Unraveling regulatory networks in biological nutrient removal (BNR) microbiomes

University Of Wisconsin-Madison, Madison WI

Investigators

Abstract

A large fraction of the energy cost in operating a wastewater treatment plant is supplying oxygen to the microorganisms performing the treatment process. These costs would decrease if treatment plants could be operated under low-oxygen conditions. It remains unknown how microorganisms adapt to low-oxygen conditions and whether treatment plants can be effectively operated with minimal oxygen aeration. This project will reveal how key microorganisms in wastewater treatment systems regulate their internal gene networks in response to plant operations with minimal aeration. Other impacts of the project include educational activities related to mentoring high school, undergraduate, and graduate students, and advancing the understanding of gene regulation in natural and engineered environments. If successful, this project will provide transformative information that can help protect the Nation's water security through advanced treatment processes to remove both reactive nitrogen and phosphorus. This proposal seeks to develop and test a framework to study transcriptional regulation in complex microbiomes. Specifically, the framework will be used to investigate transcriptional regulatory networks (TRNs) in an ammonia-oxidizing bacterium and a polyphosphate accumulating organism identified as major contributors to a biological nutrient removal (BNR) microbiome. This microbiome is maintained in a high-rate BNR bioreactor operated using cycles of anaerobic and micro-aerobic conditions. The proposed framework involves: (i) the use of metagenomic data to assemble genomes of key organisms; time-series metatranscriptomic data to identify gene expression patterns in the organisms of interest; (ii) chromatin immunoprecipitation (ChIP) followed by genome-scale DNA sequencing (ChIP-seq) to identify genes directly targeted by a specific transcription factor (TF); and (iii) an in silico phylogenomic approach to predict sets of co-regulated genes in assembled genomes. The project consists of four Tasks. Task 1 focuses on improving genome assembly by using novel tools, obtaining long DNA sequence reads with an emerging sequencing technology, and creating enrichments of the organisms of interest. Task 2 relates to the use ChIP-seq to experimentally identify genes in Nitrosomonas strain UW-LDO-1 targeted by ANR. Task 3 will use ChIP-seq to target RegA, ANR, and DNR - three TFs hypothesized to control aerobic and anoxic respiration in Accumulibacter strain UW-TNR-IC. Task 4 consists of an in silico reconstruction of large-scale TRNs of Nitrosomonas UW-LDO-1 and Accumulibacter UW-TNR-IC. The effective use of this integrative systems biology methodology is transformational because it is the first attempt at assembling a TRN in uncultured microbes using a combination of experimental and computational approaches. This project will provide the first analysis of how respiration is regulated at the transcription level in two key organisms. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.