Table_1_RNA and mRNA Nitration as a Novel Metabolic Link in Potato Immune Response to Phytophthora infestans.DOCX

<p>Peroxynitrite (ONOO^-) exhibits a well-documented nitration activity in relation to proteins and lipids; however, the interaction of ONOO^- with nucleic acids remains unknown in plants. The study uncovers RNA and mRNA nitration as an integral event in plant metabolism intensified during immune response. Using potato-avr/vr Phytophthora infestans systems and immunoassays we documented that potato immunity is accompanied by two waves of boosted ONOO^- formation affecting guanine nucleotides embedded in RNA/mRNA and protein tyrosine residues. The early ONOO^- generation was orchestrated with an elevated level of protein nitration and a huge accumulation of 8-nitroguanine (8-NO₂-G) in RNA and mRNA pools confirmed as a biomarker of nucleic acid nitration. Importantly, potato cells lacking ONOO^- due to scavenger treatment and attacked by the avr pathogen exhibited a low level of 8-NO₂-G in the mRNA pool correlated with reduced symptoms of programmed cell death (PCD). The second burst of ONOO^- coincided both with an enhanced level of tyrosine-nitrated proteins identified as subtilisine-like proteases and diminished protease activity in cells surrounding the PCD zone. Nitration of both RNA/mRNA and proteins via NO/ONOO^- may constitute a new metabolic switch in redox regulation of PCD, potentially limiting its range in potato immunity to avr P. infestans.</p

Image_3_RNA and mRNA Nitration as a Novel Metabolic Link in Potato Immune Response to Phytophthora infestans.JPEG

<p>Peroxynitrite (ONOO^-) exhibits a well-documented nitration activity in relation to proteins and lipids; however, the interaction of ONOO^- with nucleic acids remains unknown in plants. The study uncovers RNA and mRNA nitration as an integral event in plant metabolism intensified during immune response. Using potato-avr/vr Phytophthora infestans systems and immunoassays we documented that potato immunity is accompanied by two waves of boosted ONOO^- formation affecting guanine nucleotides embedded in RNA/mRNA and protein tyrosine residues. The early ONOO^- generation was orchestrated with an elevated level of protein nitration and a huge accumulation of 8-nitroguanine (8-NO₂-G) in RNA and mRNA pools confirmed as a biomarker of nucleic acid nitration. Importantly, potato cells lacking ONOO^- due to scavenger treatment and attacked by the avr pathogen exhibited a low level of 8-NO₂-G in the mRNA pool correlated with reduced symptoms of programmed cell death (PCD). The second burst of ONOO^- coincided both with an enhanced level of tyrosine-nitrated proteins identified as subtilisine-like proteases and diminished protease activity in cells surrounding the PCD zone. Nitration of both RNA/mRNA and proteins via NO/ONOO^- may constitute a new metabolic switch in redox regulation of PCD, potentially limiting its range in potato immunity to avr P. infestans.</p

Table_2_RNA and mRNA Nitration as a Novel Metabolic Link in Potato Immune Response to Phytophthora infestans.DOCX

<p>Peroxynitrite (ONOO^-) exhibits a well-documented nitration activity in relation to proteins and lipids; however, the interaction of ONOO^- with nucleic acids remains unknown in plants. The study uncovers RNA and mRNA nitration as an integral event in plant metabolism intensified during immune response. Using potato-avr/vr Phytophthora infestans systems and immunoassays we documented that potato immunity is accompanied by two waves of boosted ONOO^- formation affecting guanine nucleotides embedded in RNA/mRNA and protein tyrosine residues. The early ONOO^- generation was orchestrated with an elevated level of protein nitration and a huge accumulation of 8-nitroguanine (8-NO₂-G) in RNA and mRNA pools confirmed as a biomarker of nucleic acid nitration. Importantly, potato cells lacking ONOO^- due to scavenger treatment and attacked by the avr pathogen exhibited a low level of 8-NO₂-G in the mRNA pool correlated with reduced symptoms of programmed cell death (PCD). The second burst of ONOO^- coincided both with an enhanced level of tyrosine-nitrated proteins identified as subtilisine-like proteases and diminished protease activity in cells surrounding the PCD zone. Nitration of both RNA/mRNA and proteins via NO/ONOO^- may constitute a new metabolic switch in redox regulation of PCD, potentially limiting its range in potato immunity to avr P. infestans.</p

Image_2_RNA and mRNA Nitration as a Novel Metabolic Link in Potato Immune Response to Phytophthora infestans.JPEG

<p>Peroxynitrite (ONOO^-) exhibits a well-documented nitration activity in relation to proteins and lipids; however, the interaction of ONOO^- with nucleic acids remains unknown in plants. The study uncovers RNA and mRNA nitration as an integral event in plant metabolism intensified during immune response. Using potato-avr/vr Phytophthora infestans systems and immunoassays we documented that potato immunity is accompanied by two waves of boosted ONOO^- formation affecting guanine nucleotides embedded in RNA/mRNA and protein tyrosine residues. The early ONOO^- generation was orchestrated with an elevated level of protein nitration and a huge accumulation of 8-nitroguanine (8-NO₂-G) in RNA and mRNA pools confirmed as a biomarker of nucleic acid nitration. Importantly, potato cells lacking ONOO^- due to scavenger treatment and attacked by the avr pathogen exhibited a low level of 8-NO₂-G in the mRNA pool correlated with reduced symptoms of programmed cell death (PCD). The second burst of ONOO^- coincided both with an enhanced level of tyrosine-nitrated proteins identified as subtilisine-like proteases and diminished protease activity in cells surrounding the PCD zone. Nitration of both RNA/mRNA and proteins via NO/ONOO^- may constitute a new metabolic switch in redox regulation of PCD, potentially limiting its range in potato immunity to avr P. infestans.</p

Image_1_RNA and mRNA Nitration as a Novel Metabolic Link in Potato Immune Response to Phytophthora infestans.JPEG

<p>Peroxynitrite (ONOO^-) exhibits a well-documented nitration activity in relation to proteins and lipids; however, the interaction of ONOO^- with nucleic acids remains unknown in plants. The study uncovers RNA and mRNA nitration as an integral event in plant metabolism intensified during immune response. Using potato-avr/vr Phytophthora infestans systems and immunoassays we documented that potato immunity is accompanied by two waves of boosted ONOO^- formation affecting guanine nucleotides embedded in RNA/mRNA and protein tyrosine residues. The early ONOO^- generation was orchestrated with an elevated level of protein nitration and a huge accumulation of 8-nitroguanine (8-NO₂-G) in RNA and mRNA pools confirmed as a biomarker of nucleic acid nitration. Importantly, potato cells lacking ONOO^- due to scavenger treatment and attacked by the avr pathogen exhibited a low level of 8-NO₂-G in the mRNA pool correlated with reduced symptoms of programmed cell death (PCD). The second burst of ONOO^- coincided both with an enhanced level of tyrosine-nitrated proteins identified as subtilisine-like proteases and diminished protease activity in cells surrounding the PCD zone. Nitration of both RNA/mRNA and proteins via NO/ONOO^- may constitute a new metabolic switch in redox regulation of PCD, potentially limiting its range in potato immunity to avr P. infestans.</p

Image_2_Insight into metabolic sensors of nitrosative stress protection in Phytophthora infestans.jpeg

Phytophthora infestans, a representative of phytopathogenic oomycetes, have been proven to cope with redundant sources of internal and host-derived reactive nitrogen species (RNS). To gain insight into its nitrosative stress resistance mechanisms, metabolic sensors activated in response to nitrosative challenge during both in vitro growth and colonization of the host plant were investigated. The conducted analyses of gene expression, protein accumulation, and enzyme activity reveal for the first time that P. infestans (avirulent MP946 and virulent MP977 toward potato cv. Sarpo Mira) withstands nitrosative challenge and has an efficient system of RNS elimination. The obtained data indicate that the system protecting P. infestans against nitric oxide (NO) involved the expression of the nitric oxide dioxygenase (Pi-NOD1) gene belonging to the globin family. The maintenance of RNS homeostasis was also supported by an elevated S-nitrosoglutathione reductase activity and upregulation of peroxiredoxin 2 at the transcript and protein levels; however, the virulence pattern determined the expression abundance. Based on the experiments, it can be concluded that P. infestans possesses a multifarious system of metabolic sensors controlling RNS balance via detoxification, allowing the oomycete to exist in different micro-environments flexibly.</p

Image_5_Insight into metabolic sensors of nitrosative stress protection in Phytophthora infestans.tif

Phytophthora infestans, a representative of phytopathogenic oomycetes, have been proven to cope with redundant sources of internal and host-derived reactive nitrogen species (RNS). To gain insight into its nitrosative stress resistance mechanisms, metabolic sensors activated in response to nitrosative challenge during both in vitro growth and colonization of the host plant were investigated. The conducted analyses of gene expression, protein accumulation, and enzyme activity reveal for the first time that P. infestans (avirulent MP946 and virulent MP977 toward potato cv. Sarpo Mira) withstands nitrosative challenge and has an efficient system of RNS elimination. The obtained data indicate that the system protecting P. infestans against nitric oxide (NO) involved the expression of the nitric oxide dioxygenase (Pi-NOD1) gene belonging to the globin family. The maintenance of RNS homeostasis was also supported by an elevated S-nitrosoglutathione reductase activity and upregulation of peroxiredoxin 2 at the transcript and protein levels; however, the virulence pattern determined the expression abundance. Based on the experiments, it can be concluded that P. infestans possesses a multifarious system of metabolic sensors controlling RNS balance via detoxification, allowing the oomycete to exist in different micro-environments flexibly.</p

Image_1_Insight into metabolic sensors of nitrosative stress protection in Phytophthora infestans.tif

Phytophthora infestans, a representative of phytopathogenic oomycetes, have been proven to cope with redundant sources of internal and host-derived reactive nitrogen species (RNS). To gain insight into its nitrosative stress resistance mechanisms, metabolic sensors activated in response to nitrosative challenge during both in vitro growth and colonization of the host plant were investigated. The conducted analyses of gene expression, protein accumulation, and enzyme activity reveal for the first time that P. infestans (avirulent MP946 and virulent MP977 toward potato cv. Sarpo Mira) withstands nitrosative challenge and has an efficient system of RNS elimination. The obtained data indicate that the system protecting P. infestans against nitric oxide (NO) involved the expression of the nitric oxide dioxygenase (Pi-NOD1) gene belonging to the globin family. The maintenance of RNS homeostasis was also supported by an elevated S-nitrosoglutathione reductase activity and upregulation of peroxiredoxin 2 at the transcript and protein levels; however, the virulence pattern determined the expression abundance. Based on the experiments, it can be concluded that P. infestans possesses a multifarious system of metabolic sensors controlling RNS balance via detoxification, allowing the oomycete to exist in different micro-environments flexibly.</p

Image_4_Insight into metabolic sensors of nitrosative stress protection in Phytophthora infestans.tif

Phytophthora infestans, a representative of phytopathogenic oomycetes, have been proven to cope with redundant sources of internal and host-derived reactive nitrogen species (RNS). To gain insight into its nitrosative stress resistance mechanisms, metabolic sensors activated in response to nitrosative challenge during both in vitro growth and colonization of the host plant were investigated. The conducted analyses of gene expression, protein accumulation, and enzyme activity reveal for the first time that P. infestans (avirulent MP946 and virulent MP977 toward potato cv. Sarpo Mira) withstands nitrosative challenge and has an efficient system of RNS elimination. The obtained data indicate that the system protecting P. infestans against nitric oxide (NO) involved the expression of the nitric oxide dioxygenase (Pi-NOD1) gene belonging to the globin family. The maintenance of RNS homeostasis was also supported by an elevated S-nitrosoglutathione reductase activity and upregulation of peroxiredoxin 2 at the transcript and protein levels; however, the virulence pattern determined the expression abundance. Based on the experiments, it can be concluded that P. infestans possesses a multifarious system of metabolic sensors controlling RNS balance via detoxification, allowing the oomycete to exist in different micro-environments flexibly.</p

Table_1_Insight into metabolic sensors of nitrosative stress protection in Phytophthora infestans.docx

Phytophthora infestans, a representative of phytopathogenic oomycetes, have been proven to cope with redundant sources of internal and host-derived reactive nitrogen species (RNS). To gain insight into its nitrosative stress resistance mechanisms, metabolic sensors activated in response to nitrosative challenge during both in vitro growth and colonization of the host plant were investigated. The conducted analyses of gene expression, protein accumulation, and enzyme activity reveal for the first time that P. infestans (avirulent MP946 and virulent MP977 toward potato cv. Sarpo Mira) withstands nitrosative challenge and has an efficient system of RNS elimination. The obtained data indicate that the system protecting P. infestans against nitric oxide (NO) involved the expression of the nitric oxide dioxygenase (Pi-NOD1) gene belonging to the globin family. The maintenance of RNS homeostasis was also supported by an elevated S-nitrosoglutathione reductase activity and upregulation of peroxiredoxin 2 at the transcript and protein levels; however, the virulence pattern determined the expression abundance. Based on the experiments, it can be concluded that P. infestans possesses a multifarious system of metabolic sensors controlling RNS balance via detoxification, allowing the oomycete to exist in different micro-environments flexibly.</p