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SUMMARY: The genomic integrity of every organism is endangered by various intrinsic and extrinsic stresses. To maintain genomic integrity, a sophisticated DNA damage response (DDR) network is activated rapidly after DNA damage. Notably, the fundamental DDR mechanisms are conserved in eukaryotes. However, knowledge about many regulatory aspects of the plant DDR is still limited. Important, yet little understood, regulatory factors of the DDR are the long non‐coding RNAs (lncRNAs). In humans, 13 lncRNAs functioning in DDR have been characterized to date, whereas no such lncRNAs have been characterized in plants yet. By meta‐analysis, we identified the putative long intergenic non‐coding RNA induced by DNA damage (LINDA) that responds strongly to various DNA double‐strand break‐inducing treatments, but not to replication stress induced by mitomycin C. After DNA damage, LINDA is rapidly induced in an ATM‐ and SOG1‐dependent manner. Intriguingly, the transcriptional response of LINDA to DNA damage is similar to that of its flanking hypothetical protein‐encoding gene. Phylogenetic analysis of putative Brassicales and Malvales LINDA homologs indicates that LINDA lncRNAs originate from duplication of a flanking small protein‐encoding gene followed by pseudogenization. We demonstrate that LINDA is not only needed for the regulation of this flanking gene but also fine‐tuning of the DDR after the occurrence of DNA double‐strand breaks. Moreover, Δlinda mutant root stem cells are unable to recover from DNA damage, most likely due to hyper‐induced cell death. Significant Statement: We unraveled the functional relevance of the first putative lncRNA within the DNA damage response network of Arabidopsis thaliana. This putative lncRNA, termed LINDA, is an important part of the DNA damage response network, as it is needed for accurate regulation of cell death and cell cycle progression. [ABSTRACT FROM AUTHOR]

Survival of living organisms is fully dependent on their maintenance of genome integrity, being permanently threatened by replication stress in proliferating cells. Although the plant DNA damage response (DDR) regulator SOG1 has been demonstrated to cope with replication defects, accumulating evidence points to other pathways functioning independent of SOG1. Here, we report the roles of the Arabidopsis E2FA and EF2B transcription factors, two well-characterized regulators of DNA replication, in plant response to replication stress. Through a combination of reverse genetics and chromatin immunoprecipitation approaches, we show that E2FA and E2FB share many target genes with SOG1, providing evidence for their involvement in the DDR. Analysis of double- and triple-mutant combinations revealed that E2FB, rather than E2FA, plays the most prominent role in sustaining plant growth in the presence of replication defects, either operating antagonistically or synergistically with SOG1. Conversely, SOG1 aids in overcoming the replication defects of E2FA/E2FB-deficient plants. Collectively, our data reveal a complex transcriptional network controlling the replication stress response in which E2Fs and SOG1 act as key regulatory factors. This study investigates the respective roles of E2FA and E2FB transcription factors during plant response to replication stress. The results show that E2FA and E2FB function together with SOG1 to control the expression of DDR genes and that E2FB plays a unique role in allowing cell cycle progression from G2/M despite replication stress. [ABSTRACT FROM AUTHOR]

Plant roots possess remarkable regenerative potential owing to their ability to replenish damaged or lost stem cells. ETHYLENE RESPONSE FACTOR 115 (ERF115), one of the key molecular elements linked to this potential, plays a predominant role in the activation of regenerative cell divisions. However, the downstream operating molecular machinery driving wound-activated cell division is largely unknown. Here, we biochemically and genetically identified the GRAS-domain transcription factor SCARECROW-LIKE 5 (SCL5) as an interaction partner of ERF115 in Arabidopsis thaliana. Although nonessential under control growth conditions, SCL5 acts redundantly with the related PHYTOCHROME A SIGNAL TRANSDUCTION 1 (PAT1) and SCL21 transcription factors to activate the expression of the DNA-BINDING ONE FINGER 3.4 (DOF3.4) transcription factor gene. DOF3.4 expression is wound-inducible in an ERF115-dependent manner and, in turn, activates D3-type cyclin expression. Accordingly, ectopic DOF3.4 expression drives periclinal cell division, while its downstream D3-type cyclins are essential for the regeneration of a damaged root. Our data highlight the importance and redundant roles of the SCL5, SCL21, and PAT1 transcription factors in wound-activated regeneration processes and pinpoint DOF3.4 as a key downstream element driving regenerative cell division. [ABSTRACT FROM AUTHOR]

The anaphase-promoting complex/cyclosome (APC/C) marks key cell cycle proteins for proteasomal breakdown, thereby ensuring unidirectional progression through the cell cycle. Its target recognition is temporally regulated by activating subunits, one of which is called CELL CYCLE SWITCH 52 A2 (CCS52A2). We sought to expand the knowledge on the APC/C by using the severe growth phenotypes of CCS52A2-deficient Arabidopsis (Arabidopsis thaliana) plants as a readout in a suppressor mutagenesis screen, resulting in the identification of the previously undescribed gene called PIKMIN1 (PKN1). PKN1 deficiency rescues the disorganized root stem cell phenotype of the ccs52a2-1 mutant, whereas an excess of PKN1 inhibits the growth of ccs52a2-1 plants, indicating the need for control of PKN1 abundance for proper development. Accordingly, the lack of PKN1 in a wild-type background negatively impacts cell division, while its systemic overexpression promotes proliferation. PKN1 shows a cell cycle phase-dependent accumulation pattern, localizing to microtubular structures, including the preprophase band, the mitotic spindle, and the phragmoplast. PKN1 is conserved throughout the plant kingdom, with its function in cell division being evolutionarily conserved in the liverwort Marchantia polymorpha. Our data thus demonstrate that PKN1 represents a novel, plant-specific protein with a role in cell division that is likely proteolytically controlled by the CCS52A2-activated APC/C. [ABSTRACT FROM AUTHOR]

Summary Because seed yield is the major factor determining the commercial success of grain crop cultivars, there is a large interest to obtain more understanding of the genetic factors underlying this trait. Despite many studies, mainly in the model plant Arabidopsis thaliana, have reported transgenes and mutants with effects on seed number and/or seed size, knowledge about seed yield parameters remains fragmented. This study investigated the effect of 46 genes, either in gain- and/or loss-of-function situations, with a total of 64 Arabidopsis lines being examined for seed phenotypes such as seed size, seed number per silique, number of inflorescences, number of branches on the main inflorescence and number of siliques. Sixteen of the 46 genes, examined in 14 Arabidopsis lines, were reported earlier to directly affect in seed size and/or seed number or to indirectly affect seed yield by their involvement in biomass production. Other genes involved in vegetative growth, flower or inflorescence development or cell division were hypothesized to potentially affect the final seed size and seed number. Analysis of this comprehensive data set shows that of the 14 lines previously described to be affected in seed size or seed number, only nine showed a comparable effect. Overall, this study provides the community with a useful resource for identifying genes with effects on seed yield and candidate genes underlying seed QTL. In addition, this study highlights the need for more thorough analysis of genes affecting seed yield. [ABSTRACT FROM AUTHOR]

We report on the construction of sex-specific high-density linkage maps and identification of sex-linked markers for the black tiger shrimp (Penaeus monodon). Overall, we identified 44 male and 43 female linkage groups (2n = 88) from the analysis of 2306 AFLP markers segregating in three full-sib families, covering 2378 and 2362 cM, respectively. Twenty-one putatively homologous linkage groups, including the sex-linkage groups, were identified between the female and male linkage maps. Six sex-linked AFLP marker alleles were inherited from female parents in the three families, suggesting that the P. monodon adopts a WZ-ZZ sex-determining system. Two sex-linked AFLP markers, one of which we converted into an allele-specific assay, confirmed their association with sex in a panel of 52 genetically unrelated animals. [ABSTRACT FROM AUTHOR]

The WEE1 and ATM AND RAD3-RELATED (ATR) kinases are important regulators of the plant intra-S-phase checkpoint; consequently, WEE1KO and ATRKO roots are hypersensitive to replication-inhibitory drugs. Here, we report on a loss-of-function mutant allele of the FASCIATA1 (FAS1) subunit of the chromatin assembly factor 1 (CAF-1) complex that suppresses the phenotype of WEE1- or ATR-deficient Arabidopsis (Arabidopsis thaliana) plants. We demonstrate that lack of FAS1 activity results in the activation of an ATAXIA TELANGIECTASIA MUTATED (ATM)- and SUPPRESSOR OF GAMMA-RESPONSE 1 (SOG1)-mediated G2/M-arrest that renders the ATR and WEE1 checkpoint regulators redundant. This ATM activation accounts for the telomere erosion and loss of ribosomal DNA that are described for fas1 plants. Knocking out SOG1 in the fas1 wee1 background restores replication stress sensitivity, demonstrating that SOG1 is an important secondary checkpoint regulator in plants that fail to activate the intra-S-phase checkpoint. [ABSTRACT FROM AUTHOR]

The ataxia-telangiectasia mutated (ATM) and ATM and Rad3-related (ATR) kinases coordinate the DNA damage response. The roles described for Arabidopsis thaliana ATR and ATM are assumed to be conserved over other plant species, but molecular evidence is scarce. Here, we demonstrate that the functions of ATR and ATM are only partially conserved between Arabidopsis and maize (Zea mays). In both species, ATR and ATM play a key role in DNA repair and cell cycle checkpoint activation, but whereas Arabidopsis plants do not suffer from the absence of ATR under control growth conditions, maize mutant plants accumulate replication defects, likely due to their large genome size. Moreover, contrarily to Arabidopsis, maize ATM deficiency does not trigger meiotic defects, whereas the ATR kinase appears to be crucial for the maternal fertility. Strikingly, ATR is required to repress premature endocycle onset and cell death in the maize endosperm. Its absence results in a reduction of kernel size, protein and starch content, and a stochastic death of kernels, a process being counteracted by ATM. Additionally, while Arabidopsis atr atm double mutants are viable, no such mutants could be obtained for maize. Therefore, our data highlight that the mechanisms maintaining genome integrity may be more important for vegetative and reproductive development than previously anticipated. [ABSTRACT FROM AUTHOR]

Aluminum (Al) toxicity and inorganic phosphate (Pi) limitation are widespread chronic abiotic and mutually enhancing stresses that profoundly affect crop yield. Both stresses strongly inhibit root growth, resulting from a progressive exhaustion of the stem cell niche. Here, we report on a casein kinase 2 (CK2) inhibitor identified by its capability to maintain a functional root stem cell niche in Arabidopsis thaliana under Al toxic conditions. CK2 operates through phosphorylation of the cell cycle checkpoint activator SUPPRESSOR OF GAMMA RADIATION1 (SOG1), priming its activity under DNA-damaging conditions. In addition to yielding Al tolerance, CK2 and SOG1 inactivation prevents meristem exhaustion under Pi starvation, revealing the existence of a low Pi-induced cell cycle checkpoint that depends on the DNA damage activator ATAXIA-TELANGIECTASIA MUTATED (ATM). Overall, our data reveal an important physiological role for the plant DNA damage response pathway under agriculturally limiting growth conditions, opening new avenues to cope with Pi limitation. [ABSTRACT FROM AUTHOR]

The anaphase promoting complex/cyclosome (APC/C) controls unidirectional progression through the cell cycle by marking key cell cycle proteins for proteasomal turnover. Its activity is temporally regulated by the docking of different activating subunits, known in plants as CELL DIVISION PROTEIN20 (CDC20) and CELL CYCLE SWITCH52 (CCS52). Despite the importance of the APC/C during cell proliferation, the number of identified targets in the plant cell cycle is limited. Here, we used the growth and meristem phenotypes of Arabidopsis (Arabidopsis thaliana) CCS52A2-deficient plants in a suppressor mutagenesis screen to identify APC/CCCS52A2 substrates or regulators, resulting in the identification of a mutant cyclin CYCA3;4 allele. CYCA3;4 deficiency partially rescues the ccs52a2-1 phenotypes, whereas increased CYCA3;4 levels enhance the scored ccs52a2-1 phenotypes. Furthermore, whereas the CYCA3;4 protein is promptly broken down after prophase in wild-type plants, it remains present in later stages of mitosis in ccs52a2-1 mutant plants, marking it as a putative APC/CCCS52A2 substrate. Strikingly, increased CYCA3;4 levels result in aberrant root meristem and stomatal divisions, mimicking phenotypes of plants with reduced RETINOBLASTOMA-RELATED PROTEIN1 (RBR1) activity. Correspondingly, RBR1 hyperphosphorylation was observed in CYCA3;4 gain-of-function plants. Our data thus demonstrate that an inability to timely destroy CYCA3;4 contributes to disorganized formative divisions, possibly in part caused by the inactivation of RBR1. [ABSTRACT FROM AUTHOR]

The rapid appearance of herbicide-resistant weeds combined with a lack of novel herbicides being brought to market reduces crop production, thereby threatening food security worldwide. Here, we report on the use of the previously identified cellulose biosynthesis-inhibiting chemical compound C17 as a potential herbicide. Toxicity tests showed that C17 efficiently inhibits the growth of various weeds and widely cultivated dicotyledonous crops, whereas only slight or no growth inhibition was observed for monocotyledonous crops. Surprisingly, when exposed to a mixture of C17 and one of two well-known cellulose biosynthesis inhibitors (CBIs), isoxaben and indaziflam, an additive growth inhibition was observed, demonstrating that C17 has a different mode of action that can be used to sensitize plants toward known CBIs. Moreover, we demonstrate that a C17-resistant CESA3 allele can be used as a positive transformation selection marker and that C17 resistance can be obtained through genome engineering of the wild-type CESA3 allele using clustered regularly interspaced short palindromic repeats-mediated base editing. This editing system allowed us to engineer C17 tolerance in an isoxaben-resistant line, resulting in double herbicide-resistant plants. [ABSTRACT FROM AUTHOR]

Somatic polyploidy caused by endoreplication is observed in arthropods, molluscs, and vertebrates but is especially prominent in higher plants, where it has been postulated to be essential for cell growth and fate maintenance. However, a comprehensive understanding of the physiological significance of plant endopolyploidy has remained elusive. Here, we modeled and experimentally verified a high-resolution DNA endoploidy map of the developing Arabidopsis thaliana root, revealing a remarkable spatiotemporal control of DNA endoploidy levels across tissues. Fitting of a simplified model to publicly available data sets profiling root gene expression under various environmental stress conditions suggested that this root endoploidy patterning may be stress-responsive. Furthermore, cellular and transcriptomic analyses revealed that inhibition of endoreplication onset alters the nuclear-to-cellular volume ratio and the expression of cell wall-modifying genes, in correlation with the appearance of cell structural changes. Our data indicate that endopolyploidy might serve to coordinate cell expansion with structural stability and that spatiotemporal endoreplication pattern changes may buffer for stress conditions, which may explain the widespread occurrence of the endocycle in plant species growing in extreme or variable environments. [ABSTRACT FROM AUTHOR]

The quiescent center (QC) plays an essential role during root development by creating a microenvironment that preserves the stem cell fate of its surrounding cells. Despite being surrounded by highly mitotic active cells, QC cells self-renew at a low proliferation rate. Here, we identified the ERF115 transcription factor as a rate-limiting factor of QC cell division, acting as a transcriptional activator of the phytosulfokine PSK5 peptide hormone. ERF115 marks QC cell division but is restrained through proteolysis by the APC/CCCS52A2 ubiquitin ligase, whereas QC proliferation is driven by brassinosteroid-dependent ERF115 expression. Together, these two antagonistic mechanisms delimit ERF115 activity, which is called upon when surrounding stem cells are damaged, revealing a cell cycle regulatory mechanism accounting for stem cell niche longevity. [ABSTRACT FROM AUTHOR]

We report on the construction of sex-specific linkage maps, the identification of sex-linked markers and the genome size estimation for the brine shrimp Artemia franciscana. Overall, from the analysis of 433 AFLP markers segregating in a 112 full-sib family we identified 21 male and 22 female linkage groups (2n = 42), covering 1,041 and 1,313 cM respectively. Fifteen putatively homologous linkage groups, including the sex linkage groups, were identified between the female and male linkage map. Eight sex-linked AFLP marker alleles were inherited from the female parent, supporting the hypothesis of a WZ–ZZ sex-determining system. The haploid Artemia genome size was estimated to 0.93 Gb by flow cytometry. The produced Artemia linkage maps provide the basis for further fine mapping and exploring of the sex-determining region and are a possible marker resource for mapping genomic loci underlying phenotypic differences among Artemia species. [ABSTRACT FROM AUTHOR]

Large-scale comparison of sequence polymorphism and divergence at numerous genomic loci within and between closely related species can reveal signatures of natural selection. Here, we present a population genomics study based on direct sequencing of 61 mitotic cell cycle genes from 30 Arabidopsis thaliana accessions and comparison of the resulting data to the close relative Arabidopsis lyrata. We found that the Arabidopsis core cell cycle (CCC) machinery is not highly constrained but is subject to different modes of selection. We found patterns of purifying selection for the cyclin-dependent kinase (CDK), CDK subunit, retinoblastoma, and WEE1 gene families. Other CCC gene families often showed a mix of one or two constrained genes and relaxed purifying selection on the other genes. We found several large effect mutations in CDKB1;2 that segregate in the species. We found a strong signature of adaptive protein evolution in the Kip-related protein KRP6 and departures from equilibrium at CDKD;1 and CYCA3;3 consistent with the operation of selection in these gene regions. Our data suggest that within Arabidopsis, the genetic robustness of cell cycle-related processes is more due to functional redundancy than high selective constraint. [ABSTRACT FROM AUTHOR]

Diatoms, an evolutionarily successful group of microalgae, display high levels of intraspecific genetic variability in natural populations. However, the contribution of various mechanisms generating such diversity is unknown. Here we estimated the genetic micro-diversity within a natural diatom population and mapped the genomic changes arising within clonally propagated diatom cell cultures. Through quantification of haplotype diversity by next-generation sequencing and amplicon re-sequencing of selected loci, we documented a rapid accumulation of multiple haplotypes accompanied by the appearance of novel protein variants in cell cultures initiated from a single founder cell. Comparison of the genomic changes between mother and daughter cells revealed copy number variation and copy-neutral loss of heterozygosity leading to the fixation of alleles within individual daughter cells. The loss of heterozygosity can be accomplished by recombination between homologous chromosomes. To test this hypothesis, we established an endogenous readout system and estimated that the frequency of interhomolog mitotic recombination was under standard growth conditions 4.2 events per 100 cell divisions. This frequency is increased under environmental stress conditions, including treatment with hydrogen peroxide and cadmium. These data demonstrate that copy number variation and mitotic recombination between homologous chromosomes underlie clonal variability in diatom populations. We discuss the potential adaptive evolutionary benefits of the plastic response in the interhomolog mitotic recombination rate, and we propose that this may have contributed to the ecological success of diatoms. [Display omitted] • Diatom clonal cultures rapidly accumulate genomic diversity • At the single-cell level loss of heterozygosity and copy number variation are observed • Genomic changes correlate with the occurrence of mitotic interhomolog recombination • The frequency attains 4.2 events per 100 cell divisions and increases under stress Bulankova et al. show that recombination between homologous chromosomes in vegetative cells of diatom Phaeodactylum tricornutum results in genomic diversity in clonal populations. This diversity includes the appearance of recombined haplotypes, loss of heterozygosity, and copy number variation between individual cells. [ABSTRACT FROM AUTHOR]