TY - JOUR
T1 - Exploring microRNA Signatures of DNA Damage Response Using an Innovative System of Genotoxic Stress in Medicago truncatula Seedlings
AU - Gualtieri, Carla
AU - Gianella, Maraeva
AU - Pagano, Andrea
AU - Cadeddu, Tiziano
AU - Araújo, Susana
AU - Balestrazzi, Alma
AU - Macovei, Anca
N1 - Funding Information:
This research received support from the Italian Ministry of Education, University and Research (MIUR): Dipartimenti di Eccellenza Program (2018-2022) – Department of Biology and Biotechnology “L. Spallanzani,” University of Pavia. AP is supported by a Departmental Fellowship (assegno di ricerca tipo A – COD. DBB2020-A01). SA acknowledges the support of Fundação para a Ciência e Tecnologia throughout the Research Unit “GREEN-IT” (UID/Multi/04551/2020), as well as, the support the 3i Bioeconomy project POCI-01-0246-FEDER-026758, funded by the Fundo Europeu de Desenvolvimento Regional (FEDER), under the “Programa Operacional Temático Competitivdade e Internacionalização”-COMPETE 2020 and the program NORTE 2020 through the project NORTE-06-3559-FSE-000103 funded by the Fundo Social Europeu (FSE).
Funding Information:
We are thankful to Dr. Alexander Martinkosky (Drug Synthesis and Chemistry Branch, Developmental Therapeutics Program, Division of Cancer Treatment and Diagnosis, National Cancer Institute, Bethesda, United States) for providing the NSC120686 compound. We would also like to thank Dr. Jos? Parreira (ITQB-NOVA, Portugal) for the support provided during the setting-up of the analysis of miRNA expression. Funding. This research received support from the Italian Ministry of Education, University and Research (MIUR): Dipartimenti di Eccellenza Program (2018-2022) ? Department of Biology and Biotechnology ?L. Spallanzani,? University of Pavia. AP is supported by a Departmental Fellowship (assegno di ricerca tipo A ? COD. DBB2020-A01). SA acknowledges the support of Funda??o para a Ci?ncia e Tecnologia throughout the Research Unit ?GREEN-IT? (UID/Multi/04551/2020), as well as, the support the 3i Bioeconomy project POCI-01-0246-FEDER-026758, funded by the Fundo Europeu de Desenvolvimento Regional (FEDER), under the ?Programa Operacional Tem?tico Competitivdade e Internacionaliza??o?-COMPETE 2020 and the program NORTE 2020 through the project NORTE-06-3559-FSE-000103 funded by the Fundo Social Europeu (FSE).
Publisher Copyright:
© Copyright © 2021 Gualtieri, Gianella, Pagano, Cadeddu, Araújo, Balestrazzi and Macovei.
PY - 2021/3/9
Y1 - 2021/3/9
N2 - One of the challenges that living organisms face is to promptly respond to genotoxic stress to avoid DNA damage. To this purpose, all organisms, including plants, developed complex DNA damage response (DDR) mechanisms. These mechanisms are highly conserved among organisms and need to be finely regulated. In this scenario, microRNAs (miRNAs) are emerging as active players, thus attracting the attention of the research community. The involvement of miRNAs in DDR has been investigated prominently in human cells whereas studies in plants are still scarce. To experimentally investigate the involvement of plant miRNAs in the regulation of DDR-associated pathways, an ad hoc system was developed, using the model legume Medicago truncatula. Specific treatments with camptothecin (CPT) and/or NSC120686 (NSC), targeting distinct components of DDR, namely topoisomerase I (TopI) and tyrosyl-DNA phosphodiesterase 1 (TDP1), were used. Phenotypic (germination percentage and speed, seedling growth) and molecular (cell death, DNA damage, and gene expression profiles) analyses demonstrated that the imposed treatments impact DDR. Our results show that these treatments do not influence the germination process but rather inhibit seedling development, causing an increase in cell death and accumulation of DNA damage. Moreover, treatment-specific changes in the expression of suppressor of gamma response 1 (SOG1), master-regulator of plant DDR, were observed. Additionally, the expression of multiple genes playing important roles in different DNA repair pathways and cell cycle regulation were differentially expressed in a treatment-specific manner. Subsequently, specific miRNAs identified from our previous bioinformatics approaches as putatively targeting genes involved in DDR processes were investigated alongside their targets. The obtained results indicate that under most conditions when a miRNA is upregulated the corresponding candidate target gene is downregulated, providing an indirect evidence of miRNAs action over these targets. Hence, the present study extends the present knowledge on the information available regarding the roles played by miRNAs in the post-transcriptional regulation of DDR in plants.
AB - One of the challenges that living organisms face is to promptly respond to genotoxic stress to avoid DNA damage. To this purpose, all organisms, including plants, developed complex DNA damage response (DDR) mechanisms. These mechanisms are highly conserved among organisms and need to be finely regulated. In this scenario, microRNAs (miRNAs) are emerging as active players, thus attracting the attention of the research community. The involvement of miRNAs in DDR has been investigated prominently in human cells whereas studies in plants are still scarce. To experimentally investigate the involvement of plant miRNAs in the regulation of DDR-associated pathways, an ad hoc system was developed, using the model legume Medicago truncatula. Specific treatments with camptothecin (CPT) and/or NSC120686 (NSC), targeting distinct components of DDR, namely topoisomerase I (TopI) and tyrosyl-DNA phosphodiesterase 1 (TDP1), were used. Phenotypic (germination percentage and speed, seedling growth) and molecular (cell death, DNA damage, and gene expression profiles) analyses demonstrated that the imposed treatments impact DDR. Our results show that these treatments do not influence the germination process but rather inhibit seedling development, causing an increase in cell death and accumulation of DNA damage. Moreover, treatment-specific changes in the expression of suppressor of gamma response 1 (SOG1), master-regulator of plant DDR, were observed. Additionally, the expression of multiple genes playing important roles in different DNA repair pathways and cell cycle regulation were differentially expressed in a treatment-specific manner. Subsequently, specific miRNAs identified from our previous bioinformatics approaches as putatively targeting genes involved in DDR processes were investigated alongside their targets. The obtained results indicate that under most conditions when a miRNA is upregulated the corresponding candidate target gene is downregulated, providing an indirect evidence of miRNAs action over these targets. Hence, the present study extends the present knowledge on the information available regarding the roles played by miRNAs in the post-transcriptional regulation of DDR in plants.
KW - camptothecin
KW - DNA damage response
KW - genotoxicity
KW - microRNA
KW - NSC120686
KW - seedling development
KW - tyrosyl-DNA phosphodiesterase 1
UR - http://www.scopus.com/inward/record.url?scp=85102923700&partnerID=8YFLogxK
U2 - 10.3389/fpls.2021.645323
DO - 10.3389/fpls.2021.645323
M3 - Article
AN - SCOPUS:85102923700
SN - 1664-462X
VL - 12
JO - Frontiers in Plant Science
JF - Frontiers in Plant Science
M1 - 645323
ER -