Journal
MSYSTEMS
Volume 6, Issue 1, Pages -Publisher
AMER SOC MICROBIOLOGY
DOI: 10.1128/mSystems.00002-21
Keywords
genome-scale metabolic model; C-13 metabolic flux analysis; nonmodel fungus; Neocallimastigomycota; flux balance analysis; Neocallimastix lanati; anaerobes; anaerobic fungi
Categories
Funding
- National Science Foundation (NSF) [MCB-1553721]
- Office of Biological and Environmental Research of the DOE Office of Science [DE-AC02-05CH11231]
- Army Research Office [W911NF-19-1-0010]
- Office of Science of the U.S. Department of Energy [DE-AC02-05CH11231]
- Dow Discovery Fellowship
- University of California, Santa Barbara
- University of California, Office of the President
- National Science Foundation [CNS-1725797]
- California NanoSystems Institute
- Materials Research Science and Engineering Center (MRSEC) at UC Santa Barbara [NSF DMR 1720256]
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This study presents a high-quality genome of the anaerobic fungus Neocallimastix lanati, from which the first genome-scale metabolic model of an anaerobic fungus was constructed. The model accurately predicts experimentally validated substrate utilization requirements and metabolic flux predictions were verified. The model sheds light on key aspects of hydrogenosomal metabolism and provides a platform for quantitatively studying these important but poorly understood early-branching fungi.
Anaerobic gut fungi in the phylum Neocallimastigomycota typically inhabit the digestive tracts of large mammalian herbivores, where they play an integral role in the decomposition of raw lignocellulose into its constitutive sugar monomers. However, quantitative tools to study their physiology are lacking, partially due to their complex and unresolved metabolism that includes the largely uncharacterized fungal hydrogenosome. Modern omits approaches combined with metabolic modeling can be used to establish an understanding of gut fungal metabolism and develop targeted engineering strategies to harness their degradation capabilities for lignocellulosic bioprocessing. Here, we introduce a high-quality genome of the anaerobic fungus Neocallimastix lanati from which we constructed the first genome-scale metabolic model of an anaerobic fungus. Relative to its size (200 Mbp, sequenced at 62 x depth), it is the least fragmented publicly available gut fungal genome to date. Of the 1,788 lignocellulolytic enzymes annotated in the genome, 585 are associated with the fungal cellulosome, underscoring the powerful lignocellulolytic potential of N. lanati. The genome-scale metabolic model captures the primary metabolism of N. lanati and accurately predicts experimentally validated substrate utilization requirements. Additionally, metabolic flux predictions are verified by C-13 metabolic flux analysis, demonstrating that the model faithfully describes the underlying fungal metabolism. Furthermore, the model clarifies key aspects of the hydrogenosomal metabolism and can be used as a platform to quantitatively study these biotechnologically important yet poorly understood early-branching fungi. IMPORTANCE Recent genomic analyses have revealed that anaerobic gut fungi possess both the largest number and highest diversity of lignocellulolytic enzymes of all sequenced fungi, explaining their ability to decompose lignocellulosic substrates, e.g., agricultural waste, into fermentable sugars. Despite their potential, the development of engineering methods for these organisms has been slow due to their complex life cycle, understudied metabolism, and challenging anaerobic culture requirements. Currently, there is no framework that can be used to combine multi-omic data sets to understand their physiology. Here, we introduce a high-quality PacBio-sequenced genome of the anaerobic gut fungus Neocallimastix lanati. Beyond identifying a trove of lignocellulolytic enzymes, we use this genome to construct the first genome-scale metabolic model of an anaerobic gut fungus. The model is experimentally validated and sheds light on unresolved metabolic features common to gut fungi. Model-guided analysis will pave the way for deepening our understanding of anaerobic gut fungi and provides a systematic framework to guide strain engineering efforts of these organisms for biotechnological use.
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