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Embryo development in Arabidopsis (Arabidopsis thaliana) starts off with an asymmetric division of the zygote to generate the precursors of the embryo proper and the supporting extraembryonic suspensor. The suspensor degenerates as the development of the embryo proper proceeds beyond the heart stage. Until the globular stage, the suspensor maintains embryonic potential and can form embryos in the absence of the developing embryo proper. We report a mutant called meerling-1 (mrl-1), which shows a high penetrance of suspensor-derived polyembryony due to delayed development of the embryo proper. Eventually, embryos from both apical and suspensor lineages successfully develop into normal plants and complete their life cycle. We identified the causal mutation as a genomic rearrangement altering the promoter of the Arabidopsis U3 SMALL NUCLEOLAR RNA-ASSOCIATED PROTEIN 18 (UTP18) homolog that encodes a nucleolar-localized WD40-repeat protein involved in processing 18S preribosomal RNA. Accordingly, root-specific knockout of UTP18 caused growth arrest and accumulation of unprocessed 18S pre-rRNA. We generated the mrl-2 loss-of-function mutant and observed asynchronous megagametophyte development causing embryo sac abortion. Together, our results indicate that promoter rearrangement decreased UTP18 protein abundance during early stage embryo proper development, triggering suspensor-derived embryogenesis. Our data support the existence of noncell autonomous signaling from the embryo proper to prevent direct reprogramming of the suspensor toward embryonic fate. Arabidopsis meerling-1 mutant exhibits suspensor-derived polyembryony due to delayed processing of the 18S preribosomal RNA, resulting in reprogramming of the suspensor toward embryonic fate. [ABSTRACT FROM AUTHOR]

Transcription termination has important regulatory functions, impacting mRNA stability, localization and translation potential. Failure to appropriately terminate transcription can also lead to read-through transcription and the synthesis of antisense RNAs which can have profound impact on gene expression. The Transcription-Export (THO/TREX) protein complex plays an important role in coupling transcription with splicing and export of mRNA. However, little is known about the role of the THO/TREX complex in the control of transcription termination. In this work, we show that two proteins of the THO/TREX complex, namely TREX COMPONENT 1 (TEX1 or THO3) and HYPER RECOMBINATION1 (HPR1 or THO1) contribute to the correct transcription termination at several loci in Arabidopsis thaliana. We first demonstrate this by showing defective termination in tex1 and hpr1 mutants at the nopaline synthase (NOS) terminator present in a T-DNA inserted between exon 1 and 3 of the PHO1 locus in the pho1-7 mutant. Read-through transcription beyond the NOS terminator and splicing-out of the T-DNA resulted in the generation of a near full-length PHO1 mRNA (minus exon 2) in the tex1 pho1-7 and hpr1 pho1-7 double mutants, with enhanced production of a truncated PHO1 protein that retained phosphate export activity. Consequently, the strong reduction of shoot growth associated with the severe phosphate deficiency of the pho1-7 mutant was alleviated in the tex1 pho1-7 and hpr1 pho1-7 double mutants. Additionally, we show that RNA termination defects in tex1 and hpr1 mutants leads to 3'UTR extensions in several plant genes. These results demonstrate that THO/TREX complex contributes to the regulation of transcription termination. Author summary: Production of messenger RNAs (mRNAs) involves numerous steps including initiation of transcription, elongation, splicing, termination, as well as export out of the nucleus. All these steps are highly coordinated and failure in any steps has a profound impact on the level and identity of mRNAs produced. The THO/TREX protein complex is associated with nascent RNAs and contributes to several mRNA biogenesis steps, including splicing and export. However, the contribution of the THO/TREX complex to mRNA termination was poorly defined. We have identified a role for two components of the THO/TREX complex, namely the proteins TEX1 and HPR1, in the control of transcription termination in the plant Arabidopsis thaliana. We show that the tex1 and hpr1 mutants have defects in terminating mRNA at the nopaline synthase (NOS) terminator found in a T-DNA insertion mutant leading to the transcriptional read-through pass the NOS terminator. We also show that tex1 and hpr1 mutants have defects in mRNA termination at several endogenous genes, leading to the production of 3'UTR extensions. Together, these results highlight a role for the THO/TREX complex in mRNA termination. [ABSTRACT FROM AUTHOR]

Structural variants (SVs) are an important class of genetic variation implicated in a wide array of genetic diseases including cancer. Despite the advances in whole genome sequencing, comprehensive and accurate detection of SVs in short-read data still poses some practical and computational challenges. We present sv-callers, a highly portable workflow that enables parallel execution of multiple SV detection tools, as well as provide users with example analyses of detected SV callsets in a Jupyter Notebook. This workflow supports easy deployment of software dependencies, configuration and addition of new analysis tools. Moreover, porting it to different computing systems requires minimal effort. Finally, we demonstrate the utility of the workflow by performing both somatic and germline SV analyses on different high-performance computing systems. [ABSTRACT FROM AUTHOR]

Membrane traffic at the trans-Golgi network (TGN) is crucial for correctly distributing various membrane proteins to their destination. Polarly localized auxin efflux proteins, including PIN-FORMED1 (PIN1), are dynamically transported between the endosomes and the plasma membrane (PM) in the plant cells. The intracellular trafficking of PIN1 protein is sensitive to the fungal toxin brefeldin A (BFA), which is known to inhibit guanine nucleotide exchange factors for ADP ribosylation factors (ARF GEFs) such as GNOM. However, the molecular details of the BFA-sensitive trafficking pathway have not been fully revealed. In a previous study, we identified an Arabidopsis mutant BFA-visualized endocytic trafficking defective 3 (ben3) which exhibited reduced sensitivity to BFA in terms of BFA-induced intracellular PIN1 agglomeration. Here, we show that BEN3 encodes a member of BIG family ARF GEFs, BIG2. BEN3/BIG2 tagged with fluorescent proteins co-localized with markers for the TGN/early endosome (EE). Inspection of conditionally induced de novo synthesized PIN1 confirmed that its secretion to the PM is BFA sensitive, and established BEN3/BIG2 as a crucial component of this BFA action at the level of the TGN/EE. Furthermore, ben3 mutation alleviated BFAinduced agglomeration of another TGN-localized ARF GEF, BEN1/MIN7. Taken together, our results suggest that BEN3/ BIG2 is an ARF GEF component, which confers BFA sensitivity to the TGN/EE in Arabidopsis. [ABSTRACT FROM AUTHOR]

Organ formation in animals and plants relies on precise control of cell state transitions to turn stem cell daughters into fully differentiated cells. In plants, cells cannot rearrange due to shared cell walls. Thus, differentiation progression and the accompanying cell expansion must be tightly coordinated across tissues. PLETHORA (PLT) transcription factor gradients are unique in their ability to guide the progression of cell differentiation at different positions in the growing Arabidopsis thaliana root, which contrasts with well-described transcription factor gradients in animals specifying distinct cell fates within an essentially static context. To understand the output of the PLT gradient, we studied the gene set transcriptionally controlled by PLTs. Our work reveals how the PLT gradient can regulate cell state by region-specific induction of cell proliferation genes and repression of differentiation. Moreover, PLT targets include major patterning genes and autoregulatory feedback components, enforcing their role as master regulators of organ development. [ABSTRACT FROM AUTHOR]

Natural genetic variation is crucial for adaptability of plants to different environments. Seed dormancy prevents precocious germination in unsuitable conditions and is an adaptation to a major macro-environmental parameter, the seasonal variation in temperature and day length., Here we report the isolation of IBO, a quantitative trait locus ( QTL) that governs c. 30% of germination rate variance in an Arabidopsis recombinant inbred line ( RIL) population derived from the parental accessions Eilenburg-0 (Eil-0) and Loch Ness-0 (Lc-0)., IBO encodes an uncharacterized phosphatase 2C-related protein, but neither the Eil-0 nor the Lc-0 variant, which differ in a single amino acid, have any appreciable phosphatase activity in in vitro assays. However, we found that the amino acid change in the Lc-0 variant of the IBO protein confers reduced germination rate. Moreover, unlike the Eil-0 variant of the protein, the Lc-0 variant can interfere with the activity of the phosphatase 2C ABSCISIC ACID INSENSITIVE 1 in vitro., This suggests that the Lc-0 variant possibly interferes with abscisic acid signaling, a notion that is supported by physiological assays. Thus, we isolated an example of a QTL allele with a nonsynonymous amino acid change that might mediate local adaptation of seed germination timing. [ABSTRACT FROM AUTHOR]

Eukaryotic mRNA transcription and turnover is controlled by an enzymatic machinery that includes RNA polymerase II and the 3' to 5' exosome. The activity of these protein complexes is modulated by additional factors, such as the nuclear RNA polymerase II-associated factor 1 (Paf1c) and the cytoplasmic Superkiller (SKI) complex, respectively. Their components are conserved across uni- as well as multi-cellular organisms, including yeast, Arabidopsis, and humans. Among them, SKI8 displays multiple facets on top of its cytoplasmic role in the SKI complex. For instance, nuclear yeast ScSKI8 has an additional function in meiotic recombination, whereas nuclear human hSKI8 (unlike ScSKI8) associates with Paf1c. The Arabidopsis SKI8 homolog VERNALIZATION INDEPENDENT 3 (VIP3) has been found in Paf1c as well; however, whether it also has a role in the SKI complex remains obscure so far. We found that transgenic VIP3-GFP, which complements a novel vip3 mutant allele, localizes to both nucleus and cytoplasm. Consistently, biochemical analyses suggest that VIP3-GFP associates with the SKI complex. A role of VIP3 in the turnover of nuclear encoded mRNAs is supported by random-primed RNA sequencing of wild-type and vip3 seedlings, which indicates mRNA stabilization in vip3. Another SKI subunit homolog mutant, ski2, displays a dwarf phenotype similar to vip3. However, unlike vip3, it displays neither early flowering nor flower development phenotypes, suggesting that the latter reflect VIP3's role in Paf1c. Surprisingly then, transgenic ScSKI8 rescued all aspects of the vip3 phenotype, suggesting that the dual role of SKI8 depends on species-specific cellular context. [ABSTRACT FROM AUTHOR]

Casparian strips are precisely localized and aligned ring-like cell wall modifications in the root of all higher plants. They set up an extracellular diffusion barrier analogous to animal tight junctions, and are crucial for maintaining the homeostatic capacity of plant roots. Casparian strips become localized because of the formation of a highly stable plasma membrane domain, consisting of a family of small transmembrane proteins called Casparian strip membrane domain proteins (CASPs). Here we report a large-scale forward genetic screen directly visualizing endodermal barrier function, which allowed us to identify factors required for the formation and integrity of Casparian strips. We present the identification and characterization of one of the mutants, schengen1 (sgn1), a receptor-like cytoplasmic kinase that we show localizes in a strictly polar fashion to the outer plasma membrane of endodermal cells and is required for the positioning and correct formation of the centrally located CASP domain.

Ultra-high-throughput sequencing (UHTS) techniques are evolving rapidly and may soon become an affordable and routine tool for sequencing plant DNA, even in smaller plant biology labs. Here we review recent insights into intraspecific genome variation gained from UHTS, which offers a glimpse of the rather unexpected levels of structural variability among Arabidopsis thaliana accessions. The challenges that will need to be addressed to efficiently assemble and exploit this information are also discussed.

Identification of small polymorphisms from next generation sequencing short read data is relatively easy, but detection of larger deletions is less straightforward. Here, we analyzed four divergent Arabidopsis accessions and found that intersection of absent short read coverage with weak tiling array hybridization signal reliably flags deletions. Interestingly, individual deletions were frequently observed in two or more of the accessions examined, suggesting that variation in gene content partly reflects a common history of deletion events.