DSpace Collection:https://hdl.handle.net/10171/425352024-03-28T11:00:05Z2024-03-28T11:00:05ZResponse of Tempranillo (Vitis vinifera L.) clones to climate change-related factors (elevated temperature, high CO2 and water deficit): plant performance and berry compositionhttps://hdl.handle.net/10171/637642023-03-20T12:51:48Z2022-06-30T00:00:00ZTitle: Response of Tempranillo (Vitis vinifera L.) clones to climate change-related factors (elevated temperature, high CO2 and water deficit): plant performance and berry composition
Abstract: SUMMARY Climate change is expected to modify future environmental conditions, therefore affecting agriculture.
Tempranillo, a largely cultivated worldwide grapevine (Vitis vinifera L.) red variety, will be affected by
the increase of global mean temperature and atmospheric CO2 levels and the decrease of water
availability in its cultivation area. The use of the intra-varietal diversity has been proposed as a strategy
for keeping wine typicity and regional varieties cultivation under future growing conditions by shifting
the ripening phase to more favourable environmental conditions. The aim of the thesis was to
determine the response of different clones of Tempranillo to simulated 2100 environmental
conditions, focusing on plant growth and development, as well as on berry composition. Fruit-bearing
cuttings of Tempranillo clones, which differed in the length of their reproductive cycle, were exposed
from fruit set to maturity to different scenarios of climate change in temperature gradient
greenhouses (TGG) and growth chamber greenhouses (GCG). The impact of elevated temperature (+4
°C), elevated CO2 (700 ppm) and water deficit, both in combination or independently, were evaluated.
The results show an increment of vegetative growth and a reduction of yield due to high temperatures.
Elevated CO2 concentration also increased vegetative growth and photosynthetic activity, even though
an acclimation process was observed, being stronger when combined with high temperature. Water
deficit reduced severely the photosynthetic activity and vegetative growth, overshadowing the
temperature and CO2 effects. Elevated temperature, both individually and combined with high CO2
levels, hastened sugar accumulation and advanced maturity, but these effects were mitigated by water
deficit. Malic acid degradation was also enhanced by high temperature, especially when combined
with elevated CO2 and water deficit. Amino acid concentration and profile were affected by high
temperature, elevated atmospheric CO2 and, especially, water deficit. Elevated CO2 reduced the effect
of temperature decoupling the anthocyanin and TSS accumulation; however, the combination of
elevated temperature, high CO2 and water deficit led to the imbalance between these two grape
components. Anthocyanin profile was modified by climate change, high temperature increasing the
relative abundance of acylated forms and both elevated CO2 and drought favouring the relative
content of malvidin and acylated, methylated and tri-hydroxylated forms. The clones studied showed
differences in their phenological development, vegetative and reproductive growth, as well as in their
grape composition. In addition, the results reveal the existence of a differential response of
Tempranillo clones to the environmental conditions projected for 2100 in relation to plant
performance and grape composition. In general, RJ43 was the most affected by the future growing
conditions (high temperature, elevated CO2 and water deficit) among the clones studied in terms of
phenology and anthocyanin concentration and profile. Conversely, VN31 maintained the highest
anthocyanin and anthoycianin:TSS ratio, whereas 1084 had the lowest sugar, malic acid and
anthocyanin levels. The differences observed in the response of the clones to climate change not
always depended on their reproductive cycle length.
Keywords: Climate change; Grapevine; Tempranillo; Clones; Intra-varietal diversity; Vegetative
development; Grape composition; Anthocyanin profile.; RESUMEN Se espera que el cambio climático modifique las condiciones ambientales futuras, afectando a la
agricultura. Tempranillo, una variedad tinta de vid (Vitis vinifera L.) cultivada en todo el mundo, se verá
afectada por el aumento de la temperatura media global y la concentración atmosférica de CO2 así
como porla disminución del agua disponible en el área de cultivo de esta variedad. Entre las estrategias
para mitigar el impacto del cambio climático sobre las características del vino y mantener en el futuro
el cultivo de variedades ligadas a una región determinada, se encuentra el uso de la diversidad
intravarietal para trasladar el momento de maduración a épocas con condiciones ambientales más
favorables. El objetivo de esta tesis fue determinar la respuesta de distintos clones de Tempranillo a
las condiciones ambientales previstas para 2100, centrándonos en el crecimiento y desarrollo de las
plantas, así como en la composición de las bayas. Para ello, se utilizaron esquejes fructíferos de clones
de Tempranillo caracterizados con distinta duración del ciclo reproductivo, siendo expuestos a
distintas condiciones ambientales en túneles de gradiente térmico (Temperature gradient
greenhouses, TGG) e invernaderos cámara (Growth chamber greenhouses, GCG) desde cuajado a
madurez. Se evaluó el impacto del aumento de la temperatura (+4 °C), del CO2 elevado (700 ppm) y
del déficit hídrico, tanto de forma individual como combinada. Los resultados muestran un aumento
del crecimiento vegetativo y una reducción de la producción de fruto debido a las altas temperaturas.
El CO2 elevado también aumentó el crecimiento vegetativo y la actividad fotosintética, aunque se
observó un proceso de aclimatación, siendo más intenso en combinación con altas temperaturas. La
sequía redujo drásticamente la actividad fotosintética y el crecimiento vegetativo, anulando los efectos
de la temperatura y del CO2. La temperatura elevada, tanto de forma individual como combinada con
altos niveles de CO2, aceleró la acumulación de azúcares y adelantó la madurez, siendo estos efectos
mitigados por el déficit hídrico. La degradación del ácido málico también se incrementó con el aumento
de la temperatura, especialmente en combinación con alto CO2 y sequía. La concentración y el perfil
de aminoácidos se vieron afectados por el aumento de la temperatura, por el alto CO2 y,
especialmente, por el déficit hídrico. El CO2 elevado disminuyó el desacoplamiento en la acumulación
de antocianinas y azúcares inducido por la temperatura elevada; sin embargo, la combinación de
temperatura elevada, alto CO2 y sequía provocó el desequilibrio entre estos compuestos. Las
condiciones de cambio climático alteraron el perfil de antocianinas; así, la temperatura elevada
aumentó la abundancia relativa de formas aciladas y, tanto el CO2 elevado como la sequía, favorecieron
la acumulación de malvidina y de formas aciladas, metiladas y trihidroxiladas. Los clones estudiados
mostraron diferencias en su desarrollo fenológico, crecimiento vegetativo y reproductivo, así como en
la composición de la baya. Además, los resultados revelan la existencia de una respuesta diferente de
los clones de Tempranillo estudiados a las condiciones ambientales proyectadas para 2100 en términos
de crecimiento y desarrollo de la planta, como de composición de la uva. En términos generales, el
clon más afectado por las condiciones ambientales que se esperan para finales del presente siglo (alta
temperatura, alto CO2 y déficit hídrico) fue el RJ43 en cuanto a su desarrollo fenológico, la
concentración de antocianinas y la relación antocianinas:azúcares. Además, bajo dichas condiciones,
VN31 presentó la concentración de antocianinas y la relación antocianinas:azúcares más alta, mientras
que las bayas de 1084 presentaron los niveles más bajos de azúcares, ácido málico y antocianinas. Las
diferencias en la respuesta de los clones al cambio climático no siempre dependieron de la duración
de su ciclo reproductivo.; RESUMÉ Le changement climatique devrait modifier les conditions environnementales dans le future, affectant
ainsi l'agriculture. Le Tempranillo, une variété de vigne rouge (Vitis vinifera L .) largement cultivée au
niveau international, pourrait être affecté par l’augmentation des températures moyennes mondiales
et des niveaux de CO2 dans l’atmosphère, ainsi que par la diminution de la disponibilité en eau sur sa
zone traditionnelle de culture. L'utilisation de la diversité intra-variétale a été proposée comme une
stratégie pour essayer de conserver la typicité du vin et les variétés régionales dans les conditions de
culture futures, en déplaçant la phase de maturation vers des périodes aux conditions
environnementales plus favorables. L’objectif de cette thèse était donc de déterminer la réponse de
différents clones de Tempranillo aux conditions environnementales simulées de 2100, en se
concentrant sur la croissance et le développement des plantes, ainsi que sur la composition des baies.
Des boutures fructifères de clones de Tempranillo, dont la longueur du cycle de reproduction était
différente, ont été exposées à différents scénarios climatiques dans des serres à gradient de
température (TGG) et des serres de chambre de croissance (GCG) depuis la fructification jusqu’à la
maturité. Les impacts de la température élevée (+4 ° C), du CO2 élevé (700 ppm) et du déficit en eau,
combinés ou non, ont été évalués. Les résultats montrent une augmentation de la croissance
végétative et une réduction de la production dues aux températures élevées. La concentration élevée
de CO2 a également augmenté la croissance végétative et l'activité photosynthétique. Néanmoins, un
processus d'acclimatation a été observé, celui-ci étant plus fort lorsqu’un haut niveau de CO2 est
combiné à une température élevée. Le déficit en eau a fortement réduit l'activité photosynthétique et
la croissance végétative, occultant les effets de la température et du CO2. La température élevée, que
ce soit individuellement ou associée à des niveaux élevés de CO2, a accéléré l'accumulation de sucres
et la date de maturité a été avancée, mais ces effets ont été atténués par le déficit en eau. La
dégradation de l’acide malique a également été favorisée par l’augmentation de la température, en
particulier lorsque cette dernière est associée à une concentration élevée de CO2 et à un déficit en eau.
La concentration et le profil des acides aminés ont été influencés par les températures élevées, un
niveau de CO2 élevé et, en particulier, par un déficit en eau. L'augmentation de CO2 a réduit l'effet de
la température sur le découplage de l’accumulation des anthocyanes par rapport à celle des sucres ;
cependant, la combinaison d’une température élevée, d’un haut niveau de CO2 et d’un déficit en eau
a conduit à un déséquilibre entre ces deux composés du raisin. Le profil des anthocyanes a été modifié
par le changement climatique, une température élevée augmentant la proportion des formes acylées
tandis qu’un haut niveau de CO2 et un déficit hydrique ont favorisé quant à eux l'abondance relative
de la malvidine, et des formes acylées, méthylées et trihydroxylées. Les clones étudiés ont montré des
différences dans leur développement phénologique, leur croissance végétative et reproductive, ainsi
que dans la composition de leurs raisins. En outre, les résultats révèlent l’existence d’une réponse
différentielle des clones de Tempranillo aux conditions environnementales prévues pour 2100 en
termes de performance de la plante et de composition du raisin. De façon générale, parmi les clones
étudiés, RJ43 fut le plus affecté par les conditions de croissance futures (températures élevées, haut
niveau de CO2 et déficit en eau) aussi bien en termes de développement phénologique qu’en termes
de concentration en anthocyanes et de leur profil. A l’inverse, VN31 a maintenu la plus haute teneur
en anthocyanes et le ratio anthoycyanes sur TSS les plus élevé tandis que 1084 a montré les teneurs
les plus faibles en sucres, acide malique et en anthocyanes et le ratio anthocyanes sur TSS le plus bas.
Les différences observées dans la réponse des clones au changement climatique ne dépendent pas
toujours de la longueur du cycle de reproduction.2022-06-30T00:00:00ZEstudio de la implicación de la presentación cruzada de antígenos en la actividad antitumoral de anticuerpos monoclonaleshttps://hdl.handle.net/10171/627762022-01-21T02:03:29Z2022-01-20T00:00:00ZTitle: Estudio de la implicación de la presentación cruzada de antígenos en la actividad antitumoral de anticuerpos monoclonales
Abstract: This PhD project has served to uncover the essential role cDC1s and cross-presentation play in the success of the immunotherapeutic agents anti-PD-1 and anti-CD137, analogous to those available in the clinic and that have revolutionized treatment of cancer. We have done so in loss-of-function settings using mouse genetically deficient for Batf3 and devoid of cDC1s, which displayed complete unresponsiveness to immunotherapy. Next, we have devised gain-of-function experiments aimed to systemically and locally expand cDC1 populations, while at the same time providing local activation signals to mature them. In the first chapter, we chose to expand cDC1s by systemically administering sFlt3L through hydrodynamic injection of sFlt3L-coding plasmid, and to locally activate them by intratumoral injection of Hiltonol®, Poly-ICLC, a TLR3 agonist available in the clinic. In the second chapter, we cloned XCL1 and sFlt3L into a Semliki Forest Virus vector (SFV-XF) for intratumoral administration. In this setting, both transgenes were intended to cause chemoattraction and differentiation of cDC1s, while viral RNA would provide the activation signals to drive DC maturation and potentiate CD8 T-cell cross-priming. Although we did not manage to detect increased cDC1 infiltration into injected tumors, SFV-XF showed robust antitumor efficacy against different tumor models in mice and promoted accumulation of conventional DCs in tumor-draining and distant lymph nodes.2022-01-20T00:00:00ZAclimatación fotosintética, producción y calidad de plantas de vid (Vitis vinifera L.) de la variedad Tempranillo Tinto y Blanco, cultivadas en diferentes escenarios de cambio climáticohttps://hdl.handle.net/10171/627742022-01-20T02:04:22Z2022-01-19T00:00:00ZTitle: Aclimatación fotosintética, producción y calidad de plantas de vid (Vitis vinifera L.) de la variedad Tempranillo Tinto y Blanco, cultivadas en diferentes escenarios de cambio climático
Abstract: Atmospheric CO2 concentration has increased from preindustrial level of 280 μmol CO2 mol-1 air (ppm) to currently more than 400 ppm. Intergovernmental Panel on Climate Change foresees that atmospheric CO2 concentration will continue increasing to reach at the end of this century 700 ppm. Due to its greenhouse effect, elevated atmospheric CO2 concentration is leading to higher atmospheric temperatures, a phenomenon that is being accompanied by episodes of less water availability or drought periods. Grapevine (Vitis vinifera L.) is a plant species very sensitive to those environmental factors. Fruit-bearing cuttings of red and white Tempranillo were grown under elevated CO2 (around 700 ppm, versus 400), high temperature (ambient temperature + 4ºC, versus ambient) and water deficit (cyclic drought, versus well irrigated) in temperature gradient greenhouses located at the University of Navarra (Pamplona, Spain) for three consecutive growing seasons (years 2013, 2014 and 2015). Vegetative growth (total vegetative mass and leaf area) was significantly reduced by drought (consistent the three years) and was more associated to a worse substrate water status than to decreases in leaf water content. In spite of decreasing leaf water content in both cultivars, elevated CO2 stimulated more vegetative than reproductive growth. The largest increases of elevated CO2 were observed in leaf and root growth in white and red Tempranillo, respectively. There was a clear interaction between temperature and water availability. Yield was significantly reduced by drought and was year-dependent, being especially low in 2015 due to eventual heat shocks episodes. The high temperatures of the heat shocks (above 35ºC) induced berry burn and browning and finally loss of 50% of the berries. The response of berry quality to climate change-related factors was highly variable and depended on the year. However, some general conclusions can be reached from the three years of experimentation. Thus, the simulated climate change scenarios affected to a greater extent the technological maturity parameters (primary metabolism) than the phenolic maturity (secondary metabolism). In particular, high temperature and drought significantly and consistently increased must pH, due to the decrease in malic acid. On the contrary, elevated CO2 decreased pH associated with significant increases in tartaric acid. These changes of the must affect its quality and potentially that of the resulting wine. Under elevated CO2 concentration, grapevine photosynthesis increases, modulated by temperature and water availability. However, under prolonged exposure to elevated CO2, grapevine down-regulates photosynthesis, decreasing photosynthetic capacity. Grapevine plants underwent photosynthetic acclimation after a long exposure to elevated CO2, regardless of temperature and water availability. Evidence comes from photosynthetic capacity decreases, leaf starch accumulation, and increases in leaf carbon/nitrogen ratio. Photosynthetic acclimation was well correlated to leaf starch, but not to soluble sugars. The white Tempranillo has altered its response to prolonged exposure to elevated CO2. For any given sink size or any given leaf starch accumulation, white Tempranillo always had higher levels of photosynthetic acclimation than the red one. Data suggest that mutation in white Tempranillo has affected loci other than grape color.2022-01-19T00:00:00ZIdentification of novel regulators of transcription in iPSC-derived cardiovascular progenitorshttps://hdl.handle.net/10171/588882024-01-24T09:29:36Z2020-03-16T00:00:00ZTitle: Identification of novel regulators of transcription in iPSC-derived cardiovascular progenitors
Abstract: Stem cells allow to investigate about the basic mechanisms that regulate embryonic development, cellular plasticity, and organ maintenance and regeneration. Induced pluripotent stem cells (iPSCs) are a powerful source of cells for diverse applications such as developmental and disease modeling, drug discovery and regenerative medicine. Cardiac development is coordinated by complex interactions between cardiac progenitor cell populations, different molecular signaling pathways, and spatially and temporally regulated gene expression. Cardiovascular progenitors (CVPs), with similar potential to these present in early stages of embryonic development, can be obtained from iPSCs by mimicking signaling during cardiogenesis, creating an ideal cell source to treat the damaged heart. However, the mechanisms and conditions for long-term self-renewal and maintenance of CVPs remain elusive. The generation of new iPSC models for tracing CVP lineages permits to delve into the biology of CVPs and discover novel potential regulators of their fate. We have established three different Cre/LoxP mouse models for lineage tracing of CVPs and their cell progeny by the expression of ZsGreen (ZsG) protein: Ai6-Mesp1-Cre (Mesp1 tracer), Ai6-Isl1-Cre (Isl1 tracer) and Ai6-Mef2c-AHF-Cre (AHF tracer) mice. Multiple iPSC clones have been derived from Ai6-Isl1-Cre and Ai6-Mef2c-AHF-Cre reporter mice. Several generated iPSC lines have been fully characterized, demonstrating embryonic stem-like features. iPSCs encoded the expected genomic insertions, showed normal karyotypes, transgenes were silenced, and expressed endogenous pluripotency-associated markers. Moreover, iPSCs were capable to differentiate into the three germ layers both in vitro and in vivo. We have verified the utility of established AHFiPSCs to track CVPs and their differentiated progeny. Upon differentiation, ZsG+ cells derived from AHFiPSCs appeared from embryoid body (EB) day 6 onwards, expressed cardiovascular-related markers, and were able to differentiate into cardiomyocytes, endothelial and smooth muscle cells. Comparative gene expression analysis using four different AHFiPSC lines revealed distinct molecular signatures in three particular stages of differentiation: undifferentiated iPSCs (AHFiPS-D0), sorted ZsG+ cells at day 6 (AHFiPS-D6.ZsG+) and sorted ZsG+ cells at day 13 of differentiation (AHFiPS-D13.ZsG+), that expressed pluripotency-, CVP- and cardiac/vascular lineages-associated markers, respectively. Gapdh and Polr2a have been determined the most stable housekeeping genes along the differentiation of AHFiPSCs, being optimal for accurate normalization of gene expression in our samples. We have identified novel regulators of transcription specifically upregulated in CVPs: Lin28a (and its paralog Lin28b), Lhx1 and Nr6a1. The expression of these genes was also found increased in the corresponding CVP-enriched samples from two different public analyses using mouse and human pluripotent stem cells. Moreover, using bioinformatic analysis of biological pathways we have found p53 as an interconnecting molecule of all these selected regulators of transcription. In order to explore novel insights into the biological role of the selected regulators in CVP fate, we have generated an inducible vector (pTRE-CDS-IRES-Puro-REX1-Blast) to carry out gain-of-function (GOF) analyses. This Tet-On system worked properly in AHFiPSC clones, but unfortunately it failed to work in EBs of certain size along differentiation. In contrast, this GOF system correctly functioned in human iPSCs differentiated in monolayer cultures. We have established four CBiPS1sv-4F-5 cell lines carrying Tet-On systems for the inducible expression of LIN28A, LIN28B, NR6A1 and LHX1, and preliminary results indicated that these regulators of transcription might have a role in CVP fate determination.2020-03-16T00:00:00Z