Nuclear cooperation, sharing of public goods and coordination of plant biomass utilization: probing functions unique to multinucleate syncytial fungi
EMSL Project ID
51549
Abstract
Filamentous fungi are essential for carbon recycling in ecosystems, in symbiotic interactions essential for plant productivity and in biotechnology as superb systems for protein production utilized for biofuel and bioproducts manufacturing and development. Filamentous fungi grow as multinucleate syncytial networks that show differentiation. For example, in mature colonies, apical hyphae, trunk hyphae, branch hyphae and aerial hyphae are present (analogous to meristems, stems, branches, and leaves in plants). Rules associated with regulatory aspects governing expression of genes, production of proteins and metabolic functions of these differentiated parts of a filamentous colony are essentially unknown. Upon exposure to heterogenous environments (such as complex structure of a plant cell wall), it is unclear which nuclei in the syncytium respond to environmental/nutritional cues, where mRNA is positioned relative to nucleus of origin, when/where mRNA is translated and how complex transcriptional programs that require precise levels of state variables, like the circadian clock and protein/enzyme production, are coordinated. One hypothesized cooperative advantage for multinucleate syncytia is that nuclei within a compartment can temporarily and flexibly adopt different roles in response to nutritional and environmental cues. Data from Ashbya gossypii and Neurospora crassa using single molecule fluorescence in situ hybridization (smFISH) and on colony transcriptional profiling in N. crassa suggests heterogeneity in transcription in hyphal compartments. In this proposal, we test the hypothesis that fungal multinucleate syncytia have unique regulatory processes essential for coordination of metabolic activities for optimal production of proteins associated with utilization of plant biomass, and which is coordinated with the circadian clock. Using a combination of transcriptomic, proteomic, and metabolomic approaches in collaboration with JGI and EMSL, we will dissect different regions of N. crassa colonies to determine the extent of heterogeneity of metabolic programs over circadian time. Using laser capture microdissection, nanoproteomic and spatial metabolomics, heterogeneity in different hyphal compartments will assessed, including transcriptional states using a JGI nuclear RNA-seq approach. Nuclei in multinucleate syncytia will be differentially labeled and nuclei-of-origin for transcripts encoding components required for the circadian clock and for plant biomass deconstruction will be determined using fluctuation localization imaging-based fluorescence in situ hybridization (fliFISH) and catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) at EMSL. This work will be complemented by protein localization data provided by the PI/co-PIs using a bank of fluorescently-tagged proteins. The availability of these data will enable us to discover the rules of regulatory processes across syncytial hyphal compartments in fungal colonies. We hypothesize that syncytial filamentous fungi, which are essential for carbon recycling and as industrial workhorses in the biotechnology sector, utilize unique rules that govern hyphal compartment sufficiency, nuclear transcriptional state, and mRNA, translation and protein localization. Elucidating these rules of organization is critical for understanding how syncytial organisms accomplish plant biomass deconstruction, carbon cycling, and nutrient transport in ecosystems. Importantly, defining the rules of multinucleate syncytia will enable new engineering strategies for strain-improvement of industrially important filamentous fungi for biofuels and bioproduct production.
Project Details
Project type
FICUS Research
Start Date
2020-10-01
End Date
2022-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members