Determination of the gene regulatory network of a genome-reduced bacterium highlights alternative regulation independent of transcription factors

Open Access

Abstract

Here, we determined the relative importance of different transcriptional mechanisms in the genome-reduced bacterium Mycoplasma pneumoniae, by employing an array of experimental techniques under multiple genetic and environmental perturbations. Of the 143 genes tested (21% of the bacterium's annotated proteins), only 55% showed an altered phenotype, highlighting the robustness of biological systems. We identified nine transcription factors (TFs) and their targets, representing 43% of the genome, and 16 regulators that indirectly affect transcription. Only 20% of transcriptional regulation is mediated by canonical TFs when responding to perturbations. Using a Random Forest, we quantified the non-redundant contribution of different mechanisms such as supercoiling, metabolic control, RNA degradation, and chromosome topology to transcriptional changes. Model-predicted gene changes correlate well with experimental data in 95% of the tested perturbations, explaining up to 70% of the total variance when also considering noise. This analysis highlights the importance of considering non-TF-mediated regulation when engineering bacteria. ; The project was supported by funds from the Fundación Marcelino Botín and the Spanish Ministerio de Economía y Competitividad (BIO2007-61762). This project was financed by Instituto de Salud Carlos III and co-financed by Federación Española de Enfermedades Raras under grant agreement PI10/01702 and the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program, under grant agreements no 634942 (MycoSynVac) and 670216 (MYCOCHASSIS). We acknowledge support from the Spanish Ministry of Economy and Competitiveness to the EMBL partnership, Centro de Excelencia Severo Ochoa.