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Tweaking proline metabolism in plants to increase their tolerance to stress

Alfalfa is the main forage legume used worldwide to feed cattle and proline accumulation is known to be a conserved response of plants to stress having multiple beneficial roles but this response has been poorly exploited for the development of more resilient crop varieties. Therefore, in this project we propose to develop an alfalfa resistant genotype by adjusting proline metabolism through CRISPR/Cas9 gene editing. The edited plants will be transgenesis free making them more acceptable by the society. Future applications of these results include the use of this legume in the livestock industry. As a sub-product we will also obtain knock-out plants for the gene P5CS2 (i.e. with reduced proline accumulation), which will be a valuable resource to support different ongoing basic science projects in our group and the group of our collaborators.

CRP - S. Signorelli. ICGEB, Italy.


Food and Plant Biology: CSIC Group Programe

By 2050 there will be a large gap between the world's capacity to produce food and its demand. This is mainly due to population growth, which results in higher demand, and soil erosion in combination with climate change, which compromises food production. Since plants respond to environmental changes to maintain their growth, understanding the responses to them is necessary either to enhance them through breeding or to establish management practices that favor crop productivity. The research group (GI) "Plant and Food Biology" aims to contribute to the biochemical knowledge of how plants respond to environmental factors, as well as the development of strategies to optimise these responses in order to improve the production/quality of crops. Our investigations use different crops, both for human consumption and forage, as well as the arabidopsis model plant. The crop in question varies depending on the specific question we sought to answer, always with the aim of understanding biochemical responses to environmental stimuli. Our group also contributes to HDR, teaching activities in undergraduate and postgraduate courses. 

Wheat type logo ea copy.jpg

CSIC Groups - S. Signorelli. CSIC Uruguay.

Proline accumulation in plants: a multidisciplinary approach to elucidate 

Proline accumulation is an evocatively conserved response of plants to stress. The biochemical and physiological relevance of this response is yet unclear and thus it has not been used as a molecular marker for the development of more resilient crops. This project aims to expand our current knowledge about the role of proline accumulation in plants acclimation to abiotic stress.

We will evaluate the proteome of KO mutant plants unable to accumulate proline under drought and saline stress conditions to determine which biological process are affected by proline accumulation. In these mutants we will also evaluate their photosynthetic capacity and chloroplastic oxidative damage under drought and saline stress conditions. Moreover, if proline catabolism contributes to the onset of induced autophagy. Finally, we will study the physico-chemical properties of proline to determine its osmolite and comsmotropic capacity relative to other compatible osmolites of plants. Understanding the role of proline accumulation under stress would expand our current knowledge in a topic of agronomical relevance. Furthermore, it would provide new concepts for breeders to consider in conventional breeding programmes or through biotechnology, to mitigate the negative impact caused by environmental stresses  on yield and quality of crops.

CSIC I+D Project - S. Signorelli. - A. H. Millar - CSIC, Uruguay.

Image by Rodion Kutsaev

Revealing the role of proline accumulation under abiotic stress in plants

Abiotic stresses (eg. drought) are consistent constraints to Uruguayan cropping yield and qualities. Proline is a relevant amino acid in plants, and its accumulation is linked to a plants’ response to abiotic stress. However, the biochemical and physiological relevance of this response remains poorly understood, and hence poorly exploited for the development of more resilient crop varieties. This is partially attributed to the fact that most researches have focused on proline itself, and not in the benefits of the activation of its biosynthesis and degradation. In this project, we will use an innovative approach focused on proline metabolism as a strategy of plants to enable effective acclimation to abiotic stress. The main hypothesis of this project is that proline accumulation is necessary for chloroplastic NADP regeneration, which is essential for sustaining photosynthetic activity. Hence, proline biosynthesis would play a major role in plants wellbeing under stress conditions. To prove this, we will evaluate the photosynthetic activity and the chloroplastic redox status in two independent KO-mutants for proline accumulation and the wild type plant under control, saline and drought conditions. Using proteomic approaches, we will determine the subcellular localization of proline metabolism enzymes, and identify differentially expressed proteins in the mutant lines to determine which metabolic process are affected by proline biosynthesis. The intended outcome of this project is to contribute to the understanding of this key stress response in plant biology and, in longer-term, provide significant benefits for plant breeders.

DT Project - S. Signorelli. CSIC Uruguay.

Test Tubes

Ecology, selection, formulation and use of benefit micro-organisms in agriculture

The need for reaching a sustainable agriculture has empowered the interest in biological nitrogen fixation (BNF) and thus the interest in developing new inoculants for the rhizobium-legume symbiosis, in particular for those of relevance in forage and food. 

In Uruguay, the current inoculants used for lotus, alfalfa and clover were selected in the 60s', in agronomical environments different to the ones that we have. At least, half of the producers do not inoculate lotus and clover, claiming that they do not observed any benefits from inoculation.

In Uruguayan fields, most of the nodules are occupied by native strains, and this was also observed in Australia and New Zealand. Ineficient-parasit populations can be the main cause for the failure of grasslands. Alfalfa is often used in marginal edaphic conditions with acidic soils which negatively affect the symbiosis and plant growth. Regional researchers have identified strains tolerant to acidic soils, yet not evaluated in field.  

In Uruguay, works from Mallarino et al. (1990 a,b,c) have suggested that the principal component determined by the N fixated is the produced area of the legume. However, the productivity of legumes in Uruguay is also limited by sub-optimal levels of soil pH and the nutrition of phosphorous, potassium and sulfur has a direct effect on the BNF.

This project aim to improve the sustainability of our pastoral animal production system by improving the BNF through (i) developing new inoculants based on efficient strains selected by their competitiveness and persistence, and (ii) improve our understanding of the effect of agronomical factors on the BNF.

J. Monza - S. Signorelli. FPTA-INIA Uruguay.

Cattle in Pasture

Editing of chili (Capsicum annuum) eIF4E gene to develop Potyvirus-resistant cultivar

Chili (Capsicum spp.) is one of the most important herbs in many cultures of the world. However, the supply is often inadequate to meet the demand due to diseases, such as potyvirus. This family of phyto-viruses is widespread globally and uses aphids as its transmission vectors. Chili and peppers are prone to infection of at least 11 types of potyvirus. Hence, this research aims to develop potyvirus-resistant cultivar of Chili (Capsicum annuum) through gene editing method of CRISPR-Cas9. Eukaryotic translation initiation factor 4E (eIF4E) is known to confer plant susceptibility towards the viruses, particularly potyvirus. The emergence of plant resistance trait to potyvirus by silencing (Knock Out) the eIF4Egene has been proven successful in several types of crops, such as tomatoes, cucumbers, and soybeans. Unfortunately, this strategy has negative drawbacks such as sacrificing plant fitness, because the susceptibility genes often also have important cellular roles. One way to solve this problem is to design a variant of the susceptibility gene that is identical to the resistant allele in nature. Potyvirus-resistant eIF4E allele in some chili cultivars were known to differ only at a few amino acids, compared to the potyvirus-susceptible eIF4E. The expected result from this research is a Chili cultivar with edited eIF4E gene that is resistant to potyvirus infection with unaltered morphologies.

Red Chili Pods

K. Meitha - RISET ITB 2021

Development of an alfalfa inoculant persistent to the acidic soils of the Uruguayan dairy basin

Dairy farmers use alfalfa (Medical sativa) in long-cycle pastures, although in a low proportion compared to legumes with lower productive capacity and/or persistence. Even though the edaphic conditions of the Diary Basin are marginal for alfalfa, its cultivation has a high productive potential, so that the expansion of the area can improve the competitiveness of the sector by reducing the costs of pastoral feeding. The reduced preference for this pasture is due to the variability of results obtained at the farm levels since the establishment, at which stage the nodulation is key for the later development. ​The soil acidity negatively affects the alfalfa-rizhobia symbiosis, and restricts the growth and persistence of Sinorhizobium meliloti, which conditions the production to the inoculation success. For this reason, it is relevant for Uruguayan dairy production to have persistent strains, such as those that nodulate Lotus corniculatus, Trifolium pratense and T. repent. In this project we propose (i) to develop an efficient inoculant for acid pH that persists in the soils of the Dairy Basin, (ii) to generate information of on how much the soil temperature affects the establishment of the symbiosis, which is applicable to recommendations on sowing dates. The prospection and evaluation of strains in the Dairy Basin will allow the fist approximation to a problem without dimension, which is the presence of parasite strains type Oregon in those soils.

J. Monza - FMV-ANII

Cows in Pasture
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