Plant autoimmunity-fresh insights into an old phenomenon.

The plant immune system is well equipped to ward off the attacks of different types of phytopathogens. It primarily relies on two types of immune sensors-plasma membrane-resident receptor-like kinases and intracellular nucleotide-binding domain leucine-rich repeat (NLRs) receptors that engage preferentially in pattern- and effector-triggered immunity, respectively. Delicate fine-tuning, in particular of the NLR-governed branch of immunity, is key to prevent inappropriate and deleterious activation of plant immune responses. Inadequate NLR allele constellations, such as in the case of hybrid incompatibility, and the mis-activation of NLRs or the absence or modification of proteins guarded by these NLRs can result in the spontaneous initiation of plant defense responses and cell death-a phenomenon referred to as plant autoimmunity. Here, we review recent insights augmenting our mechanistic comprehension of plant autoimmunity. The recent findings broaden our understanding regarding hybrid incompatibility, unravel candidates for proteins likely guarded by NLRs and underline the necessity for the fine-tuning of NLR expression at various levels to avoid autoimmunity. We further present recently emerged tools to study plant autoimmunity and draw a cross-kingdom comparison to the role of NLRs in animal autoimmune conditions.

Cyclin G2 upregulation impairs migration, invasion, and network formation through RNF123/Dvl2/JNK signaling in the trophoblast cell line HTR8/SVneo, a possible role in preeclampsia.

Disruption of extravillous trophoblast (EVT) migration and invasion is considered to be responsible for pathological placentation in preeclampsia (PE). Cyclin G2 (CCNG2) is an atypical cyclin that inhibits cell cycle progression. However, its biological function and underlying molecular mechanism in PE are poorly understood. In this study, clinical data demonstrated that CCNG2 was significantly upregulated in PE placenta and associated with invasive EVT dysfunction. Additionally, Ccng2 knockout led to an attenuation of PE-like symptoms in the PE mouse model produced via treatment with NG-nitro-L-arginine methyl ester (L-NAME). In vitro, CCNG2 inhibited the migration, invasion, and endothelial-like network formation of human trophoblast cell line HTR8/SVneo. Mechanically, CCNG2 suppressed JNK-dependent Wnt/PCP signaling and its downstream indicators including epithelial-to-mesenchymal transition (EMT) markers and matrix metalloproteinases (MMPs) via promoting the polyubiquitination degradation of dishevelled 2 (Dvl2) protein in HTR8/SVneo cells. We also discovered that the E3 ligase Ring finger protein 123 (RNF123), as a novel CCNG2 target among HTR8/SVneo cells, interacted with Dvl2 and participated in CCNG2-induced polyubiquitination degradation of Dvl2. Moreover, we verified that the treatment of HTR8/SVneo cells with RNF123-specific siRNA improved polyubiquitination-induced degradation of Dvl2 and the activity of Wnt/PCP-JNK signaling mediated by CCNG2. Taken together, our results reveal that the CCNG2/RNF123/Dvl2/JNK axis may be involved in the pathogenesis and progression of PE through trophoblastic cell function modulation, thus probably providing us with new therapeutic strategies for PE treatment.

Cyclin G2 regulates canonical Wnt signalling via interaction with Dapper1 to attenuate tubulointerstitial fibrosis in diabetic nephropathy.

Cyclin G2 (CCNG2) is an atypical cyclin that inhibits cell cycle progression and is often dysregulated in human cancers. Cyclin G2 in the occurrence and development of diabetic nephropathy (DN), one of the most severe diabetic complications, has not been fully identified. In this study, we investigated the function and regulatory mechanism of cyclin G2 in DN. In vivo studies revealed that a deficiency of cyclin G2 significantly increased albuminuria and promoted tubulointerstitial fibrosis in established DN. Cyclin G2 regulated the expression of fibrosis-related proteins via the canonical Wnt signalling pathway in renal tubular epithelial cells. Moreover, the binding of cyclin G2 to Dapper1 (Dpr1/DACT1), a protein involved in Wnt signalling, decreased the phosphorylation of Dpr1 at Ser762 by casein kinase 1 (CK1) and suppressed the Wnt signalling pathway. These findings reveal that cyclin G2 can protect against renal injury and fibrosis associated with DN and, thus, is a new target for the prevention and treatment of diabetic complications.

Cyclin G2 Is Involved in the Proliferation of Placental Trophoblast Cells and Their Interactions with Endothelial Cells.

BACKGROUND Remodeling of maternal spiral arteries after embryo implantation relies on well-regulated trophoblast functions. Although cyclin G2 (CCNG2) is thought to be involved in placental development and function, its role in trophoblasts and the mechanisms underlying placental development and function remain unclear. The present study investigated the potential role of CCNG2 in trophoblast cell proliferation and their interactions with endothelial cells. MATERIAL AND METHODS CCNG2 levels were modified by stable infection of HTR8/SVneo cells with lentiviruses overexpressing and silencing CCNG2. Cell proliferation was measured using CCK-8 assays. Network formation assays were performed using trophoblasts alone and co-cultured trophoblasts and endothelial cells to measure angiogenesis of trophoblasts and trophoblast-endothelial interactions. Levels of angiogenic factors (VEGF and sFlt-1) in the supernatant were measured by ELISA, and the expression of cell cycle regulatory (cyclin D1) and invasive (MMP2, MMP3, MMP9) markers implicated in artery remodeling were measured by western blotting. RESULTS Ectopic expression of CCNG2 blocked the proliferation of HTR8/SVneo cells, as well as their abilities to form networks and integrate into human umbilical vein endothelial cells, whereas CCNG2 inhibition had the opposite effects. CCNG2 upregulation significantly reduced the expression of VEGF, cyclin D1, MMP2, MMP3, and MMP9, but enhanced the expression of sFlt-1. In contrast, CCNG2 downregulation had the opposite effects. CONCLUSIONS CCNG2 plays a critical role in trophoblast proliferation and trophoblast-endothelial cell interactions by significant affecting cell cycle, angiogenic, and invasive markers. CCNG2 may thus be a novel marker for the treatment of placental disorders.

MYB75 Phosphorylation by MPK4 Is Required for Light-Induced Anthocyanin Accumulation in Arabidopsis.

Light is a major environmental cue affecting various physiological and metabolic processes in plants. Although plant photoreceptors are well characterized, the mechanisms by which light regulates downstream responses are less clear. In Arabidopsis thaliana, the accumulation of photoprotective anthocyanin pigments is light dependent, and the R2R3 MYB transcription factor MYB75/PAP1 regulates anthocyanin accumulation. Here, we report that MYB75 interacts with and is phosphorylated by MAP KINASE4 (MPK4). Their interaction is dependent on MPK4 kinase activity and is required for full function of MYB75. MPK4 can be activated in response to light and is involved in the light-induced accumulation of anthocyanins. We show that MPK4 phosphorylation of MYB75 increases its stability and is essential for light-induced anthocyanin accumulation. Our findings reveal an important role for a MAPK pathway in light signal transduction.

Direct and tunable modulation of protein levels in rice and wheat with a synthetic small molecule.

Direct control of protein level enables rapid and efficient analyses of gene functions in crops. Previously, we developed the RDDK-Shield1 (Shld1) system in the model plant Arabidopsis thaliana for direct modulation of protein stabilization using a synthetic small molecule. However, it was unclear whether this system is applicable to economically important crops. In this study, we show that the RDDK-Shld1 system enables rapid and tunable control of protein levels in rice and wheat. Accumulation of RDDK fusion proteins can be reversibly and spatio-temporally controlled by the synthetic small-molecule Shld1. Moreover, RDDK-Bar and RDDK-Pid3 fusions confer herbicide and rice blast resistance, respectively, in a Shld1-dependent manner. Therefore, the RDDK-Shld1 system provides a reversible and tunable technique for controlling protein functions and conditional expression of transgenes in crops.

MAGE-RING protein complexes comprise a family of E3 ubiquitin ligases.

The melanoma antigen (MAGE) family consists of more than 60 genes, many of which are cancer-testis antigens that are highly expressed in cancer and play a critical role in tumorigenesis. However, the biochemical and cellular functions of this enigmatic family of proteins have remained elusive. Here, we identify really interesting new gene (RING) domain proteins as binding partners for MAGE family proteins. Multiple MAGE family proteins bind E3 RING ubiquitin ligases with specificity. The crystal structure of one of these MAGE-RING complexes, MAGE-G1-NSE1, reveals structural insights into MAGE family proteins and their interaction with E3 RING ubiquitin ligases. Biochemical and cellular assays demonstrate that MAGE proteins enhance the ubiquitin ligase activity of RING domain proteins. For example, MAGE-C2-TRIM28 is shown to target p53 for degradation in a proteasome-dependent manner, consistent with its tumorigenic functions. These findings define a biochemical and cellular function for the MAGE protein family.

Characterisation of human RING finger protein TRIM69, a novel testis E3 ubiquitin ligase and its subcellular localisation.

The E3 ubiquitin ligase activity and subcellular localisation of human TRIM69 (hTRIM69) gene were studied. It was found that hTRIM69 mediated ubiquitination in an E2 conjugating enzyme selective fashion in vitro and an intact RING finger domain was indispensible for the process. Further evidences showed that hTRIM69 could mediate ubiquitination in vivo, which could be enhanced by a proteasome inhibitor. hTRIM69 was found to localise in both the cytoplasm and the nucleus in a speckled aggregating pattern, which also required an intact RING finger domain. Collectively, hTRIM69 is a novel E3 ubiquitin ligase identified from human testis and may function to ubiquitinate its particular substrates during spermatogenesis.

Purification and crystallization of the ABC-type transport substrate-binding protein OppA from Thermoanaerobacter tengcongensis.

Di- and oligopeptide- binding protein OppAs play important roles in solute and nutrient uptake, sporulation, biofilm formation, cell wall muropeptides recycling, peptide-dependent quorum-sensing responses, adherence to host cells, and a variety of other biological processes. Soluble OppA from Thermoanaerobacter tengcongensis was expressed in Escherichia coli. The protein was found to be >95% pure with SDS-PAGE after a series of purification steps and the purity was further verified by mass spectrometry. The protein was crystallized using the sitting-drop vapour-diffusion method with PEG 400 as the precipitant. Crystal diffraction extended to 2.25 Å. The crystal belonged to space group C222(1), with unit-cell parameters of a=69.395, b=199.572, c=131.673 Å, and α=ß=γ=90°.

Purification, crystallization and preliminary X-ray crystallographic analysis of the CIDE-N domain of Fsp27.

Fsp27, a member of the CIDE protein family which is selectively expressed in adipocytes, has emerged as a novel regulator for unilocular lipid droplet (LD) formation, lipid metabolism, differentiation of adipocytes and insulin sensitivity. An LD is a subcellular compartment that is used by adipocytes for the efficient storage of fats. The CIDE-N domain of Fsp27 functions as a recruitment platform that induces the correct configuration of the Fsp27 CIDE-C domain to facilitate LD fusion. This study reports the high-yield expression of the mouse Fsp27 CIDE-N domain in Escherichia coli; a two-step purification protocol with high efficiency was established and crystallographic analysis was performed. The purity of the recombinant Fsp27 was >95% as assessed by SDS-PAGE. Crystals were obtained at 291 K using 28% polyethylene glycol 4000 as a precipitant. Diffraction data were collected to 1.92 Šresolution and the crystal belonged to space group P6(5), with unit-cell parameters a=b=63.3, c=37.4 Å, α=ß=90, γ=120°. The components of the crystal were identified by ion-trap LC/MS/MS spectrometric analysis. The structure has been solved by molecular replacement and refinement is in progress.

Cyclin G2 in macrophages triggers CTL-mediated antitumor immunity and antiangiogenesis via interferon-gamma.

BACKGROUND: IFN-γ is a key mediator of tumor immunity that can induce macrophage polarization to suppress tumor growth. Cyclin G2 functions as a tumor suppressor in various cancer cells; however, its role in macrophages remains unclear. This study aimed to investigate the role and underlying mechanisms of cyclin G2 in macrophages in vitro and in vivo. METHODS: Mouse tumor models were used to determine the effect of cyclin G2 in macrophages on tumor growth in vivo following IFN-γ treatment. Immunohistochemistry staining, immunofluorescence staining and flow cytometry were used to evaluate the number of cytotoxic T lymphocytes (CTLs) and blood vessels in the mouse tumors. Moreover, the biological roles of cyclin G2 in macrophages with regard to CTL chemotaxis, cytotoxic function, and vascular endothelial cell tube formation were assessed using in vitro functional experiments. Immunoprecipitation (IP), real-time PCR, and enzyme-linked immunosorbent assays (ELISAs) were conducted to investigate the underlying mechanisms by which cyclin G2 regulates CTLs and vascular endothelial cells. RESULTS: We found that cyclin G2 expression was upregulated in macrophages after IFN-γ treatment. Upregulated cyclin G2 inhibited lung and colon cancer growth by increasing the secretion of its downstream effector CXCL9, which promoted CTL chemotaxis and suppressed vascular endothelial cell tube formation. Moreover, cyclin G2 increased CXCL9 mRNA levels by promoting STAT1 nuclear translocation. In addition, cyclin G2 promoted the activation of the STAT1 signaling pathway, which was dependent on PP2Ac. CONCLUSIONS: Cyclin G2 is upregulated by IFN-γ in macrophages, promotes the secretion of CXCL9 to increase CTL chemotaxis and inhibit angiogenesis to suppress tumor growth. Our findings suggest that targeting cyclin G2 could benefit future immunotherapy.

Highly efficient genome editing in Xanthomonas oryzae pv. oryzae through repurposing the endogenous type I-C CRISPR-Cas system.

Efficient and modular genome editing technologies that manipulate the genome of bacterial pathogens will facilitate the study of pathogenesis mechanisms. However, such methods are yet to be established for Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of rice bacterial blight. We identified a single type I-C CRISPR-Cas system in the Xoo genome and leveraged this endogenous defence system for high-efficiency genome editing in Xoo. Specifically, we developed plasmid components carrying a mini-CRISPR array, donor DNA, and a phage-derived recombination system to enable the efficient and programmable genome editing of precise deletions, insertions, base substitutions, and gene replacements. Furthermore, the type I-C CRISPR-Cas system of Xoo cleaves target DNA unidirectionally, and this can be harnessed to generate large genomic deletions up to 212 kb efficiently. Therefore, the genome-editing strategy we have developed can serve as an excellent tool for functional genomics of Xoo, and should also be applicable to other CRISPR-harbouring bacterial plant pathogens.

Cyclin G2 reverses immunosuppressive tumor microenvironment and potentiates PD-1 blockade in glioma.

BACKGROUND: Expression of aberrant cyclin G2 is a key factor contributing to cancer biological processes, including glioma. However, the potential underlying mechanisms of cyclin G2 in the glioma tumor immune microenvironment remain unclear. METHODS: Co-immunoprecipitation (co-IP), in situ proximity ligation assay (PLA), and in vitro kinase assay were conducted to reveal the underlying mechanism by which cyclin G2 regulates Y10 phosphorylation of LDHA. Further, the biological roles of cyclin G2 in cell proliferation, migration, invasion capacity, apoptosis, glycolysis, and immunomodulation were assessed through in vitro and in vivo functional experiments. Expressions of cyclin G2 and Foxp3 in glioma specimens was determined by immunohistochemistry. RESULTS: In this study, we found that cyclin G2 impeded the interaction between LDHA and FGFR1, thereby decreasing Y10 phosphorylation of LDHA through FGFR1 catalysis. Cyclin G2 inhibited proliferation, migration, invasion capacity, and glycolysis and promoted apoptosis glioma cells via suppressing Y10 phosphorylation of LDHA. Moreover, we further verified that cyclin G2 reversed the immunosuppressive to antitumor immune microenvironment through inhibiting lactate production by glioma cells. Besides, cyclin G2 potentiated PD-1 blockade and exerted strong antitumor immunity in the glioma-bearing mice model. CONCLUSIONS: Cyclin G2 acts as a potent tumor suppressor in glioma and enhances responses to immunotherapy. Our findings may be helpful in selecting glioma patients for immunotherapy trials in the future.

Cyclin G2 promotes the formation of smooth muscle cells derived foam cells in atherosclerosis via PP2A/NF-κB/LOX-1 pathway.

BACKGROUND: To investigate the role and underlying mechanism of cyclin G2 (G2-type cyclin) in the formation of vascular smooth muscle cells (VSMCs) derived foam cells. METHODS: The levels of α-SMA (alpha-SM-actin), p-NF-κB (phosphorylation nuclear transcription factors kappa B), and LOX-1 (lectin-like oxidized low-density lipoprotein receptor-1) were measured by immunohistochemistry and western blotting. The mouse aortic root smooth muscle cell line MOVAS was transfected to over-express cyclin G2, which were then stimulated with 80 µg/mL ox-LDL (oxidized low-density lipoprotein) to induce foam cell formation. DT-061 an activator of PP2A (protein phosphatase 2A) agonist was used to verify the role of PP2A in the process. RESULTS: Knocking out the Ccng2 gene in Apoe-/- mice alleviated aortic lipid plaque, foam cell formulation, ameliorative body weight, and LDL-cholesterol. We observed that the number of α-SMA positive cells was significantly decreased in Apoe-/-Ccng2-/- mice compared to Apoe-/- mice. Also, the protein levels of p-NF-κB and LOX-1 were markedly reduced in the aortic root of Apoe-/-Ccng2-/- mice. Upon stimulation with ox-LDL, upregulated cyclin G2 increased the intracellular lipid accumulation in MOVAS cells. Also, it suppressed the activity of PP2A but up-regulated LOX-1. Additionally, the cell nuclear translocation of p-NF-κB was increased. Interestingly, DT-061 intervention, re-activating the activity of PP2A, reduced the levels of nuclear p-NF-κB and LOX-1. This led to decreased lipid endocytosis reducing the formation of VSMCs- derived foam cells. CONCLUSIONS: Cyclin G2 increases the nuclear translocation of p-NF-κB by reducing the enzymatic activity of PP2A and upregulating LOX-1, thereby promotes the formation of VSMCs -derived foam cells in atherosclerosis.

Purification, characterization, and crystallization of the adhesive domain of SdrD from Staphylococcus aureus.

The adhesive domain of SdrD from Staphylococcus aureus was solubly expressed in Escherichia coli in high yield. After a series of purification steps, the purified protein was >95% pure, which was SdrD from S. aureus identified by SDS-PAGE and MALDI-TOF MS. Crystals were grown at 18 degrees C using 25% polyethylene glycol 3350 as precipitant. Diffraction by the crystal extends to 1.65A resolution, and the crystal belongs to the space group C2, with the unit cell parameters a=133.3, b=58.3, c=112.3A, alpha=90.00, beta=111.14, gamma=90.00.

Cyclin G2 Inhibits Oral Squamous Cell Carcinoma Growth and Metastasis by Binding to IGFBP3 and Regulating the FAK-SRC-STAT Signaling Pathway.

The cell cycle protein cyclin G2 is considered a tumor suppressor. However, its regulatory effects and potential mechanisms in oral cancers are not well understood. This study aimed to investigate the effect of cyclin G2 on oral squamous cell carcinoma (OSCC). The data from 80 patients with OSCC were utilized to predict the abnormal expression of cyclin G2. The proliferation and metastasis were determined by a cell counting Kit-8 assay, flow cytometry, a wound-healing assay, and a cell invasion assay. The expression of key proteins and genes associated with the cyclin G2 signaling pathways was determined by western blotting and real-time PCR, respectively. The orthotopic nude mice model was established by a mouth injection of SCC9 cells overexpressing cyclin G2. We showed that the low level of cyclin G2 in OSCC, which is negatively correlated with clinical staging, was a negative prognostic factor for the disease. We also found that cyclin G2 inhibited the proliferation, metastasis, and blocked the cell cycle at G1/S of OSCC cells, suggesting that cyclin G2 has an inhibitory effect in OSCC. Mechanistically, cyclin G2 inhibited the growth and metastasis of OSCC by binding to insulin-like growth factor binding protein 3 (IGFBP3) and regulating the focal adhesion kinase (FAK) -SRC-STAT signal transduction pathway. Cyclin G2 competed with integrin to bind to IGFBP3; the binding between integrin and IGFBP3 was reduced after cyclin G2 overexpression, thereby inhibiting the phosphorylation of FAK and SRC. These results showed that cyclin G2 inhibited the progression of OSCC by interacting with IGFBP3 and that it may be a new target for OSCC treatment.

To control or to be controlled? Dual roles of CDK2 in DNA damage and DNA damage response.

CDK2 (cyclin-dependent kinase 2), a member of the CDK family, has been shown to play a role in many cellular activities including cell cycle progression, apoptosis and senescence. Recently, accumulating evidence indicates that CDK2 is involved in DNA damage and DNA repair response (DDR). When DNA is damaged by internal or external genotoxic stresses, CDK2 activity is required for proper DNA repair in vivo and in vitro, whereas inactivation of CDK2 by siRNA techniques or by inhibitors could result in DNA damage and stimulate DDR. Hence, CDK2 seems to play dual roles in DNA damage and DDR. On one aspect, it is activated and stimulates DDR to repair DNA damage when DNA damage occurs; on the other hand, its inactivation directly leads to DNA damage and evokes DDR. Here, we describe the roles of CDK2 in DNA damage and DDR, and discuss the potential application of CDK2 inhibitors as anti-cancer agents.

Cyclin G2 Inhibits the Warburg Effect and Tumour Progression by Suppressing LDHA Phosphorylation in Glioma.

Cyclin G2 has been identified as a tumour suppressor in several cancers. However, its regulatory roles and underlying mechanisms in tumours are still unknown. In this study, we demonstrated that cyclin G2 was expressed at low levels in glioma, which was as a poor prognostic factor for this disease. We also found that, cyclin G2 could suppress cell proliferation, initiate cell apoptosis and reduce aerobic glycolysis, suggesting that cyclin G2 plays a tumour suppressive role in glioma. Mechanistically, cyclin G2 could negatively regulate tyrosine-10 phosphorylation of a critical glycolytic enzyme, lactate dehydrogenase A, through direct interaction. Taken together, these results indicate that cyclin G2 acts as a tumour suppressor in glioma by repressing glycolysis and tumour progression through its interaction with LDHA.

Corrigendum to "Cyclin G2 Suppresses Glomerulosclerosis by Regulating Canonical Wnt Signalling".

[This corrects the article DOI: 10.1155/2018/6938482.].

Structure and Function of the Photosystem Supercomplexes.

Photosynthesis converts solar energy into chemical energy to sustain all life on earth by providing oxygen and food, and controlling the atmospheric carbon dioxide. During this process, the water-splitting and oxygen-evolving reaction is catalyzed by photosystem II (PSII), while photosystem I (PSI) generates the reducing power for the reduction of NADP+ to NADPH. Together with their peripheral light-harvesting complexes (LHCs), photosystems function as multisubunit supercomplexes located in the thylakoid membranes of cyanobacteria, algae, and plants. Recent advances in single-particle cryo-electron microscopy (cryoEM), X-ray free electron laser (XFEL) and other techniques have revealed unprecedented structural and catalytic details concerning the two supercomplexes. Several high-resolution structures of the complexes from plants were solved, and serial time-resolved crystallography and "radiation-damage-free" femtosecond XFEL also provided important insights into the mechanism of water oxidation. Here, we review these exciting advances in the studies of the photosystem supercomplexes with an emphasis on PSII-LHCII, propose presently unresolved problems in this field, and suggest potential tendencies for future studies.