scholarly journals LPMO-oxidized cellulose oligosaccharides evoke immunity in Arabidopsis conferring resistance towards necrotrophic fungus B. cinerea

2021 ◽  
Author(s):  
Marco Zarattini ◽  
Massimiliano Corso ◽  
Marco Antonio Kadowaki ◽  
Antonielle Monclaro ◽  
Silvia Magri ◽  
...  
Keyword(s):  
Cell Wall ◽  
Crucial Role ◽  
Plant Cell Wall ◽  
Pathogenic Fungus ◽  
Pairwise Comparison ◽  
Transcriptional Reprogramming ◽  
Specific Effects ◽  
Lytic Polysaccharide Monooxygenases ◽  
Polysaccharide Monooxygenases ◽  
Plant Pathogen Interactions

AbstractLytic Polysaccharide Monooxygenases (LPMOs) are powerful redox enzymes able to oxidatively cleave cellulose polymers. Widely conserved across biological kingdoms, LPMOs of the AA9 family are deployed by phytopathogens during necrotrophic attack of plant cell wall. In response, plants have evolved sophisticated mechanisms to sense cell wall damage and thus self-triggering Damage Triggered Immunity (DTI) responses. Here, we show that Arabidopsis plants exposed to LPMO products responds by activating the innate immunity ultimately leading to increased resistance to pathogenic fungus Botrytis cinerea. We demonstrated with microarray hybridization that plants undergo a deep transcriptional reprogramming upon elicitation with AA9 derived cellulose- or cello-oligosaccharides (AA9_COS). To decipher the specific effects of native and oxidized LPMO-generated cello-oligosaccharides, a pairwise comparison with cellobiose, the smallest non-oxidized unit constituting cellulose, is presented. Moreover, we identified two leucine-rich repeat receptor-like kinases, namely STRESS INDUCED FACTOR 2 and 4, playing a crucial role in signaling the AA9_COS-dependent responses such as camalexin production. We observed an increased production of ethylene, jasmonic and salicylic acid hormones, and finally deposition of callose in cell wall. Collectively, our data reveal that LPMOs might play a crucial role in plant-pathogen interactions.

scholarly journals LPMO-oxidized cellulose oligosaccharides evoke immunity in Arabidopsis conferring resistance towards necrotrophic fungus B. cinerea

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Marco Zarattini ◽  
Massimiliano Corso ◽  
Marco Antonio Kadowaki ◽  
Antonielle Monclaro ◽  
Silvia Magri ◽  
...  
Keyword(s):  
Cell Wall ◽  
Crucial Role ◽  
Pairwise Comparison ◽  
Leucine Rich Repeat ◽  
Redox Enzymes ◽  
Transcriptional Reprogramming ◽  
Specific Effects ◽  
Lytic Polysaccharide Monooxygenases ◽  
Polysaccharide Monooxygenases ◽  
Plant Pathogen Interactions

AbstractLytic Polysaccharide Monooxygenases (LPMOs) are powerful redox enzymes able to oxidatively cleave recalcitrant polysaccharides. Widely conserved across biological kingdoms, LPMOs of the AA9 family are deployed by phytopathogens to deconstruct cellulose polymers. In response, plants have evolved sophisticated mechanisms to sense cell wall damage and thus self-triggering Damage Triggered Immunity responses. Here, we show that Arabidopsis plants exposed to LPMO products triggered the innate immunity ultimately leading to increased resistance to the necrotrophic fungus Botrytis cinerea. We demonstrated that plants undergo a deep transcriptional reprogramming upon elicitation with AA9 derived cellulose- or cello-oligosaccharides (AA9_COS). To decipher the specific effects of native and oxidized LPMO-generated AA9_COS, a pairwise comparison with cellobiose, the smallest non-oxidized unit constituting cellulose, is presented. Moreover, we identified two leucine-rich repeat receptor-like kinases, namely STRESS INDUCED FACTOR 2 and 4, playing a crucial role in signaling the AA9_COS-dependent responses such as camalexin production. Furthermore, increased levels of ethylene, jasmonic and salicylic acid hormones, along with deposition of callose in the cell wall was observed. Collectively, our data reveal that LPMOs might play a crucial role in plant-pathogen interactions.

scholarly journals Secreted pectin monooxygenases drive plant infection by pathogenic oomycetes

Science ◽  
2021 ◽  
Vol 373 (6556) ◽  
pp. 774-779
Author(s):  
Federico Sabbadin ◽  
Saioa Urresti ◽  
Bernard Henrissat ◽  
Anna O. Avrova ◽  
Lydia R. J. Welsh ◽  
...  
Keyword(s):  
Plant Cell Wall ◽  
Crop Protection ◽  
Model Organism ◽  
Plant Infection ◽  
Lytic Polysaccharide Monooxygenases ◽  
Polysaccharide Monooxygenases ◽  
Encoding Gene ◽  
Plant Pathogen Interactions ◽  
Encoding Genes ◽  
Secreted Enzymes

The oomycete Phytophthora infestans is a damaging crop pathogen and a model organism to study plant-pathogen interactions. We report the discovery of a family of copper-dependent lytic polysaccharide monooxygenases (LPMOs) in plant pathogenic oomycetes and its role in plant infection by P. infestans. We show that LPMO-encoding genes are up-regulated early during infection and that the secreted enzymes oxidatively cleave the backbone of pectin, a charged polysaccharide in the plant cell wall. The crystal structure of the most abundant of these LPMOs sheds light on its ability to recognize and degrade pectin, and silencing the encoding gene in P. infestans inhibits infection of potato, indicating a role in host penetration. The identification of LPMOs as virulence factors in pathogenic oomycetes opens up opportunities in crop protection and food security.

scholarly journals Specific Xylan Activity Revealed for AA9 Lytic Polysaccharide Monooxygenases of the Thermophilic Fungus Malbranchea cinnamomea by Functional Characterization

2019 ◽  
Vol 85 (23) ◽  
Author(s):  
Silvia Hüttner ◽  
Anikó Várnai ◽  
Dejan M. Petrović ◽  
Cao Xuan Bach ◽  
Dang Thi Kim Anh ◽  
...  
Keyword(s):  
Cell Wall ◽  
Plant Cell ◽  
Plant Cell Wall ◽  
Functional Characterization ◽  
Cellulose Microfibrils ◽  
Cell Wall Polysaccharide ◽  
Poor Growth ◽  
Content Type ◽  
Lytic Polysaccharide Monooxygenases ◽  
Polysaccharide Monooxygenases

ABSTRACT The thermophilic biomass-degrader Malbranchea cinnamomea exhibits poor growth on cellulose but excellent growth on hemicelluloses as the sole carbon source. This is surprising considering that its genome encodes eight lytic polysaccharide monooxygenases (LPMOs) from auxiliary activity family 9 (AA9), enzymes known for their high potential in accelerating cellulose depolymerization. We characterized four of the eight (M. cinnamomea AA9s) McAA9s, namely, McAA9A, McAA9B, McAA9F, and McAA9H, to gain a deeper understanding about their roles in the fungus. The characterized McAA9s were active on hemicelluloses, including xylan, glucomannan, and xyloglucan, and furthermore, in accordance with transcriptomics data, differed in substrate specificity. Of the McAA9s, McAA9H is unique, as it preferentially cleaves residual xylan in phosphoric acid-swollen cellulose (PASC). Moreover, when exposed to cellulose-xylan blends, McAA9H shows a preference for xylan and for releasing (oxidized) xylooligosaccharides. The cellulose dependence of the xylan activity suggests that a flat conformation, with rigidity similar to that of cellulose microfibrils, is a prerequisite for productive interaction between xylan and the catalytic surface of the LPMO. McAA9H showed a similar trend on xyloglucan, underpinning the suggestion that LPMO activity on hemicelluloses strongly depends on the polymers’ physicochemical context and conformation. Our results support the notion that LPMO multiplicity in fungal genomes relates to the large variety of copolymeric polysaccharide arrangements occurring in the plant cell wall. IMPORTANCE The Malbranchea cinnamomea LPMOs (McAA9s) showed activity on a broad range of soluble and insoluble substrates, suggesting their involvement in various steps of biomass degradation besides cellulose decomposition. Our results indicate that the fungal AA9 family is more diverse than originally thought and able to degrade almost any kind of plant cell wall polysaccharide. The discovery of an AA9 that preferentially cleaves xylan enhances our understanding of the physiological roles of LPMOs and enables the use of xylan-specific LPMOs in future applications.

scholarly journals In vitro biosynthesis of poly-β-1,4-glucan derivatives using a pro-miscuous glycosyltransferase

2020 ◽  
Author(s):  
Gregory S. Bulmer ◽  
Ashley P. Mattey ◽  
Fabio Parmeggiani ◽  
Ryan Williams ◽  
Helene Ledru ◽  
...  
Keyword(s):  
Plant Cell Wall ◽  
Chemical Probes ◽  
Cell Wall Degrading Enzymes ◽  
Lytic Polysaccharide Monooxygenases ◽  
Natural Cellulose ◽  
Polysaccharide Monooxygenases ◽  
In Vitro Biosynthesis ◽  
New Applications ◽  
Biosynthetic Machinery

AbstractThe β-1,4-glucose linkage of cellulose is the most abundant polymeric linkage on earth and as such is of considerable interest in biology and biotechnology. It remains challenging to synthesize this linkage in vitro due to a lack of suitable biocatalysts; the natural cellulose biosynthetic machinery is a membrane-associated complex with processive activity that cannot be easily manipulated to synthesize tailor-made oligosaccharides and their derivatives. Here we identify a promiscuous activity of a soluble recombinant biocatalyst, Neisseria meningitidis glycosyltransferase LgtB, suitable for the polymerization of glucose from UDP-glucose via the generation of β-1,4-glycosidic linkages. We employed LgtB to synthesize natural and derivatized cello-oligosaccharides and we demonstrate how LgtB can be incorporated in biocatalytic cascades and chemo-enzymatic strategies to synthesize cello-oligosaccharides with tailored functionalities. We also show how the resulting glycan structures can be applied as chemical probes to report on activity and selectivity of plant cell wall degrading enzymes, including lytic polysaccharide monooxygenases. We anticipate that this biocatalytic approach to derivatized cello-oligosaccharides via glucose polymerization will open up new applications in biology and nanobiotechnology.

scholarly journals Identification of the molecular determinants driving the substrate specificity of fungal lytic polysaccharide monooxygenases (LPMOs)

2020 ◽  
pp. jbc.RA120.015545
Author(s):  
Kristian E. H. Frandsen ◽  
Mireille Haon ◽  
Sacha Grisel ◽  
Bernard Henrissat ◽  
Leila Lo Leggio ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Time Course ◽  
Plant Cell Wall ◽  
Plant Biomass ◽  
Substrate Binding ◽  
Glycoside Hydrolases ◽  
Sequence Alignments ◽  
Lytic Polysaccharide Monooxygenases ◽  
Molecular Determinants ◽  
Polysaccharide Monooxygenases

Understanding enzymatic breakdown of plant biomass is crucial to develop nature-inspired biotechnological processes. Lytic polysaccharide monooxygenases (LPMOs) are microbial enzymes secreted by fungal saprotrophs involved in carbon recycling. LPMOs modify biomass by oxidatively cleaving polysaccharides thereby enhancing the efficiency of glycoside hydrolases. Fungal AA9 LPMOs are active on cellulose but some members also display activity on hemicelluloses and/or oligosaccharides. Although the active site subsites are well defined for a few model LPMOs, the molecular determinants driving broad substrate specificity are still not easily predictable. Based on bioinformatic clustering and sequence alignments, we selected seven fungal AA9 LPMOs that differ in the amino-acid residues constituting their subsites. Investigation of their substrate specificities revealed that all these LPMOs are active on cellulose and cello-oligosaccharides, as well as plant cell wall-derived hemicellulosic polysaccharides and carry out C4 oxidative cleavage. The product profiles from cello-oligosaccharides degradation suggests that the subtle differences in amino acids sequence within the substrate-binding loop regions lead to different preferred binding modes. Our functional analyses allowed us to probe the molecular determinants of substrate binding within two AA9 LPMO sub-clusters. Many wood-degrading fungal species rich in AA9 genes have at least one AA9 enzyme with structural loop features that allow recognition of short β-(1,4)-linked glucan chains. Time-course monitoring of these AA9 LPMOs on cello-oligosaccharides also provides a useful model system for mechanistic studies of LPMO catalysis. These results are valuable for the understanding of LPMO contribution to wood decaying process in nature and for the development of sustainable biorefineries.

Learning from microbial strategies for polysaccharide degradation

2016 ◽  
Vol 44 (1) ◽  
pp. 94-108 ◽  
Author(s):  
Glyn R. Hemsworth ◽  
Guillaume Déjean ◽  
Gideon J. Davies ◽  
Harry Brumer
Keyword(s):  
Energy Source ◽  
Plant Cell Wall ◽  
Biomass Conversion ◽  
Human Gut ◽  
Human Diet ◽  
Healthy Human ◽  
Polysaccharide Degradation ◽  
Lytic Polysaccharide Monooxygenases ◽  
Polysaccharide Monooxygenases ◽  
Polysaccharide Utilization Loci

Complex carbohydrates are ubiquitous in all kingdoms of life. As major components of the plant cell wall they constitute both a rich renewable carbon source for biotechnological transformation into fuels, chemicals and materials, and also form an important energy source as part of a healthy human diet. In both contexts, there has been significant, sustained interest in understanding how microbes transform these substrates. Classical perspectives of microbial polysaccharide degradation are currently being augmented by recent advances in the discovery of lytic polysaccharide monooxygenases (LPMOs) and polysaccharide utilization loci (PULs). Fundamental discoveries in carbohydrate enzymology are both advancing biological understanding, as well as informing applications in industrial biomass conversion and modulation of the human gut microbiota to mediate health benefits.

scholarly journals Host Cell Wall Damage during Pathogen Infection: Mechanisms of Perception and Role in Plant-Pathogen Interactions

Plants ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 399
Author(s):  
Riccardo Lorrai ◽  
Simone Ferrari
Keyword(s):  
Cell Wall ◽  
Plant Cell Wall ◽  
Complex Structure ◽  
Host Cell Wall ◽  
Infection Mechanisms ◽  
Phytopathogenic Microorganisms ◽  
Plant Pathogen Interactions ◽  
Molecular Patterns ◽  
Damage Associated Molecular Patterns ◽  
Fine Tune

The plant cell wall (CW) is a complex structure that acts as a mechanical barrier, restricting the access to most microbes. Phytopathogenic microorganisms can deploy an arsenal of CW-degrading enzymes (CWDEs) that are required for virulence. In turn, plants have evolved proteins able to inhibit the activity of specific microbial CWDEs, reducing CW damage and favoring the accumulation of CW-derived fragments that act as damage-associated molecular patterns (DAMPs) and trigger an immune response in the host. CW-derived DAMPs might be a component of the complex system of surveillance of CW integrity (CWI), that plants have evolved to detect changes in CW properties. Microbial CWDEs can activate the plant CWI maintenance system and induce compensatory responses to reinforce CWs during infection. Recent evidence indicates that the CWI surveillance system interacts in a complex way with the innate immune system to fine-tune downstream responses and strike a balance between defense and growth.

scholarly journals Quantifying oxidation of cellulose-associated glucuronoxylan by two lytic polysaccharide monooxygenases from Neurospora crassa

2021 ◽  
Author(s):  
Olav A. Hegnar ◽  
Heidi Østby ◽  
Dejan M. Petrović ◽  
Lisbeth Olsson ◽  
Anikó Várnai ◽  
...  
Keyword(s):  
Neurospora Crassa ◽  
Plant Cell Wall ◽  
Plant Biomass ◽  
Hydrolytic Enzymes ◽  
Structural Features ◽  
Cell Wall Polysaccharides ◽  
Functional Variation ◽  
Lytic Polysaccharide Monooxygenases ◽  
Polysaccharide Monooxygenases ◽  
Oxidized Products

Family AA9 lytic polysaccharide monooxygenases (LPMOs) are abundant in fungi where they catalyze oxidative depolymerization of recalcitrant plant biomass. These AA9 LPMOs cleave cellulose, and some also act on hemicelluloses, primarily other (substituted) β-(1→4)-glucans. Oxidative cleavage of xylan has been shown for only a handful AA9 LPMOs, and it remains unclear whether this activity is a minor side reaction or primary function. Here, we show that Nc LPMO9F and the phylogenetically related, hitherto uncharacterized Nc LPMO9L from Neurospora crassa are active on both cellulose and cellulose-associated glucuronoxylan, but not on glucuronoxylan alone. A newly developed method for simultaneous quantification of xylan-derived and cellulose-derived oxidized products showed that Nc LPMO9F preferentially cleaves xylan when acting on a cellulose–beechwood glucuronoxylan mixture, yielding about three times more xylan-derived than cellulose-derived oxidized products. Interestingly, under similar conditions, Nc LPMO9L and previously characterized Mc LPMO9H from Malbranchea cinnamomea showed different xylan-to-cellulose preferences, giving oxidized product ratios of about 0.5:1 and 1:1, respectively, indicative of functional variation among xylan-active LPMOs. Phylogenetic and structural analysis of xylan-active AA9 LPMOs led to the identification of characteristic structural features, including unique features that do not occur in phylogenetically remote AA9 LPMOs, such as four AA9 LPMOs whose lack of activity towards glucuronoxylan was demonstrated in the present study. Taken together, the results provide a path towards discovery of additional xylan-active LPMOs and show that the huge family of AA9 LPMOs has members that preferentially act on xylan. These findings shed new light on the biological role and industrial potential of these fascinating enzymes. Importance Plant cell wall polysaccharides are highly resilient to depolymerization by hydrolytic enzymes, partly due to cellulose chains being tightly packed in microfibrils that are covered by hemicelluloses. Lytic polysaccharide monooxygenases (LPMOs) seem well suited to attack these resilient co-polymeric structures, but the occurrence and importance of hemicellulolytic activity among LPMOs remains unclear. Here we show that certain AA9 LPMOs preferentially cleave xylan when acting on a cellulose–glucuronoxylan mixture, and that this ability is the result of protein evolution that has resulted in a clade of AA9 LPMOs with specific structural features. Our findings strengthen the notion that the vast arsenal of AA9 LPMOs in certain fungal species provides functional versatility, and that AA9 LPMOs may have evolved to promote oxidative depolymerization of a wide variety of recalcitrant, co-polymeric plant polysaccharide structures. These findings have implications for understanding the biological roles and industrial potential of LPMOs.

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