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Seed thermoinhibition, the repression of germination under high temperatures, prevents seedling establishment under potentially fatal conditions. Thermoinhibition is relevant for phenology and agriculture, particularly in a warming globe. The temperature sensing mechanisms and signaling pathways sustaining thermoinhibition are unknown. Here we show that thermoinhibition in Arabidopsis thaliana is not autonomously controlled by the embryo but is rather implemented by the endosperm. High temperature is sensed through endospermic phyB by accelerating its reversion from the active signaling Pfr form into the inactive Pr form, as previously described in seedlings. This leads to thermoinhibition mediated by PIFs, mainly PIF1, PIF3 and PIF5. Endospermic PIF3 represses the expression of the endospermic ABA catabolic gene CYP707A1 and promotes endospermic ABA accumulation and release towards the embryo to block its growth. Furthermore, endospermic ABA represses embryonic PIF3 accumulation that would otherwise promote embryonic growth. Hence, under high temperatures PIF3 exerts opposite growth responses in the endosperm and embryo., (© 2023. The Author(s).)

Seed dormancy-the absence of seed germination under favorable germination conditions-is a plant trait that evolved to enhance seedling survival by avoiding germination under unsuitable environmental conditions. In Arabidopsis , dormancy levels are influenced by the seed coat composition, while the endosperm is essential to repress seed germination of dormant seeds upon their imbibition. Recent research has shown that the mother plant modulates its progeny seed dormancy in response to seasonal temperature changes by changing specific aspects of seed coat and endosperm development. This process involves genomic imprinting by means of epigenetic marks deposited in the seed progeny and regulators previously known to regulate flowering time. This review discusses and summarizes these discoveries and provides an update on our present understanding of the role of DOG1 and abscisic acid, two key contributors to dormancy.

Seed dormancy is a widespread and key adaptive trait that is essential for the establishment of soil seed banks and prevention of pre-harvest sprouting. Herein we demonstrate that the endosperm-expressed transcription factors ZHOUPI (ZOU) and INDUCER OF CBF EXPRESSION1 (ICE1) play a role in determining the depth of primary dormancy in Arabidopsis. We show that ice1 or zou increases seed dormancy and the double mutant has an additive phenotype. This increased dormancy is associated with increased ABA levels, and can be separated genetically from any role in endosperm maturation because loss of ABA biosynthesis or DELAY OF GERMINATION 1 reverses the dormancy phenotype without affecting the aberrant seed morphology. Consistent with these results, ice1 endosperms had an increased capacity for preventing embryo greening, a phenotype previously associated with an increase in endospermic ABA levels. Although ice1 changes the expression of many genes, including some in ABA biosynthesis, catabolism and/or signalling, only ABA INSENSITIVE 3 is significantly misregulated in ice1 mutants. We also demonstrate that ICE1 binds to and inhibits expression of ABA INSENSITIVE 3. Our data demonstrate that Arabidopsis ICE1 and ZOU determine the depth of primary dormancy during maturation independently of their effect on endosperm development., (© 2018 The Authors. The Plant Journal published by John Wiley & Sons Ltd and Society for Experimental Biology.)

Examples of transgenerational transmission of environmentally induced epigenetic traits remain rare and disputed. Abiotic stress can release the transcription of epigenetically suppressed transposons and, noticeably, this activation is only transient. Therefore, it is likely that mechanisms countering the mitotic and meiotic inheritance of stress-triggered chromatin changes must exist but are undefined. To reveal these mechanisms, we screened for Arabidopsis mutants impaired in the resetting of stress-induced loss of epigenetic silencing and found that two chromatin regulators, Decrease in DNA methylation1 (DDM1) and Morpheus' Molecule1 (MOM1), act redundantly to restore prestress state and thus erase "epigenetic stress memory". In ddm1 mutants, stress hyperactivates heterochromatic transcription and transcription persists longer than in the wild type. However, this newly acquired state is not transmitted to the progeny. Strikingly, although stress-induced transcription in mom1 mutants is as rapidly silenced as in wild type, in ddm1 mom1 double mutants, transcriptional signatures of stress are able to persist and are found in the progeny of plants stressed as small seedlings. Our results reveal an important, previously unidentified function of DDM1 and MOM1 in rapid resetting of stress induced epigenetic states, and therefore also in preventing their mitotic propagation and transgenerational inheritance.

Leaf primordia are generated at the periphery of the shoot apex, developing into flat symmetric organs with adaxial-abaxial polarity, in which the indeterminate state is repressed. Despite the crucial role of the ASYMMETRIC LEAVES1 (AS1)-AS2 nuclear-protein complex in leaf adaxial-abaxial polarity specification, information on mechanisms controlling their downstream genes has remained elusive. We systematically analyzed transcripts by microarray and chromatin immunoprecipitation assays and performed genetic rescue of as1 and as2 phenotypic abnormalities, which identified a new target gene, ETTIN (ETT)/AUXIN RESPONSE FACTOR3 (ARF3), which encodes an abaxial factor acting downstream of the AS1-AS2 complex. While the AS1-AS2 complex represses ETT by direct binding of AS1 to the ETT promoter, it also indirectly activates miR390- and RDR6-dependent post-transcriptional gene silencing to negatively regulate both ETT and ARF4 activities. Furthermore, AS1-AS2 maintains the status of DNA methylation in the ETT coding region. In agreement, filamentous leaves formed in as1 and as2 plants treated with a DNA methylation inhibitor were rescued by loss of ETT and ARF4 activities. We suggest that negative transcriptional, post-transcriptional and epigenetic regulation of the ARFs by AS1-AS2 is important for stabilizing early leaf partitioning into abaxial and adaxial domains.

It is necessary to use algorithms to analyze gene expression data from DNA microarrays, such as in clustering and machine learning. Previously, we developed the knowledge-based fuzzy adaptive resonance theory (KB-FuzzyART), a clustering algorithm suitable for analyzing gene expression data, to find clues for identifying gene networks. Leaf primordia form around the shoot apical meristem (SAM), which consists of indeterminate stem cells. Upon initiation of leaf development, adaxial-abaxial patterning is crucial for lateral expansion, via cellular proliferation, and the formation of flat symmetric leaves. Many regulatory genes that specify such patterning have been identified. Analysis by the KB-FuzzyART and subsequent molecular and genetic analyses previously showed that ASYMMETRIC LEAVES1 (AS1) and AS2 repress the expression of some abaxial-determinant genes, such as AUXIN RESPONSE FACTOR3 (ARF3)/ETTIN (ETT) and ARF4, which are responsible for defects in leaf adaxial-abaxial polarity in as1 and as2. In the present study, genetic analysis revealed that ARF3/ETT and ARF4 were regulated by modifier genes, BOBBER1 (BOB1) and ELONGATA3 (ELO3), together with AS1-AS2. We analyzed expression arrays with as2 elo3 and as2 bob1, and extracted genes downstream of ARF3/ETT by using KB-FuzzyART and molecular analyses. The results showed that expression of Kip-related protein (KRP) (for inhibitors of cyclin-dependent protein kinases) and Isopentenyltransferase (IPT) (for biosynthesis of cytokinin) genes were controlled by AS1-AS2 through ARF3/ETT and ARF4 functions, which suggests that the AS1-AS2-ETT pathway plays a critical role in controlling the cell division cycle and the biosynthesis of cytokinin around SAM to stabilize leaf development in Arabidopsis thaliana.

Leaf primordia are generated around the shoot apical meristem. Mutation of the ASYMMETRIC LEAVES2 (AS2) gene of Arabidopsis thaliana results in defects in repression of the meristematic and indeterminate state, establishment of adaxial-abaxial polarity and left-right symmetry in leaves. AS2 represses transcription of meristem-specific class 1 KNOX homeobox genes and of the abaxial-determinant genes ETTIN/ARF3, KANADI2 and YABBY5. To clarify the role of AS2 in the establishment of leaf polarity, we isolated mutations that enhanced the polarity defects associated with as2. We describe here the enhancer-of-asymmetric-leaves-two1 (east1) mutation, which caused the formation of filamentous leaves with abaxialized epidermis on the as2-1 background. Levels of transcripts of class 1 KNOX and abaxial-determinant genes were markedly higher in as2-1 east1-1 mutant plants than in the wild-type and corresponding single-mutant plants. EAST1 encodes the histone acetyltransferase ELONGATA3 (ELO3), a component of the Elongator complex. Genetic analysis, using mutations in genes involved in the biogenesis of a trans-acting small interfering RNA (ta-siRNA), revealed that ELO3 mediated establishment of leaf polarity independently of AS2 and the ta-siRNA-related pathway. Treatment with an inhibitor of histone deacetylases (HDACs) caused additive polarity defects in as2-1 east1-1 mutant plants, suggesting the operation of an ELO3 pathway, independent of the HDAC pathway, in the determination of polarity. We propose that multiple pathways play important roles in repression of the expression of class 1 KNOX and abaxial-determinant genes in the development of the adaxial domain of leaves and, thus, in the establishment of leaf polarity.

Viruses induce pathogenic symptoms on plants but the molecular basis is poorly understood. Here, we show that transgenic Arabidopsis expressing the pathogenesis protein betaC1 of Tomato yellow leaf curl China virus (TYLCCNV), a geminivirus, can phenocopy to a large extent disease symptoms of virus-infected tobacco plants in having upward curled leaves, radialized leaves with outgrowth tissues from abaxial surfaces, and sterile flowers. These morphological changes are paralleled by a reduction in miR165/166 levels and an increase in PHB and PHV transcript levels. Two factors, ASYMMETRIC LEAVES 1 (AS1) and ASYMMETRIC LEAVES 2 (AS2), are known to regulate leaf development as AS1/AS2 complex. Strikingly, betaC1 plants phenocopy plants overexpressing AS2 at the morphological and molecular level and betaC1 is able to partially complement as2 mutation. betaC1 binds directly to AS1, elicits morphological and gene expression changes dependent on AS1 but not AS2, and attenuates expression of selective jasmonic acid (JA)-responsive gene. Our results show that betaC1 forms a complex with AS1 to execute its pathogenic functions and to suppress a subset of JA responses.

The ASYMMETRIC LEAVES2 (AS2) gene, a member of the AS2/LOB gene family, and the ASYMMETRIC LEAVES1 (AS1) gene of Arabidopsis thaliana participate in the development of a symmetrical, expanded lamina. We report here the patterns of expression of these genes, and the importance of the sites of such expression in leaf development. Transcripts of both genes accumulated in the entire leaf primordia at early stages, but the patterns of accumulation changed as the leaves expanded. AS2 and AS1 transcripts were detected, respectively, in the adaxial domain and in the inner domain between the adaxial and abaxial domains of leaves. The ratios of numbers of adaxial cells to abaxial cells in cotyledons of corresponding mutant lines were greater than the ratios in wild-type cotyledons. The low levels of ectopic expression of AS2 under the control of the AS1 promoter in as2 mutant plants restored an almost normal phenotype in some cases, but also resulted in flatter leaves than those of wild-type plants. Strong expression of the construct in wild-type and as2 plants, but not as1 plants, resulted in the formation of narrow, upwardly curled leaves. Our results indicate that AS2 represses cell proliferation in the adaxial domain in the presence of AS1, and that adaxial expression of AS2 at an appropriate level is critical for the development of a symmetrical, expanded lamina. Real-time RT-PCR analysis revealed that mutation of either AS2 or AS1 resulted in an increase in the levels of transcripts of ETTIN (ETT; also known as AUXIN RESPONSE FACTOR3, ARF3) and KANADI2 (KAN2), which are abaxial determinants, and YABBY5 (YAB5). Thus, AS2 and AS1 might negatively regulate the expression of these genes in the adaxial domain, which might be related to the development of flat and expanded leaves.