Intracellular competition shapes plasmid population dynamics
Abstract
Conflicts between levels of biological organization are central to evolution, from populations of multicellular organisms to selfish genetic elements. Plasmids—extrachromosomal, self-replicating genetic elements that face selective pressures on their hosts, but also compete within the cell for replication—are an ideal system for studying multilevel selection. While theory indicates that within-cell selection matters for plasmid evolution, experimental measurement of these dynamics has remained elusive. Here we measure the within-cell fitness of competing plasmids and characterize drift and selective dynamics. We achieve this by the controlled splitting of synthetic plasmid dimers, creating balanced competition experiments. We find that incompatible plasmids co-occur for longer than expected due to methylation-based plasmid eclipsing. Moreover, low-transcription plasmids display a within-cell advantage over their competition and fix preferentially. Critically, fixation depends non-trivially on gene dominance. Taken together, our results show that plasmid evolution is driven by dynamics at two levels, with a transient—but critical—contribution of within-cell fitness.
Dataset DOI: 10.5061/dryad.05qfttffx
Description of the data and file structure
These files contain macroscopic images used to investigate within- and between-cell plasmid fitness components
Files and variables
File: data.zip
General description: Each folder contains macroscopic photos of petri dishes with bacterial colonies relative to the described experiments. Each Petri dish was photographed in three different channels: brightfield, red fluorescence, and green fluorescence. Each file name consists of a prefix that uniquely identifies a petri dish, followed by a channel identifier ("BF" for brightfield, "mCher" for red fluorescence, and "GFP" for green fluorescence), and finally the file extension ".jpg". For instance, the photo named 20230407_153731_AC5_BF.jpg refers to the brightfield photo of the Petri dish identified by the string "20230407_153731_AC5_". Note that, for analyses, only fluorescence images were used, but brightfield images are provided so that users know what the samples looked like to the naked eye.
Additionally, the string that identifies each Petri is composed of two parts, a numerical code that serves only to distinguish different replicates, and a alphanumerical code that carries information on the conditions of the experiment. These alphanumerical codes are specific to each of the folders. Additional information is given below for each folder.
ProC.Scar_ProC.Water: This folder contains photos of growth experiments in which dimers were composed of quasi-neutral plasmids carrying the same promoter ProC. The code CC indicates that expression of mScarlet-I is under ProC and the expression of mWatermelon is under ProC.
ProA.Scar_ProC.Water: This folder contains photos of growth experiments in which dimers were composed of plasmids carrying either the strong promoter ProC or the weak promoter ProA. The code AC indicates that expression of mScarlet-I is under ProA and the expression of mWatermelon is under ProC.
homoplasmic_cell_mix: This folder contains photos of growth experiments where the innocullum was a mix of cells carrying a single monomeric plasmid type. Once again, the code CC indicates that expression of mScarlet-I is under ProC and the expression of mWatermelon is under ProC.
DfrA_dimers: This folder contains photos of growth experiments with cells carrying dimers composed of a plasmid encoding mScarlet-I under the weak promoter ProA, and another plasmid encoding mWatermelon plus dfrA under the strong promoter ProC. Additionally dfrA could be under a strong or a weak RBS. file names describe the concentration of trimethoprim used as well as the strength of the RBS ("RBS30" for weak and "RBS1000" for strong).
DAM_neg_dfrA_dimers: Same as the above but only for strong RBS, In this folder, all plasmids had the methylation sites knocked out from their origins.
invasion: This folder contains photos of growth experiments in which cells initially contained a resident plasmid but also a chromosomally integrated plasmid that was released upon induction. As in the previous folder, file names carry information on the Trimethoprim concentration. Additionally, now plasmids can either be the resident plasmid (indicated by "pls") or the chromosomally integrated plasmid (indicated by "chr"). Here, the watermelon+dfrA plasmids are identified by the numbers "30" for the weak RBS and "1000" for the strong RBS. The plasmid encoding mScarlet-I under the weak promoter ProA was denoted by AScar. For example, the Petri dish identified by the string "20240214_223547_TMP0_chrAScar_pls30_" had no added trimethoprim ("TMP0"), had a chromosomally integrated plasmid encoding mScarlet-I under a weak promoter proA ("chrAScar"), and a resident plasmid encoding dfrA under a weak RBS ("pls30").
Code/software
Images can be viewed in any standard image viewer