Dormancy heterogeneity among Arabidopsis thaliana seeds is linked to individual seed size.

Production of morphologically and physiologically variable seeds is an important strategy that helps plants to survive in unpredictable natural conditions. However, the model plant Arabidopsis thaliana and most agronomically essential crops produce visually homogenous seeds. Using automated phenotype analysis, we observed that small seeds in Arabidopsis tend to have higher primary and secondary dormancy levels than large seeds. Transcriptomic analysis revealed distinct gene expression profiles between large and small seeds. Large seeds have higher expression of translation-related genes implicated in germination competence. By contrast, small seeds have elevated expression of many positive regulators of dormancy, including a key regulator of this process, the DOG1 gene. Differences in DOG1 expression are associated with differential production of its alternative cleavage and polyadenylation isoforms; in small seeds, the proximal poly(A) site is selected, resulting in a short mRNA isoform. Furthermore, single-seed RNA sequencing analysis demonstrated that large seeds resemble DOG1 knockout mutant seeds. Finally, on the single-seed level, expression of genes affected by seed size is correlated with expression of genes that position seeds on the path toward germination. Our results demonstrate an unexpected link between seed size and dormancy phenotypes in a species that produces highly homogenous seed pools, suggesting that the correlation between seed morphology and physiology is more widespread than initially assumed.

A non-canonical function of Arabidopsis ERECTA proteins and a role of the SWI3B subunit of the SWI/SNF chromatin remodeling complex in gibberellin signaling.

The Arabidopsis ERECTA family (ERf) of leucine-rich repeat receptor-like kinases (LRR-RLKs) comprising ERECTA (ER), ERECTA-LIKE 1 (ERL1), and ERECTA-LIKE 2 (ERL2) controls epidermal patterning, inflorescence architecture, and stomata development and patterning. These proteins are reported to be plasma membrane associated. Here we show that the er/erl1/erl2 mutant exhibits impaired gibberellin (GA) biosynthesis and perception alongside broad transcriptional changes. The ERf kinase domains were found to localize to the nucleus where they interact with the SWI3B subunit of the SWI/SNF chromatin remodeling complex (CRCs). The er/erl1/erl2 mutant exhibits reduced SWI3B protein level and affected nucleosomal chromatin structure. Similar to swi3c and brm plants with inactivated subunits of SWI/SNF CRCs, it also does not accumulate DELLA RGA and GAI proteins. The ER kinase phosphorylates SWI3B in vitro, and the inactivation of all ERf proteins leads to the decreased phosphorylation of SWI3B protein in vivo. The identified correlation between DELLA overaccumulation and SWI3B proteasomal degradation, and the physical interaction of SWI3B with DELLA proteins indicate an important role of SWI3B-containing SWI/SNF CRCs in gibberellin signaling. Co-localization of ER and SWI3B on GID1 (GIBBERELLIN INSENSITIVE DWARF 1) DELLA target gene promoter regions and abolished SWI3B binding to GID1 promoters in er/erl1/erl2 plants supports the conclusion that ERf-SWI/SNF CRC interaction is important for transcriptional control of GA receptors. Thus, the involvement of ERf proteins in the transcriptional control of gene expression, and observed similar features for human HER2 (epidermal growth family receptor member), indicate an exciting target for further studies of evolutionarily conserved non-canonical functions of eukaryotic membrane receptors.

Promoter-pervasive transcription causes RNA polymerase II pausing to boost DOG1 expression in response to salt.

Eukaryotic genomes are pervasively transcribed by RNA polymerase II. Yet, the molecular and biological implications of such a phenomenon are still largely puzzling. Here, we describe noncoding RNA transcription upstream of the Arabidopsis thaliana DOG1 gene, which governs salt stress responses and is a key regulator of seed dormancy. We find that expression of the DOG1 gene is induced by salt stress, thereby causing a delay in seed germination. We uncover extensive transcriptional activity on the promoter of the DOG1 gene, which produces a variety of lncRNAs. These lncRNAs, named PUPPIES, are co-directionally transcribed and extend into the DOG1 coding region. We show that PUPPIES RNAs respond to salt stress and boost DOG1 expression, resulting in delayed germination. This positive role of pervasive PUPPIES transcription on DOG1 gene expression is associated with augmented pausing of RNA polymerase II, slower transcription and higher transcriptional burst size. These findings highlight the positive role of upstream co-directional transcription in controlling transcriptional dynamics of downstream genes.

Single seeds exhibit transcriptional heterogeneity during secondary dormancy induction.

Seeds are highly resilient to the external environment, which allows plants to persist in unpredictable and unfavorable conditions. Some plant species have adopted a bet-hedging strategy to germinate a variable fraction of seeds in any given condition, and this could be explained by population-based threshold models. Here, in the model plant Arabidopsis (Arabidopsis thaliana), we induced secondary dormancy (SD) to address the transcriptional heterogeneity among seeds that leads to binary germination/nongermination outcomes. We developed a single-seed RNA-seq strategy that allowed us to observe a reduction in seed transcriptional heterogeneity as seeds enter stress conditions, followed by an increase during recovery. We identified groups of genes whose expression showed a specific pattern through a time course and used these groups to position the individual seeds along the transcriptional gradient of germination competence. In agreement, transcriptomes of dormancy-deficient seeds (mutant of DELAY OF GERMINATION 1) showed a shift toward higher values of the germination competence index. Interestingly, a significant fraction of genes with variable expression encoded translation-related factors. In summary, interrogating hundreds of single-seed transcriptomes during SD-inducing treatment revealed variability among the transcriptomes that could result from the distribution of population-based sensitivity thresholds. Our results also showed that single-seed RNA-seq is the method of choice for analyzing seed bet-hedging-related phenomena.

[Mechanisms controlling chromatin structure]. / Mechanizmy kontrolujace strukture chromatyny.

Genomic DNA is highly packaged in eukaryotic cells and occurs in the form of nucleoprotein complex called chromatin. Although high DNA compaction allows to store large amount of genomic information in the cell nuclei, it also restricts the access to DNA regulatory sequences. Therefore, to overcome this issue, chromatin must be subjected to various alterations which are dependent on few interrelated factors: DNA modification, histones variants and modifications, ncRNA, chromatin remodeling complexes and chromatin architecture in nuclei. They allow to multilayer regulation of fragile balance between transcriptionally active euchromatin and inactive heterochromatin. The newest research describe new chromatin elements, e.g. half nucleosomes, bivalent chromatin marker and pointed to few intermediate states between euchromatin and heterochromatin. Variety and remarkable amount of chromatin modifications require existence of multiprotein complexes reading, editing and integrating genomic information. Some of them are able to remodel nucleosomes in order to control access to particular DNA sequence. Due to the complexity of chromatin structure regulation studies describing these mechanisms are fundamental to understanding the eukaryotes life.

The Role of SWI/SNF Chromatin Remodeling Complexes in Hormone Crosstalk.

SWI/SNF-type ATP-dependent chromatin remodeling complexes (CRCs) are evolutionarily conserved multiprotein machineries controlling DNA accessibility by regulating chromatin structure. We summarize here recent advances highlighting the role of SWI/SNF in the regulation of hormone signaling pathways and their crosstalk in Arabidopsis thaliana. We discuss the functional interdependences of SWI/SNF complexes and key elements regulating developmental and hormone signaling pathways by indicating intriguing similarities and differences in plants and humans, and summarize proposed mechanisms of SWI/SNF action on target loci. We postulate that, given their viability, several plant SWI/SNF mutants may serve as an attractive model for searching for conserved functions of SWI/SNF CRCs in hormone signaling, cell cycle control, and other regulatory pathways.

SWP73 Subunits of Arabidopsis SWI/SNF Chromatin Remodeling Complexes Play Distinct Roles in Leaf and Flower Development.

Arabidopsis thaliana SWP73A and SWP73B are homologs of mammalian BRAHMA-associated factors (BAF60s) that tether SWITCH/SUCROSE NONFERMENTING chromatin remodeling complexes to transcription factors of genes regulating various cell differentiation pathways. Here, we show that Arabidopsis thaliana SWP73s modulate several important developmental pathways. While undergoing normal vegetative development, swp73a mutants display reduced expression of FLOWERING LOCUS C and early flowering in short days. By contrast, swp73b mutants are characterized by retarded growth, severe defects in leaf and flower development, delayed flowering, and male sterility. MNase-Seq, transcript profiling, and ChIP-Seq studies demonstrate that SWP73B binds the promoters of ASYMMETRIC LEAVES1 and 2, KANADI1 and 3, and YABBY2, 3, and 5 genes, which regulate leaf development and show coordinately altered transcription in swp73b plants. Lack of SWP73B alters the expression patterns of APETALA1, APETALA3, and the MADS box gene AGL24, whereas other floral organ identity genes show reduced expression correlating with defects in flower development. Consistently, SWP73B binds to the promoter regions of APETALA1 and 3, SEPALLATA3, LEAFY, UNUSUAL FLORAL ORGANS, TERMINAL FLOWER1, AGAMOUS-LIKE24, and SUPPRESSOR OF CONSTANS OVEREXPRESSION1 genes, and the swp73b mutation alters nucleosome occupancy on most of these loci. In conclusion, SWP73B acts as important modulator of major developmental pathways, while SWP73A functions in flowering time control.

DELLA-interacting SWI3C core subunit of switch/sucrose nonfermenting chromatin remodeling complex modulates gibberellin responses and hormonal cross talk in Arabidopsis.

Switch (SWI)/Sucrose Nonfermenting (SNF)-type chromatin-remodeling complexes (CRCs) are involved in regulation of transcription, DNA replication and repair, and cell cycle. Mutations of conserved subunits of plant CRCs severely impair growth and development; however, the underlying causes of these phenotypes are largely unknown. Here, we show that inactivation of SWI3C, the core component of Arabidopsis (Arabidopsis thaliana) SWI/SNF CRCs, interferes with normal functioning of several plant hormone pathways and alters transcriptional regulation of key genes of gibberellin (GA) biosynthesis. The resulting reduction of GA4 causes severe inhibition of hypocotyl and root elongation, which can be rescued by exogenous GA treatment. In addition, the swi3c mutation inhibits DELLA-dependent transcriptional activation of GIBBERELLIN-INSENSITIVE DWARF1 (GID1) GA receptor genes. Down-regulation of GID1a in parallel with the DELLA repressor gene REPRESSOR OF GA1-3 1 in swi3c indicates that lack of SWI3C also leads to defects in GA signaling. Together with the recent demonstration of function of SWI/SNF ATPase BRAHMA in the GA pathway, these results reveal a critical role of SWI/SNF CRC in the regulation of GA biosynthesis and signaling. Moreover, we demonstrate that SWI3C is capable of in vitro binding to, and shows in vivo bimolecular fluorescence complementation interaction in cell nuclei with, the DELLA proteins RGA-LIKE2 and RGA-LIKE3, which affect transcriptional activation of GID1 and GA3ox (GIBBERELLIN 3-OXIDASE) genes controlling GA perception and biosynthesis, respectively. Furthermore, we show that SWI3C also interacts with the O-GlcNAc (O-linked N-acetylglucosamine) transferase SPINDLY required for proper functioning of DELLAs and acts hypostatically to (SPINDLY) in the GA response pathway. These findings suggest that DELLA-mediated effects in GA signaling as well as their role as a hub in hormonal cross talk may be, at least in part, dependent on their direct physical interaction with complexes responsible for modulation of chromatin structure.