The PUB4 E3 Ubiquitin Ligase Is Responsible for the Variegated Phenotype Observed upon Alteration of Chloroplast Protein Homeostasis in Arabidopsis Cotyledons.

During a plant's life cycle, plastids undergo several modifications, from undifferentiated pro-plastids to either photosynthetically-active chloroplasts, ezioplasts, chromoplasts or storage organelles, such as amyloplasts, elaioplasts and proteinoplasts. Plastid proteome rearrangements and protein homeostasis, together with intracellular communication pathways, are key factors for correct plastid differentiation and functioning. When plastid development is affected, aberrant organelles are degraded and recycled in a process that involves plastid protein ubiquitination. In this study, we have analysed the Arabidopsis gun1-102 ftsh5-3 double mutant, lacking both the plastid-located protein GUN1 (Genomes Uncoupled 1), involved in plastid-to-nucleus communication, and the chloroplast-located FTSH5 (Filamentous temperature-sensitive H5), a metalloprotease with a role in photosystem repair and chloroplast biogenesis. gun1-102 ftsh5-3 seedlings show variegated cotyledons and true leaves that we attempted to suppress by introgressing second-site mutations in genes involved in: (i) plastid translation, (ii) plastid folding/import and (iii) cytosolic protein ubiquitination. Different phenotypic effects, ranging from seedling-lethality to partial or complete suppression of the variegated phenotype, were observed in the corresponding triple mutants. Our findings indicate that Plant U-Box 4 (PUB4) E3 ubiquitin ligase plays a major role in the target degradation of damaged chloroplasts and is the main contributor to the variegated phenotype observed in gun1-102 ftsh5-3 seedlings.

Genetic Interaction of SEEDSTICK, GORDITA and AUXIN RESPONSE FACTOR 2 during Seed Development.

Seed development is under the control of complex and coordinated molecular networks required for the formation of its different components. The seed coat development largely determines final seed size and shape, in addition to playing a crucial role in protecting the embryo and promoting germination. In this study, we investigated the role of three transcription factors known to be active during seed development in Arabidopsis thaliana: SEEDSTICK (STK) and GORDITA (GOA), two MADS-domain proteins, and AUXIN RESPONSE FACTOR 2 (ARF2), belonging to the ARF family. Through a reverse genetic approach, we characterized the seed phenotypes of all the single, double and triple loss-of-function mutants in relation to seed size/shape and the effects on metabolic pathways occurring in the seed coat. This approach revealed that dynamic networks involving these TFs are active throughout ovule and seed development, affecting the formation of the seed coat. Notably, while the genetic interaction among these genes results in synergies that control the promotion of cell expansion in the seed coat upon pollination and production of proanthocyanidins, functional antagonists arise in the control of cell proliferation and release of mucilage.

The Arabidopsis MADS-Domain Transcription Factor SEEDSTICK Controls Seed Size via Direct Activation of E2Fa.

Seed size is the result of complex molecular networks controlling the development of the seed coat (of maternal origin) and the two fertilization products, the embryo and the endosperm. In this study we characterized the role of Arabidopsis thaliana MADS-domain transcription factor SEEDSTICK (STK) in seed size control. STK is known to regulate the differentiation of the seed coat as well as the structural and mechanical properties of cell walls in developing seeds. In particular, we further characterized stk mutant seeds. Genetic evidence (reciprocal crosses) of the inheritance of the small-seed phenotype, together with the provided analysis of cell division activity (flow cytometry), demonstrate that STK acts in the earlier phases of seed development as a maternal activator of growth. Moreover, we describe a molecular mechanism underlying this activity by reporting how STK positively regulates cell cycle progression via directly activating the expression of E2Fa, a key regulator of the cell cycle. Altogether, our results unveil a new genetic network active in the maternal control of seed size in Arabidopsis.

Barley's Second Spring as A Model Organism for Chloroplast Research.

Barley (Hordeum vulgare) has been widely used as a model crop for studying molecular and physiological processes such as chloroplast development and photosynthesis. During the second half of the 20th century, mutants such as albostrians led to the discovery of the nuclear-encoded, plastid-localized RNA polymerase and the retrograde (chloroplast-to-nucleus) signalling communication pathway, while chlorina-f2 and xantha mutants helped to shed light on the chlorophyll biosynthetic pathway, on the light-harvesting proteins and on the organization of the photosynthetic apparatus. However, during the last 30 years, a large fraction of chloroplast research has switched to the more "user-friendly" model species Arabidopsis thaliana, the first plant species whose genome was sequenced and published at the end of 2000. Despite its many advantages, Arabidopsis has some important limitations compared to barley, including the lack of a real canopy and the absence of the proplastid-to-chloroplast developmental gradient across the leaf blade. These features, together with the availability of large collections of natural genetic diversity and mutant populations for barley, a complete genome assembly and protocols for genetic transformation and gene editing, have relaunched barley as an ideal model species for chloroplast research. In this review, we provide an update on the genomics tools now available for barley, and review the biotechnological strategies reported to increase photosynthesis efficiency in model species, which deserve to be validated in barley.

Time-Course Transcriptome Analysis of Arabidopsis Siliques Discloses Genes Essential for Fruit Development and Maturation.

Fruits protect the developing seeds of angiosperms and actively contribute to seed dispersion. Furthermore, fruit and seed development are highly synchronized and require exchange of information between the mother plant and the developing generations. To explore the mechanisms controlling fruit formation and maturation, we performed a transcriptomic analysis on the valve tissue of the Arabidopsis (Arabidopsis thaliana) silique using RNA sequencing. In doing so, we have generated a data set of differentially regulated genes that will help to elucidate the molecular mechanisms that underpin the initial phase of fruit growth and, subsequently, trigger fruit maturation. The robustness of our data set has been tested by functional genomic studies. Using a reverse genetics approach, we selected 10 differentially expressed genes and explored the consequences of their disruption for both silique growth and senescence. We found that genes contained in our data set play essential roles in different stages of silique development and maturation, indicating that our transcriptome-based gene list is a powerful tool for the elucidation of the molecular mechanisms controlling fruit formation in Arabidopsis.