SIZ1 regulates phosphate deficiency-induced inhibition of primary root growth of Arabidopsis by modulating Fe accumulation and ROS production in its roots.

Inhibition of primary root (PR) growth is a typical developmental response of Arabidopsis to phosphate (Pi) deficiency. Functional disruption of SIZ1, a SUMO E3 ligase, is known to enhance the Pi deficiency-induced inhibition of PR growth. The molecular mechanism of how SIZ1 regulates PR growth under Pi deficiency, however, remains unknown. SIZ1 was recently reported to partially SUMOylate STOP1, a transcription factor that functions in plant tolerance to aluminum toxicity and in plant responses to Pi deficiency by regulating the expression of ALMT1. ALMT1 encodes an aluminum-activated malate transporter, and its expression is induced by Pi deficiency. In siz1, the expression of ALMT1 is enhanced and the removal of Fe from Pi-deficient medium suppressed the siz1 mutant phenotype. In this report, we show that siz1 overaccumulates Fe in its root apoplasts, and consequently, produces more hydroxyl radicals, which are detrimental to root growth. Such physiological changes in siz1 can be completely suppressed by the mutation of STOP1 or ALMT1. Based on previously published work and the results of the current study, we propose that SIZ1 regulates Pi deficiency-mediated PR growth through modulating the accumulation of Fe and the production of hydroxyl radicals by controlling ALMT1 expression.

Microarray expression profile analysis of circular RNAs and their potential regulatory role in bladder carcinoma.

Dysregulated circular RNAs (circRNAs) often contribute to the occurrence and development of various tumors; however, the function and mechanism of circRNAs are largely unknown in human bladder cancer (BC). In the present study, dysregulated circRNAs between BC and adjacent non­neoplastic bladder tissues were analyzed by circRNA microarray. We randomly selected 10 upregulated and five downregulated circRNAs for validation by quantitative real­time PCR. Bioinformatics analysis was further conducted to investigate the potential function of these differentially expressed circRNAs, with the differential expression of hsa_circRNA_100876, mir­136­5p, and mRNA­chromobox 4 (CBX4) subsequently verified. A total of 512 differentially expressed circRNAs were identified after scanning and normalization (340 upregulated and 172 downregulated circRNAs), with pathway and Gene Ontology analyses revealing their association with multiple significant cancer pathways. Construction of a circRNA­microRNA­mRNA network suggested additional potential roles of these circRNAs. The expression of hsa_circRNA_100876 and CBX4 was significantly negatively correlated with the expression of miR­136­5p. Additionally, hsa_circRNA_100876 was highly positively correlated with CBX4 expression. The results revealed that hsa_circRNA_100876 inhibition suppressed BC cell proliferation and it was associated with advanced T stage and lymphatic metastasis, and poor overall survival of BC patients. In conclusion, these differentially expressed circRNAs offer novel insights into potential biological markers or new therapeutic targets for the treatment of BC. Furthermore, hsa_circRNA_100876 may increase the expression of CBX4 by competing with miR­136­5p, ultimately promoting the malignant biological behavior of BC. Aberrantly expressed hsa_circRNA_100876 could be used as a potential non­invasive biomarker for the early detection and screening of BC.

SIZ1-Mediated SUMO Modification of SEUSS Regulates Photomorphogenesis in Arabidopsis.

Small ubiquitin-like modifier (SUMO) post-translational modification (SUMOylation) plays essential roles in regulating various biological processes; however, its function and regulation in the plant light signaling pathway are largely unknown. SEUSS (SEU) is a transcriptional co-regulator that integrates light and temperature signaling pathways, thereby regulating plant growth and development in Arabidopsis thaliana. Here, we show that SEU is a substrate of SUMO1, and that substitution of four conserved lysine residues disrupts the SUMOylation of SEU, impairs its function in photo- and thermomorphogenesis, and enhances its interaction with PHYTOCHROME-INTERACTING FACTOR 4 transcription factors. Furthermore, the SUMO E3 ligase SIZ1 interacts with SEU and regulates its SUMOylation. Moreover, SEU directly interacts with phytochrome B photoreceptors, and the SUMOylation and stability of SEU are activated by light. Our study reveals a novel post-translational modification mechanism of SEU in which light regulates plant growth and development through SUMOylation-mediated protein stability.

Tomato YABBY2b controls plant height through regulating indole-3-acetic acid-amido synthetase (GH3.8) expression.

Dwarfing is a desirable agronomic trait in cultivation management. Dwarf plants are lodging-resistant, compact, and perform well under high-density planting. The use of dwarf genetic resources is one approach to improve crop yield. YABBY2b in tomato (Solanum lycopersicum) encodes a transcription factor that regulates plant height. In this study, we created YABBY2b knockout mutant lines, and the resulting yabby2b plants exhibited reduced height and smaller flowers and fruits. The RNA-seq analysis showed that 17 genes responding to gibberellin and auxin were differentially expressed. We hypothesized that indole-3-acetic acid-amido synthetase GH3.8 (GH3.8) played a crucial role in the resulting yabby2b dwarf phenotype. Further analysis showed that YABBY2b suppresses GH3.8 gene expression by directly binding to its promoter, and that this contributes to auxin-mediated repression of GH3.8. Moreover, the silencing of GH3.8 led to increased plant height. Combined, our data suggest that YABBY2b may positively regulate plant height in tomato by inhibiting the expression of growth suppressor GH3.8.

A nucleotide-switch mechanism mediates opposing catalytic activities of Rel enzymes.

Bifunctional Rel stringent factors, the most abundant class of RelA/SpoT homologs, are ribosome-associated enzymes that transfer a pyrophosphate from ATP onto the 3' of guanosine tri-/diphosphate (GTP/GDP) to synthesize the bacterial alarmone (p)ppGpp, and also catalyze the 3' pyrophosphate hydrolysis to degrade it. The regulation of the opposing activities of Rel enzymes is a complex allosteric mechanism that remains an active research topic despite decades of research. We show that a guanine-nucleotide-switch mechanism controls catalysis by Thermus thermophilus Rel (RelTt). The binding of GDP/ATP opens the N-terminal catalytic domains (NTD) of RelTt (RelTtNTD) by stretching apart the two catalytic domains. This activates the synthetase domain and allosterically blocks hydrolysis. Conversely, binding of ppGpp to the hydrolase domain closes the NTD, burying the synthetase active site and precluding the binding of synthesis precursors. This allosteric mechanism is an activity switch that safeguards against futile cycles of alarmone synthesis and degradation.

Overexpression of MsGH3.5 inhibits shoot and root development through the auxin and cytokinin pathways in apple plants.

Phytohormonal interactions are crucial for plant development. Auxin and cytokinin (CK) both play critical roles in regulating plant growth and development; however, the interaction between these two phytohormones is complex and not fully understood. Here, we isolated a wild apple (Malus sieversii Roem) GRETCHEN HAGEN3 (GH3) gene, MsGH3.5, encoding an indole-3-acetic acid (IAA)-amido synthetase. Overexpression of MsGH3.5 significantly reduced the free IAA content and increased the content of some IAA-amino acid conjugates, and MsGH3.5-overexpressing lines were dwarfed and produced fewer adventitious roots (ARs) than the control. This phenotype is consistent with the role of GH3 in conjugating excess free active IAA to amino acids in auxin homeostasis. Surprisingly, overexpression of MsGH3.5 significantly increased CK concentrations in the whole plant, and altered the expression of genes involved in CK biosynthesis, metabolism and signaling. Furthermore, exogenous CK application induced MsGH3.5 expression through the activity of the CK type-B response regulator, MsRR1a, which mediates the CK primary response. MsRR1a activated MsGH3.5 expression by directly binding to its promoter, linking auxin and CK signaling. Plants overexpressing MsRR1a also displayed fewer ARs, in agreement with the regulation of MsGH3.5 expression by MsRR1a. Taken together, we reveal that MsGH3.5 affects apple growth and development by modulating auxin and CK levels and signaling pathways. These findings provide insight into the interaction between the auxin and CK pathways, and might have substantial implications for efforts to improve apple architecture.

AtSIZ1 improves salt tolerance by maintaining ionic homeostasis and osmotic balance in Arabidopsis.

C2H2-type zinc finger proteins play important roles in plant growth, development, and abiotic stress tolerance. Here, we explored the role of the C2H2-type zinc finger protein SALT INDUCED ZINC FINGER PROTEIN1 (AtSIZ1; At3G25910) in Arabidopsis thaliana under salt stress. AtSIZ1 expression was induced by salt treatment. During the germination stage, the germination rate, germination energy, germination index, cotyledon growth rate, and root length were significantly higher in AtSIZ1 overexpression lines than in the wild type under various stress treatments, whereas these indices were significantly reduced in AtSIZ1 loss-of-function mutants. At the mature seedling stage, the overexpression lines maintained higher levels of K+, proline, and soluble sugar, lower levels of Na+ and MDA, and lower Na+/K+ ratios than the wild type. Stress-related marker genes such as SOS1, AtP5CS1, AtGSTU5, COR15A, RD29A, and RD29B were expressed at higher levels in the overexpression lines than the wild type and loss-of-function mutants under salt treatment. These results indicate that AtSIZ1 improves salt tolerance in Arabidopsis by helping plants maintain ionic homeostasis and osmotic balance.

Maintenance of Nucleolar Homeostasis by CBX4 Alleviates Senescence and Osteoarthritis.

CBX4, a component of polycomb repressive complex 1 (PRC1), plays important roles in the maintenance of cell identity and organ development through gene silencing. However, whether CBX4 regulates human stem cell homeostasis remains unclear. Here, we demonstrate that CBX4 counteracts human mesenchymal stem cell (hMSC) aging via the maintenance of nucleolar homeostasis. CBX4 protein is downregulated in aged hMSCs, whereas CBX4 knockout in hMSCs results in destabilized nucleolar heterochromatin, enhanced ribosome biogenesis, increased protein translation, and accelerated cellular senescence. CBX4 maintains nucleolar homeostasis by recruiting nucleolar protein fibrillarin (FBL) and heterochromatin protein KRAB-associated protein 1 (KAP1) at nucleolar rDNA, limiting the excessive expression of rRNAs. Overexpression of CBX4 alleviates physiological hMSC aging and attenuates the development of osteoarthritis in mice. Altogether, our findings reveal a critical role of CBX4 in counteracting cellular senescence by maintaining nucleolar homeostasis, providing a potential therapeutic target for aging-associated disorders.

The SUMO Conjugation Complex Self-Assembles into Nuclear Bodies Independent of SIZ1 and COP1.

Attachment of the small ubiquitin-like modifier (SUMO) to substrate proteins modulates their turnover, activity, or interaction partners. However, how this SUMO conjugation activity concentrates the proteins involved and the substrates into uncharacterized nuclear bodies (NBs) remains poorly understood. Here, we characterized the requirements for SUMO NB formation and for their subsequent colocalization with the E3 ubiquitin ligase CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1), a master regulator of plant growth. COP1 activity results in degradation of transcription factors, which primes the transcriptional response that underlies elongation growth induced by darkness and high ambient temperatures (skoto- and thermomorphogenesis, respectively). SUMO conjugation activity alone was sufficient to target the SUMO machinery into NBs. Colocalization of these bodies with COP1 required, in addition to SUMO conjugation activity, a SUMO acceptor site in COP1 and the SUMO E3 ligase SAP and Miz 1 (SIZ1). We found that SIZ1 docks in the substrate-binding pocket of COP1 via two valine-proline peptide motifs, which represent a known interaction motif of COP1 substrates. The data reveal that SIZ1 physically connects COP1 and SUMO conjugation activity in the same NBs that can also contain the blue-light receptors CRYPTOCHROME 1 and CRYPTOCHROME 2. Our findings thus suggest that sumoylation stimulates COP1 activity within NBs. Moreover, the presence of SIZ1 and SUMO in these NBs explains how both the timing and amplitude of the high-temperature growth response is controlled. The strong colocalization of COP1 and SUMO in these NBs might also explain why many COP1 substrates are sumoylated.

OsSIZ2 regulates nitrogen homeostasis and some of the reproductive traits in rice.

Small ubiquitin-related modifier (SUMO) is a post-translational modification of proteins that has important roles in plant growth and development as well as nutrition study. OsSIZ1, a SUMO E3 ligase in rice (Oryza sativa), exerts regulatory influence on nitrogen (N) homeostasis. Here, we investigated the biological function of OsSIZ2, a paralog of OsSIZ1, in the responses to nitrogen, anther dehiscence, and seed length using a reverse genetics approach. The expression of OsSIZ2 was increased during N deficiency. Under -N condition, total N concentration in the root of OsSIZ2-Ri plants and ossiz2 was significantly increased compared with wild type. Further, 15N-labelled uptake assay revealed the role of OsSIZ2 in acquisition and mobilization of N. Moreover, qRT-PCR analyses revealed that several genes involved in the maintenance of N homeostasis were altered in OsSIZ2 mutants. In addition, ossiz2 indicated obvious defects in anther dehiscence, pollen fertility, and seed set percentage. Interestingly, however, the seed length was longer in the mutant compared with wild type. Overall, these results suggest pivotal roles of OsSIZ2 in regulating homeostasis of N and different agronomic traits including anther and seed development.

Apple SUMO E3 ligase MdSIZ1 is involved in the response to phosphate deficiency.

In plants, SIZ1 regulates abiotic and biotic stress responses by promoting the SUMOylation of proteins. The apple MdSIZ1 protein has conserved domains similar to those of Arabidopsis AtSIZ1. Real-time fluorescent quantitative analysis showed that MdSIZ1 gene expression was induced by phosphate-deficient conditions. In addition, the level of SUMOylation was also significantly increased under these conditions. The MYB transcription factor MdPHR1 might be a target for the SUMO protein, which is a phosphorus starvation-dependent protein. Subsequently, an MdSIZ1 expression vector was constructed and transformed in Arabidopsis mutant siz1-2 and apple callus. The MdSIZ1 transgenic Arabidopsis partially complemented the defect phenotype of siz1-2 under phosphate-deficient conditions. The survival rate, length of primary root, and number or density of lateral roots were similar between the transgenic lines and wild type (WT). Under phosphate-deficient conditions, the SUMO conjugate and fresh weight of the MdSIZ1 transgenic apple callus were improved compared with WT. The MdSIZ1 transgenic apple callus grew under phosphate-deficient conditions, whereas the MdSIZ1 sense apple callus did not. Therefore, MdSIZ1 is involved in the regulation of the phosphate-deficiency response in apple.

Stochastic Turing patterns in a synthetic bacterial population.

The origin of biological morphology and form is one of the deepest problems in science, underlying our understanding of development and the functioning of living systems. In 1952, Alan Turing showed that chemical morphogenesis could arise from a linear instability of a spatially uniform state, giving rise to periodic pattern formation in reaction-diffusion systems but only those with a rapidly diffusing inhibitor and a slowly diffusing activator. These conditions are disappointingly hard to achieve in nature, and the role of Turing instabilities in biological pattern formation has been called into question. Recently, the theory was extended to include noisy activator-inhibitor birth and death processes. Surprisingly, this stochastic Turing theory predicts the existence of patterns over a wide range of parameters, in particular with no severe requirement on the ratio of activator-inhibitor diffusion coefficients. To explore whether this mechanism is viable in practice, we have genetically engineered a synthetic bacterial population in which the signaling molecules form a stochastic activator-inhibitor system. The synthetic pattern-forming gene circuit destabilizes an initially homogenous lawn of genetically engineered bacteria, producing disordered patterns with tunable features on a spatial scale much larger than that of a single cell. Spatial correlations of the experimental patterns agree quantitatively with the signature predicted by theory. These results show that Turing-type pattern-forming mechanisms, if driven by stochasticity, can potentially underlie a broad range of biological patterns. These findings provide the groundwork for a unified picture of biological morphogenesis, arising from a combination of stochastic gene expression and dynamical instabilities.

The Arabidopsis SUMO E3 ligase SIZ1 mediates the temperature dependent trade-off between plant immunity and growth.

Increased ambient temperature is inhibitory to plant immunity including auto-immunity. SNC1-dependent auto-immunity is, for example, fully suppressed at 28°C. We found that the Arabidopsis sumoylation mutant siz1 displays SNC1-dependent auto-immunity at 22°C but also at 28°C, which was EDS1 dependent at both temperatures. This siz1 auto-immune phenotype provided enhanced resistance to Pseudomonas at both temperatures. Moreover, the rosette size of siz1 recovered only weakly at 28°C, while this temperature fully rescues the growth defects of other SNC1-dependent auto-immune mutants. This thermo-insensitivity of siz1 correlated with a compromised thermosensory growth response, which was independent of the immune regulators PAD4 or SNC1. Our data reveal that this high temperature induced growth response strongly depends on COP1, while SIZ1 controls the amplitude of this growth response. This latter notion is supported by transcriptomics data, i.e. SIZ1 controls the amplitude and timing of high temperature transcriptional changes including a subset of the PIF4/BZR1 gene targets. Combined our data signify that SIZ1 suppresses an SNC1-dependent resistance response at both normal and high temperatures. At the same time, SIZ1 amplifies the dark and high temperature growth response, likely via COP1 and upstream of gene regulation by PIF4 and BRZ1.

Oryza sativa salt-induced RING E3 ligase 2 (OsSIRP2) acts as a positive regulator of transketolase in plant response to salinity and osmotic stress.

MAIN CONCLUSION: A rice gene (OsSIRP2) encoding the RING Ub E3 ligase was highly induced under salinity stress and physically interacted with a transketolase (OsTKL1). Overexpression of OsSIRP2 conferred salinity and osmotic stress tolerance in plants. The RING E3 ligases play a vital role in post transitional modification through ubiquitination-mediated protein degradation that mediate plants responses during abiotic stresses and signal transduction. In this study, we report an Oryza sativa salt induced Really Interesting New Gene (RING) finger protein 2 gene (OsSIRP2) and elucidate its role under salinity and osmotic stress. The transcript levels of OsSIRP2 in rice leaves were induced in response to different abiotic stresses, such as salt, drought, heat, and abscisic acid (ABA) exposure. In vitro ubiquitination revealed that the OsSIRP2 protein formed poly-ubiquitin products, whereas a single amino acid substitution in OsSIRP2 (OsSIRP2C149A) in the RING domain did not form ubiquitinated substrates, supporting the hypothesis that E3 ligase activity requires the functional RING domain. Using the yeast two-hybrid (Y2H) assay, O. sativa transketolase 1 (OsTKL1) was identified as an interacting partner. OsSIRP2 was localized in the nucleus, whereas its interacting partner (OsTKL1) was localized in the cytosol and plastids in the rice protoplasts. Fluorescence signals between OsSIRP2 and OsTKL1 were observed in the cytosol. The pull-down assay confirmed the physical interaction between OsSIRP2 and OsTKL1. In vitro ubiquitination assay and in vitro protein degradation assay revealed that OsSIRP2 ubiquitinates OsTKL1 and enhances the degradation of OsTKL1 through the 26S proteasomal pathway. Heterogeneous overexpression of OsSIRP2 resulted in conferring tolerance against salinity and osmotic stress. Overall, our findings suggest that OsSIRP2 may be associated with plant responses to abiotic stresses and act as a positive regulator of salt and osmotic stress tolerance.

Trivial role for NSMCE2 during in vitro proliferation and differentiation of male germline stem cells.

Spermatogenesis, starting with spermatogonial differentiation, is characterized by ongoing and dramatic alterations in composition and function of chromatin. Failure to maintain proper chromatin dynamics during spermatogenesis may lead to mutations, chromosomal aberrations or aneuploidies. When transmitted to the offspring, these can cause infertility or congenital malformations. The structural maintenance of chromosomes (SMC) 5/6 protein complex has recently been described to function in chromatin modeling and genomic integrity maintenance during spermatogonial differentiation and meiosis. Among the subunits of the SMC5/6 complex, non-SMC element 2 (NSMCE2) is an important small ubiquitin-related modifier (SUMO) ligase. NSMCE2 has been reported to be essential for mouse development, prevention of cancer and aging in adult mice and topological stress relief in human somatic cells. By using in vitro cultured primary mouse spermatogonial stem cells (SSCs), referred to as male germline stem (GS) cells, we investigated the function of NSMCE2 during spermatogonial proliferation and differentiation. We first optimized a protocol to generate genetically modified GS cell lines using CRISPR-Cas9 and generated an Nsmce2-/- GS cell line. Using this Nsmce2-/- GS cell line, we found that NSMCE2 was dispensable for proliferation, differentiation and topological stress relief in mouse GS cells. Moreover, RNA sequencing analysis demonstrated that the transcriptome was only minimally affected by the absence of NSMCE2. Only differential expression of Sgsm1 appeared highly significant, but with SGSM1 protein levels being unaffected without NSMCE2. Hence, despite the essential roles of NSMCE2 in somatic cells, chromatin integrity maintenance seems differentially regulated in the germline.

The E3 SUMO ligase AtSIZ1 functions in seed germination in Arabidopsis.

Seed germination is an important stage in the lifecycle of a plant because it determines subsequent vegetative growth and reproduction. Here, we show that the E3 SUMO ligase AtSIZ1 regulates seed dormancy and germination. The germination rates of the siz1 mutants were less than 50%, even after a short period of ripening. However, their germination rates increased to wild-type levels after cold stratification or long periods of ripening. In addition, exogenous gibberellin (GA) application improved the germination rates of the siz1 mutants to the wild-type level. In transgenic plants, suppression of AtSIZ1 caused rapid post-translational decay of SLEEPY1 (SLY1), a positive regulator of GA signaling, during germination, and inducible AtSIZ1 overexpression led to increased SLY1 levels. In addition, overexpressing wild-type SLY1 in transgenic sly1 mutants increased their germination ratios to wild-type levels, whereas the germination ratio of transgenic sly1 mutants overexpressing mSLY1 was similar to that of sly1. The germination ratios of siz1 mutant seeds in immature developing siliques were much lower than those of the wild-type. Moreover, SLY1 and DELAY OF GERMINATION 1 (DOG1) transcript levels were reduced in the siz1 mutants, whereas the transcript levels of DELLA and ABSCISIC ACID INSENSITIVE 3 (ABI3) were higher than those of the wild-type. Taken together, these results indicate that the reduced germination of the siz1 mutants results from impaired GA signaling due to low SLY1 levels and activity, as well as hyperdormancy due to high levels of expression of dormancy-related genes including DOG1.

Catalytic mechanism and allosteric regulation of an oligomeric (p)ppGpp synthetase by an alarmone.

Nucleotide-based second messengers serve in the response of living organisms to environmental changes. In bacteria and plant chloroplasts, guanosine tetraphosphate (ppGpp) and guanosine pentaphosphate (pppGpp) [collectively named "(p)ppGpp"] act as alarmones that globally reprogram cellular physiology during various stress conditions. Enzymes of the RelA/SpoT homology (RSH) family synthesize (p)ppGpp by transferring pyrophosphate from ATP to GDP or GTP. Little is known about the catalytic mechanism and regulation of alarmone synthesis. It also is unclear whether ppGpp and pppGpp execute different functions. Here, we unravel the mechanism and allosteric regulation of the highly cooperative alarmone synthetase small alarmone synthetase 1 (SAS1) from Bacillus subtilis. We determine that the catalytic pathway of (p)ppGpp synthesis involves a sequentially ordered substrate binding, activation of ATP in a strained conformation, and transfer of pyrophosphate through a nucleophilic substitution (SN2) reaction. We show that pppGpp-but not ppGpp-positively regulates SAS1 at an allosteric site. Although the physiological significance remains to be elucidated, we establish the structural and mechanistic basis for a biological activity in which ppGpp and pppGpp execute different functional roles.

Regulation of axon regeneration by the RNA repair and splicing pathway.

Mechanisms governing a neuron's regenerative ability are important but not well understood. We identify Rtca (RNA 3'-terminal phosphate cyclase) as an inhibitor of axon regeneration. Removal of Rtca cell-autonomously enhanced axon regrowth in the Drosophila CNS, whereas its overexpression reduced axon regeneration in the periphery. Rtca along with the RNA ligase Rtcb and its catalyst Archease operate in the RNA repair and splicing pathway important for stress-induced mRNA splicing, including that of Xbp1, a cellular stress sensor. Drosophila Rtca and Archease had opposing effects on Xbp1 splicing, and deficiency of Archease or Xbp1 impeded axon regeneration in Drosophila. Moreover, overexpressing mammalian Rtca in cultured rodent neurons reduced axonal complexity in vitro, whereas reducing its function promoted retinal ganglion cell axon regeneration after optic nerve crush in mice. Our study thus links axon regeneration to cellular stress and RNA metabolism, revealing new potential therapeutic targets for treating nervous system trauma.

Hijacking of the jasmonate pathway by the mycotoxin fumonisin B1 (FB1) to initiate programmed cell death in Arabidopsis is modulated by RGLG3 and RGLG4.

The mycotoxin fumonisin B1 (FB1) is a strong inducer of programmed cell death (PCD) in plants, but its underlying mechanism remains unclear. Here, we describe two ubiquitin ligases, RING DOMAIN LIGASE3 (RGLG3) and RGLG4, which control FB1-triggered PCD by modulating the jasmonate (JA) signalling pathway in Arabidopsis thaliana. RGLG3 and RGLG4 transcription was sensitive to FB1. Arabidopsis FB1 sensitivity was suppressed by loss of function of RGLG3 and RGLG4 and was increased by their overexpression. Thus RGLG3 and RGLG4 have coordinated and positive roles in FB1-elicited PCD. Mutated JA perception by coi1 disrupted the RGLG3- and RGLG4-related response to FB1 and interfered with their roles in cell death. Although FB1 induced JA-responsive defence genes, it repressed growth-related, as well as JA biosynthesis-related, genes. Consistently, FB1 application reduced JA content in wild-type plants. Furthermore, exogenously applied salicylic acid additively suppressed JA signalling with FB1 treatment, suggesting that FB1-induced salicylic acid inhibits the JA pathway during this process. All of these effects were attenuated in rglg3 rglg4 plants. Altogether, these data suggest that the JA pathway is hijacked by the toxin FB1 to elicit PCD, which is coordinated by Arabidopsis RGLG3 and RGLG4.

An ENU-induced mutation in the mouse Rnf212 gene is associated with male meiotic failure and infertility.

The ENU-induced repro57 mutation was identified in an unbiased screen for the discovery of novel genes for fertility. Male repro57 homozygous mice are infertile and exhibit significantly reduced testis weight compared with WT mice. Histological examination of mutant testes revealed that spermatocytes degenerated during late prophase, and no mature spermatozoa were found in the seminiferous epithelium, suggesting that infertility is caused by the arrest of spermatogenesis at late meiotic prophase. Consistent with this hypothesis, the number of foci with MLH1, a protein essential for crossing over, is greatly reduced in repro57 mutant spermatocytes, which also lack chiasmata between homologs and exhibit premature dissociation of XY chromosomes. In repro57 mutant mice, we identified a mutation in the Rnf212 gene, encoding Ring finger protein 212. The overall phenotype of repro57 mice is consistent with the recently reported phenotype of the Rnf212 knockout mice; slight differences may be due to genetic background effects. Thus, the repro57 nonsense mutation provides a new allele of the mouse Rnf212 gene.