Biology subjects

Penewit, K. M.

Publications and source records attributed to Penewit, K. M..

3 recordsLinked to original sources

bioRxiv2023

Discovery of a unique pathway for glutathione utilization in Francisella

Glutathione (GSH) is an abundant metabolite that can act as a signal, a nutrient source, or serve in a redox capacity for intracellular pathogens. For Francisella, GSH is thought to be a critical in vivo source of cysteine; however, the cellular pathways permitting GSH utilization by Francisella differ between strains and have remained poorly understood. Using genetic screening, we discovered a unique pathway for GSH utilization in Francisella spp. Whereas prior work suggested GSH catabolism initiates in the periplasm, the pathway we define consists of a major facilitator superfamily member that transports intact GSH and a previously unrecognized bacterial cytoplasmic enzyme that catalyzes the first step of GSH degradation. Interestingly, we find that the transporter gene for this pathway is pseudogenized in pathogenic Francisella, explaining phenotypic discrepancies in GSH utilization among Francisella spp. and revealing a critical role for GSH in the environmental niche of these bacteria.

microbiology↗

bioRxiv2021

Coordinately regulated interbacterial antagonism defense pathways constitute a bacterial innate immune system

Bacterial survival is fraught with antagonism, including that deriving from viruses and competing bacterial cells1-3 4. It is now appreciated that bacteria mount complex antiviral responses; however, whether a coordinated defense against bacterial threats is undertaken is not well understood. Previously we showed that Pseudomonas aeruginosa possess a danger sensing pathway that is a critical fitness determinant during competition against other bacteria5, 6. Here, we conducted genome-wide screens in P. aeruginosa that reveal three conserved and widespread interbacterial antagonism resistance clusters (arc1-3). We find that although arc1-3 are coordinately activated by the Gac/Rsm danger sensing system, they function independently and provide idiosyncratic defense capabilities, distinguishing them from general stress response pathways. Our findings demonstrate that Arc3 family proteins provide specific protection against phospholipase toxins by preventing the accumulation of lysophospholipids in a manner distinct from previously characterized membrane repair systems. These findings liken the response of P. aeruginosa to bacterial threats to that of eukaryotic innate immunity, wherein threat detection leads to the activation of specialized defense systems.

microbiology↗

bioRxiv2021

Genome-wide protein-DNA interaction site mapping using a double strand DNA-specific cytosine deaminase

DNA-protein interactions (DPIs) are central to such fundamental cellular processes as transcription and chromosome maintenance and organization. The spatiotemporal dynamics of these interactions dictate their functional consequences; therefore, there is great interest in facile methods for defining the sites of DPI within cells. Here, we present a general method for mapping DPI sites in vivo using the double stranded DNA-specific cytosine deaminase toxin DddA. Our approach, which we term DddA-sequencing (3D-seq), entails generating a translational fusion of DddA to a DNA binding protein of interest, inactivating uracil DNA glycosylase, modulating DddA activity via its natural inhibitor protein, and DNA sequencing for genome-wide DPI detection. We successfully applied this method to three Pseudomonas aeruginosa transcription factors that represent divergent protein families and bind variable numbers of chromosomal locations. 3D-seq offers several advantages over existing technologies including ease of implementation and the possibility to measure DPIs at single-cell resolution.

genetics↗