Punzo, Paola (2016) Genomics of osmotic stress responses in Arabidopsis and crops. [Tesi di dottorato]

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Tipologia del documento:	Tesi di dottorato
Lingua:	English
Titolo:	Genomics of osmotic stress responses in Arabidopsis and crops
Autori:	Autore Email Punzo, Paola paolapunzo86@gmail.com
Data:	31 Marzo 2016
Numero di pagine:	132
Istituzione:	Università degli Studi di Napoli Federico II
Dipartimento:	Agraria
Scuola di dottorato:	Scienze agrarie e agro-alimentari
Dottorato:	Agrobiologia e agrochimica
Ciclo di dottorato:	28
Coordinatore del Corso di dottorato:	nome email Carputo, Domenico carputo@unina.it
Tutor:	nome email Grillo, Stefania [non definito]
Data:	31 Marzo 2016
Numero di pagine:	132
Parole chiave:	abiotic stress; functional genomics; transcriptomics
Settori scientifico-disciplinari del MIUR:	Area 07 - Scienze agrarie e veterinarie > AGR/07 - Genetica agraria
Depositato il:	13 Apr 2016 14:31
Ultima modifica:	28 Apr 2019 01:00
URI:	http://www.fedoa.unina.it/id/eprint/10935

Abstract

Drought and soil salinity limit available land for agriculture and reduce crop yields by imposing osmotic stress on plants. The identification of genes involved in plant tolerance mechanisms is a key goal to develop crops better capable to handle these stresses. The aim of this project was to identify genes involved in the plant response to osmotic stress using genomic approaches both in the model plant Arabidopsis thaliana and in a major crop in the Mediterranean basin, Solanum lycopersicum. Previously, several genes whose expression was differentially regulated in Solanum tuberosum culture cells adapted to increasing concentrations of polyethylene glycol (PEG) were identified. Here, the functional role of fifty of these genes was verified by studying their orthologues in A. thaliana. Homozygous knockout lines for each gene were subjected to a large-scale phenotype screening in order to identify genes involved in adaptation to osmotic stress. Using this strategy, we have identified 3 genes whose function in stress response was so far unknown: the splicing factor DRT111, the putative negative regulator of TOR pathway TIP41-like, the subunit of Pol III SIN-like. A detailed functional characterization for these three selected genes was performed through the use of specific advanced molecular technologies. Interestingly, germination analysis on plants with alterated expression of DRT111 as well as the reported association with another important splicing factor SUA suggest that DRT111 is involved in pre-mRNA splicing of ABI3, regulating ABA-related inhibition of seed germination. Similarly, TIP41-like possibly affects several pathways in the ABA-mediated response to osmotic stress. The abolished expression of TIP41-like leads to ABA hypersensitive phenotypes at germination and seedling stage as well as reduction in root development, indicating alteration in ABA biosynthesis and/or perception. Finally, plants with abolished expression of SIN-like show severe reduction in root growth, suggesting that the mutation in SIN-like leads to phenotypes mainly associated with altered cell development that could be explained by an alteration of RNAs transcribed by Pol III. Growing evidence, indeed, demonstrates that RNA regulatory mechanisms participate in the modulation of abiotic stress responses. RNA binding proteins play an important role in such mechanisms, but relatively few have been characterized in plants so far. Among the differentially expressed genes in S.tuberosum adapted cells, the Arabidopsis ortholog AtRGGA, encoding a glycine-rich RNA-binding protein, was previously partially characterized in responses to drought and salt stress. Here we analyze the protein role in the RNA regulatory mechanisms. As expected from the presence of Arg/Gly motif, we provide evidence that AtRGGA is capable to binding ribosomal RNAs both in vitro and in vivo. To identify protein partners of AtRGGA, yeast two-hybrid screening was performed using an Arabidopsis cDNA library. Most of the identified proteins are involved in RNA processing, transport and ribosome biogenesis. Therefore, the obtained results indicate a role of AtRGGA in transcriptional and post-transcriptional control of gene expression during osmotic stress. Furthermore, this study also gives a complete picture of the mechanisms employed to maintain a cellular and whole-plant homeostasis in tomato during drought stress. Transcriptome profile was analyzed in M82 genotype during two cycles of prolonged drought stress and one of rewatering, monitoring physiological parameters as well as ABA and proline accumulation. The observed reduction in leaf gas exchanges was concomitant with a general arrest of transcriptional activities. In particular, Gene Onthology (GO) categories such as cell proliferation and cell cycle were significantly enriched in the down-regulated fraction of genes upon drought stress. As expected, ABA and proline accumulated after prolonged water deficit, driving the observed enrichment of stress related GOs in the up-regulated gene fractions, which included transcripts putatively involved in stomatal movements as well as cellular homeostasis maintenance. Taken together, our results revealed promising mechanisms of plant acclimation to stress. DRT111, TIP41-like, SIN-like and AtRGGA are active participants in the response to drought and salinity and represent interesting targets for future use in crop species. Furthermore, the analyzed interconnection between the physiological responses and gene expression in tomato has given other interesting candidate genes that could play novel roles in drought tolerance and adaptation.

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