Plant Biotechnology Laboratory
Referent: Dr. Anca Macovei
Co-workers: Conrado Jr. Dueñas (PhD student), Adriano Griffo (PhD student)
DDR (DNA DAMAGE RESPONSE) IN THE RESPONSE OF PLANTS TO GENOTOXIC STRESS.
DDR mechanisms specifically aimed at helping plants cope with the damaging effects of genotoxic stress. DDR is defined as a signal transduction pathway that detects DNA damage signals and transduces them to perform cellular responses (DNA repair, cell cycle arrest, programmed cell death, endoreduplication). Reactive oxygen species (ROS) act both as signal molecules and as harmful molecules, depending on their concentration and cellular localization. Links between ROS and DDR are hypothesized but not fully explained. In this topic we purpose to characterize at the molecular level of genes involved in DDR useful for defining the profiles of resistance to genotoxic stress in plants, for example: 1) genes that code for specific regulators of DDR - SOG1 (Suppressor of Gamma response 1); 2) genes encoding the enzymes tyrosyl-DNA phosphodiesterase 1 (TDP1) and tyrosyl-DNA phosphodiesterase 2 (TDP2) involved in the removal of the lesions induced by DNA Topoisomerase I and II and in different DNA repair mechanisms (BER, NER, NHEJ); 3) OGG1 (8-Oxoguanine DNA glycosylase) and FPG (Formamidopyrimidine (fapy) -DNA glycosylase) genes involved in the removal of oxidative DNA lesions (BER).
MICRORNAS: ROLE IN DDR AND CROSS-KINGDOM TRANSFER.
A basic aspect of transcription regulation processes concerns the activity of microRNAs. MicroRNAs contribute to the modulation of gene expression at the post-transcriptional level by inducing translation repression or gene silencing. A line of research is active which aims to investigate the role of miRNAs in the plant response to genotoxic stresses and in the context of DNA repair mechanisms. We are recently investigating the possible involvement of miRNAs in the regulation of cross-kingdom processes through bioinformatics and experimental approaches. These new activities have applications not only in the plant but also in the animal and biomedical fields.
DEVELOPMENT OF NON-INVASIVE METHODS FOR SEMEN QUALITY
An early, reliable and non-invasive assessment of seed quality is a valuable tool for optimizing seed production and safety checks, with practical and economic benefits. Currently, standard tests approved by the International Seed Testing Association (ISTA), such as germination tests, allow you to observe the behavior of seeds in the post-sowing phase. These procedures present problems with the direct use of seeds for testing and the time it takes to obtain relevant information. Despite its practicality, the use of a single test does not allow the analysis of multiple factors that can directly contribute to the quality of the semen or be indirectly predictive for its evaluation, so the information obtained is inevitably partial. In this case, we propose specific tests (detection of ROS molecules) and tools (biosensors to detect volatile molecules) to evaluate the quality of the seeds in a non-invasive way, together with treatments to increase their quality (e.g. seed priming).
Plant systems used in these works include Medicago truncatula, M. sativa, Glycine max, Solanum lycopersicum, Petunia hybrida, Arabidopsis thaliana, Oryza sativa, Zea mays. The main methodological approaches include: bioinformatics analysis (use of specific tools and database), PCR, RT-PCR, qRT-PCR (genes, miRNA), Comet assay, Diffusion assay, ROS detection assays (FOX, DCFH-DA, DAB, NBT), microscopy (optical, fluorescent, TEM), given 'omics' analysis, germination test.
National collaborations:
- M. Biggiogera (piattaforma di microscopia elettronica, UNIPV-DBB),
- E. Doria (biochimica vegetale, UNIPV-DBB),
- C. Calvio (genetica dei microorganismi, UNIPV-DBB),
- L.Pasotti (Modellistica Matematica e Biologia Sintetica, DIII),
- M. Conti (Laboratorio di Meccanica Computazionale e Materiali Avanzati, DICAR),
- D. Dondi (Lab. Chimica delle Radiazioni/Spettroscopia EPR, Dip. Chimica),
- A. Mondoni, G. Rossi (DSTA), P. Leonetti (IPSP-CNR, Bari),
- P. Santamaria (Università degli Studi di Bari),
- A. Moschetta (Università degli Studi di Bari)
- V. Locato (Università Campus Bio-Medico di Roma).
International collaborations:
- S. Araújo (ITQB, BCL3, Portogallo),
- C. Dobrota (UBB, Romania),
- C. Reynaud (IPS2, France),
- N. Tuteja (ICGEB, India),
- S.S. Gill (Center of Biotechnology MD University-Rohtak, India),
- Vishal Pandey (Varanasi University, India),
- Lata Shukla (Pondicherry University, India),
- I. Slamet-Loedin (IRRI, Filippine),
- J. Ray (Diliman University, Filippine).