlpD. In this study we explored this link in further detail. To our surprise DolP did not modulate amidase activity in vitro and was unable to interact with NlpD in pull-down and MST (MicroScale Thermophoresis) assays. Next, we excluded the hypothesis that ΔdolP phenocopied ΔnlpD in a range of envelope stresses. However, morphological analysis of double deletion mutants of amidases (AmiA, AmiB AmiC) and amidase regulators with dolP revealed that ΔamiAΔdolP and ΔenvCΔdolP mutants display longer chain length compared to their parental strains indicating a role for DolP in cell division. Overall, we present evidence that DolP does not affect NlpD function in vitro, implying that DolP is not an upstream regulator of NlpD. However, DolP may impact daughter cell separation by interacting directly with AmiA or AmiC, or by a yet undiscovered mechanism.

Biogenesis of the bacterial outer membrane is key to survival and antibiotic resistance. Central to this is the beta-barrel assembly machine (Bam) complex and its associated chaperones, which are responsible for outer membrane protein (OMP) transport and insertion. The Escherichia coli Bam complex consists of two essential subunits, BamA and BamD, and three non-essential lipoproteins, BamB, BamC and BamE. Optimal Bam function is further dependent on the non-essential chaperones DegP, Skp and SurA. Despite intensive study, the specific function of these non-essential Bam-associated proteins remains unknown. Here, we analysed knockout strains for each gene by phenotypic screening, conservation analysis and high-throughput genetics. We reveal that Bam activity is affected by outer membrane lipid composition and that enterobacterial common antigen is essential in the absence of the chaperone SurA. We also show that components of peptidoglycan are conditionally essential with Bam accessory lipoproteins and that DNA replication is perturbed in the absence of BamB. Together, our data indicates potential mechanisms for coordination of OMP biogenesis with processes such as LPS and peptidoglycan biogenesis, and DNA replication.