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- Institut für Biochemie und Biologie (655) (remove)
Development of a CRISPR/Cas gene editing technique for the coccolithophore Chrysotila carterae
(2024)
Light is the essential energy source for plants to drive photosynthesis. In nature, light availability is highly variable and often fluctuates on very short time scales. As a result, plants developed mechanisms to cope with these fluctuations. Understanding how to improve light use efficiency in natural fluctuating light (FL) conditions is a major target for agronomy.
In the first project, we identified an Arabidopsis thaliana plant that showed reduced levels of rapidly inducible non-photochemical quenching (NPQ). This plant was devoid of any T-DNA insertion. Using a mapping-by-sequencing approach, we successfully located the causal genomic region near the end of chromosome 4. Through variant investigations in that region, we identified a deletion of about 20 kb encompassing 9 genes. By complementation analysis, we confirmed that one of the deleted genes, VTC2, is the causal gene responsible for the low NPQ. Loss of VTC2 decreased NPQ particularly in old leaves, with young leaves being only slightly affected. Additionally, ascorbate levels were almost abolished in old leaves, likely causing the NPQ decrease by reducing the activity of the xanthophyll cycle. Although ascorbate levels in younger leaves were reduced compared to wild-type plants, they remained at a comparably higher level. This difference may be due to the VTC2 paralog VTC5, which is expressed at a higher level in young leaves than in old ones.
Plants require the PROTON GRADIENT REGULATION 5 (PGR5) protein for survival in FL. pgr5 mutants die because they fail to increase the luminal proton concentration in response to high light (HL) phases. A rapid elevation in ∆pH is needed to slow down electron transport through the Cytochrome b6 f complex (photosynthetic control). In FL, such lack of control in the pgr5 mutants results in photosystem I (PSI) overreduction, reactive oxygen species (ROS) production, and cell death. Decreases in photosystem II (PSII) activity introduced by crossing pgr5 with PSII deficient mutants
rescued the lethality of pgr5 in FL. PGR5 was suggested to act as part of the ferredoxin-plastoquinone reductase (FQR), involved in cyclic electron transfer around PSI. However, the proposed molecular role of PGR5 remains highly debated. To learn more about PGR5 function, we performed a forward genetic screen in Arabidopsis thaliana to identify EMS-induced suppressor mutants surviving longer when grown in FL compared to pgr5 mutants (referred to as ”suppressor of pgr5 lethality in fluctuating light”, splf ). 11 different candidate genes were identified in a total of 22 splf plants.
Mutants of seven of these genes in the pgr5 background showed low Fv/Fm values when grown in non-fluctuating low light (LL). Five of these 4genes were previously reported to have a role in PSII biogenesis or function. Two others, RPH1 and a DEAD/DEAH box helicase (AT3G02060), have not been linked to PSII function before. Three of splf candidate genes link to primary metabolism, fructose-2,6-bisphosphatase (F2KP ), udp-glucose pyrophosphorylase 1 (UGP1 ) and ferredoxin-dependent glutamate synthase (Fd-GOGAT ). They are characterized by the fact that they survive longer in FL than pgr5 mutants but do not procede beyond the early vegetative
phase and then die.
Plant metabolism serves as the primary mechanism for converting assimilated carbon into essential compounds crucial for plant growth and ultimately, crop yield. This renders it a focal point of research with significant implications. Despite notable strides in comprehending the genetic principles underpinning metabolism and yield, there remains a dearth of knowledge regarding the genetic factors responsible for trait variation under varying environmental conditions. Given the burgeoning global population and the advancing challenges posed by climate change, unraveling the intricacies of metabolic and yield responses to water scarcity became increasingly important in safeguarding food security.
Our research group has recently started to work on the genetic resources of legume species. To this end, the study presented here investigates the metabolic diversity across five different legume species at a tissue level, identifying species-specific biosynthesis of alkaloids as well as iso-/flavonoids with diverse functional groups, namely prenylation, phenylacylation as well as methoxylation, to create a resource for follow up studies investigation the metabolic diversity in natural diverse populations of legume species.
Following this, the second study investigates the genetic architecture of drought-induced changes in a global common bean population. Here, a plethora of quantitative trait loci (QTL) associated with various traits are identified by performing genome-wide association studies (GWAS), including for lipid signaling. On this site, overexpression of candidates highlighted the induction of several oxylipins reported to be pivotal in coping with harsh environmental conditions such as water scarcity.
Diverging from the common bean and GWAS, the following study focuses on identifying drought-related QTL in tomato using a bi-parental breeding population. This descriptive study highlights novel multi-omic QTL, including metabolism, photosynthesis as well as fruit setting, some of which are uniquely assigned under drought. Compared to conventional approaches using the bi-parental IL population, the study presented improves the resolution by assessing further backcrossed ILs, named sub-ILs.
In the final study, a photosynthetic gene, namely a PetM subunit of the cytochrome b6f complex encoding gene, involved in electron flow is characterized in an horticultural important crop. While several advances have been made in model organisms, this study highlights the transition of this fundamental knowledge to horticultural important crops, such as tomato, and investigates its function under differing light conditions. Overall, the presented thesis combines different strategies in unveiling the genetic components in multi-omic traits under drought using conventional breeding populations as well as a diverse global population. To this end, it allows a comparison of either approach and highlights their strengths and weaknesses.
The development of seeds in angiosperms starts with a complex process of double fertilization, involving the fusion of the maternal egg cell and central cell with two paternal sperm cells. This gives rise to the embryo and the nourishing endosperm, which are then enclosed by the seed coat, derived from the maternal integuments. The growth of the seed coat in Arabidopsis thaliana (Arabidopsis) is actively inhibited before fertilization by epigenetic regulators known as Polycomb Group (PcG) proteins. These proteins deposit a repressive histone mark called H3K27me3, which must be removed to enable seed coat formation. In this thesis, I explored the mechanism of removal of H3K27me3 marks from the integument cells following fertilization, which allows for seed coat formation. We hypothesized that this removal should be primarily facilitated by histone demethylases from the JMJ family and potentially influenced by the plant hormones Brassinosteroids (BRs). This hypothesis was supported by the expression patterns of the JMJ protein REF6 and of BR related genes, which are specifically expressed in the integuments and in the seed coat. Moreover, mutations in both these pathways lead to developmental defects, such as reduced ovule viability and delayed seed coat growth. Our research provides evidence suggesting that BR signalling is likely involved in recruiting JMJ-type histone demethylases to target loci responsible for seed coat growth. Moreover, we have discovered an additional pathway through which BRs regulate seed coat development, independent of their influence on H3K27me3 marks. This finding emphasizes the diverse roles of BRs in coordinating seed development, extending beyond their well-known involvement in plant growth and development. Furthermore, I explored the role of another epigenetic mark, DNA methylation, in fertilization-independent (or autonomous) seed formation in Arabidopsis. For this, we utilized epigenetic Recombinant Inbred Lines (epiRILs) and thus identified an epigenetic Quantitative Trait Locus (epiQTL) on chromosome II, potentially responsible for the larger autonomous seed size observed in DNA methylation mutants. Overall, this thesis significantly enhances our comprehension of the intricate relationship between epigenetic modifications, hormonal signaling, and plant reproductive processes. It offers valuable insights into the genetic mechanisms governing both sexual and asexual seed formation, while also presenting potential avenues for the engineer of advantageous traits in agricultural crops.