Tumor immune dysfunction and exclusion subtypes in bladder cancer and pan-cancer: a novel molecular subtyping strategy and immunotherapeutic prediction model.

BACKGROUND: Molecular subtyping is expected to enable precise treatment. However, reliable subtyping strategies for clinical application remains defective and controversial. Given the significance of tumor immune dysfunction and exclusion (TIDE), we aimed to develop a novel TIDE-based subtyping strategy to guide personalized immunotherapy in the bladder cancer (BC). METHODS: Transcriptome data of BC was used to evaluate the heterogeneity and the status of TIDE patterns. Subsequently, consensus clustering was applied to classify BC patients based on TIDE marker-genes. Patients' clinicopathological, molecular features and signaling pathways of the different TIDE subtypes were well characterized. We also utilize the deconvolution algorithms to analyze the tumor microenvironment, and further explore the sensitivity and mechanisms of each subtype to immunotherapy. Furthermore, BC patient clinical information, real-world BC samples and urine samples were collected for the validation of our findings, which were used for RNA-seq analysis, H&E staining, immunohistochemistry and immunofluorescence staining, and enzyme-linked immunosorbent assay. Finally, we also explored the conservation of our novel TIDE subtypes in pan-cancers. RESULTS: We identified 69 TIDE biomarker genes and classified BC samples into three subtypes using consensus clustering. Subtype I showed the lowest TIDE status and malignancy with the best prognosis and highest sensitivity to immune checkpoint blockade (ICB) treatment, which was enriched of metabolic related signaling pathways. Subtype III represented the highest TIDE status and malignancy with the poorest prognosis and resistance to ICB treatment, resulting from its inhibitory immune microenvironment and T cell terminal exhaustion. Subtype II was in a transitional state with intermediate TIDE level, malignancy, and prognosis. We further confirmed the existence and characteristics of our novel TIDE subtypes using real-world BC samples and collected patient clinical data. This subtyping method was proved to be more efficient than previous known methods in identifying non-responders to immunotherapy. We also propose that combining our TIDE subtypes with known biomarkers can potentially improve the sensitivity and specificity of these biomarkers. Moreover, besides guiding ICB treatment, this classification approach can assist in selecting the frontline or recommended drugs. Finally, we confirmed that the TIDE subtypes are conserved across the pan-tumors. CONCLUSIONS: Our novel TIDE-based subtyping method can serve as a powerful clinical tool for BC and pan-cancer patients, and potentially guiding personalized therapy decisions for selecting potential beneficiaries and excluding resistant patients of ICB therapy.

Construction of UCNPs-aptamer-AuNPs luminescence energy transfer probe for ratio detection of Staphylococcus aureus.

A ratio luminescence probe was developed for detecting Staphylococcus aureus (S. aureus) based on luminescence energy transfer (LET) using double-wavelength emission (550 nm and 812 nm) upconversion nanoparticles (UCNPs) as donor, gold nanoparticles (AuNPs) as acceptor and the aptamer for S. aureus as the specific recognition and link unit. The LET process could cause luminescence quenching because of the spectral overlap between the acceptor and the donor at 550 nm. In the presence of S. aureus, S. aureus selectively combined with the aptamer, and the AuNPs left the surface of UCNPs, which weakened the quenching effect and restored the luminescence of UCNPs. Based on this, the ratio detection was realized by monitoring the change of the luminescence signal of the probe at 550 nm and taking the luminescence signal at 812 nm as the reference signal. Crucially, the probe has a fast reaction speed, with a reaction time of 25 min, and the detection of S. aureus is realized in the concentration range of 5.0 × 103-3.0 × 105 CFU/ml, with the detection limit of 106 CFU/ml. Therefore, the ratio probe has great potential for detecting of S. aureus in food because of its high sensitivity, fast speed and good selectivity.

Cyanine dye-assembled composite upconversion nanoparticles for the sensing and cell imaging of nitrite based on a single particle imaging method.

In this work, a single particle imaging method based on the total internal reflection (TIR) imaging platform for the sensing and cell imaging of nitrite (NO2-) in the near-infrared region using cyanine dye-assembled composite upconversion nanoparticles (UCNPs) was developed. The NaYF4:Yb,Tm@NaGdF4 UCNPs were synthesized as energy donors, and the cyanine dye IR-798 was prepared as an energy acceptor. Since the absorption spectrum of the cyanine dye IR-798 and the luminescence spectrum of upconversion nanoparticles overlapped effectively, IR-798 quenched the luminescence of the UCNPs. When NO2- was added to the cyanine dye-assembled composite upconversion nanoparticle system, NO2- destroyed the conjugate structure of IR-798, so that the luminous intensity of UCNPs could be restored. Based on the mechanism, a quantitative image analysis with high sensitivity, low sample usage, and fast response time using the TIR single particle imaging platform was developed to determine the content of nitrite in human serum samples. In addition, the UCNPs-IR-798 probe was applied to image the exogenous NO2- content in HeLa living cells based on the single particle imaging platform.

A hybrid boronate affinity probe for the selective detection of cis-diol containing compounds in tea beverages.

UiO-66-NH2 nanocomposite was post-modified with 4-mercaptophenylboronic acid (MPBA) by the method of in situ hybridization reaction. The hybrid boronate affinity material UiO-NH2 @P (TEPIC-co-MPBA) was characterized by scanning electron microscopy, X-ray diffraction and Fourier-transform infrared spectroscopy. The material was applied as fluorescent probe for the detection of cis-diol containing compounds based on the boronate affinity mechanism, and exhibited high specific selectively. The proposed method exhibited good linearity for the detection of catechol in the range of 0.50 to 8.00 µg ml-1 . The detection limit was 0.13 µg ml-1 . The tactic was successfully applied to analyze the total polyphenols in tea beverages for catechol, and relative recovery was in 98.86-106.00%. Therefore, this work provided a promising strategy for the recognition of cis-diol containing compounds.

OsWUS promotes tiller bud growth by establishing weak apical dominance in rice.

Two branching strategies are exhibited in crops: enhanced apical dominance, as in maize; or weak apical dominance, as in rice. However, the underlying mechanism of weak apical dominance remains elusive. OsWUS, an ortholog of Arabidopsis WUSCHEL (WUS) in rice, is required for tiller development. In this study, we identified and functionally characterized a low-tillering mutant decreased culm number 1 (dc1) that resulted from loss-of-function of OsWUS. The dc1 tiller buds are viable but repressed by the main culm apex, leading to stronger apical dominance than that of the wild-type (WT). Auxin response is enhanced in the dc1 mutant, and knocking out the auxin action-associated gene ABERRANT SPIKELET AND PANICLE 1 (ASP1) de-repressed growth of the tiller buds in the dc1 mutant, suggesting that OsWUS and ASP1 are both involved in outgrowth of the rice tiller bud. Decapitation triggers higher contents of cytokinins in the shoot base of the dc1 mutant compared with those in the WT, and exogenous application of cytokinin is not sufficient for sustained growth of the dc1 tiller bud. Transcriptome analysis indicated that expression levels of transcription factors putatively bound by ORYZA SATIVA HOMEOBOX 1 (OSH1) are changed in response to decapitation and display a greater fold change in the dc1 mutant than that in the WT. Collectively, these findings reveal an important role of OsWUS in tiller bud growth by influencing apical dominance, and provide the basis for an improved understanding of tiller bud development in rice.

IL-36γ-armed oncolytic virus exerts superior efficacy through induction of potent adaptive antitumor immunity.

In this study, we aimed to apply the cytokine IL-36γ to cancer immunotherapy by constructing new oncolytic vaccinia viruses (OV) expressing interleukin-36γ (IL-36γ-OVs), leveraging unique synergism between OV and IL-36γ's ability to promote antitumor adaptive immunity and modulate tumor microenvironment (TME). IL-36γ-OV had dramatic therapeutic efficacies in multiple murine tumor models, frequently leading to complete cancer eradication in large fractions of mice. Mechanistically, IL-36-γ-armed OV induced infiltration of lymphocytes and dendritic cells, decreased myeloid-derived suppressor cells and M2-like tumor-associated macrophages, and T cell differentiation into effector cells. Further study showed that IL-36γ-OV increased the number of tumor antigen-specific CD4+ and CD8+ T cells and the therapeutic efficacy depended on both CD8+ and CD4+ T cells. These results demonstrate that these IL36γ-armed OVs exert potent therapeutic efficacy mainly though antitumor immunity and they may hold great potential to advance treatment in human cancer patients.

Sensitive detection of sulfide ions in red region based on luminescence resonance energy transfer between upconversion nanoparticles and dye-670.

Many sulfides are toxic substances that easily harm the respiratory tract, therefore affecting respiratory function or damaging other organs of the body, leading to its failure. Therefore, there is a pressing need to develop methods for sensitive detection of sulfur ions (S2- ). Based on luminescence resonance energy transfer (LRET) theory, we report the construction of a near-infrared (NIR) excitation luminescence probe using NaGdF4 :Yb3+ ,Er3+ @NaYF4 upconversion nanoparticles (UCNPs) as the donor and dye-670 as the receptor for detection of S2- . When UCNPs and dye-670 molecules were combined using ligand exchange and electrostatic attraction, LRET occurred and UCNP luminescence was quenched. When S2- was added to the system, sulfide ions were able to destroy the double bond of the dye, inhibiting LRET and restoring UCNP luminescence. Under optimum condition, the linear range of S2- detection was 0.65-18.2 µM, and the detection limit was 34.2 nM. This method was applied for determination of S2- in water with satisfactory results.

A single-particle enumeration method for the detection of Fe2+ based on a near-infrared core-shell upconversion nanoparticle and IR-808 dye composite nanoprobe.

Ferrous ion (Fe2+) is an important component of hemoglobin and plays a role in transporting O2 to human tissues. If iron deficiency is present, iron deficiency anemia may occur, so it is critical to develop sensitive and accurate methods to detect Fe2+. Herein, a novel luminescence energy transfer (ET) system has been designed for the sensitive detection of Fe2+ by a single-particle enumeration (SPE) method in the near-infrared (NIR) region through combining NIR-to-NIR ß-NaGdF4:Yb,Tm@NaYF4 upconversion nanoparticles (UCNPs) and IR-808 dye. IR-808 dye can quench the luminescence of the UCNPs because of the efficient overlap between the absorption spectrum of IR-808 and the emission spectrum of the UCNPs. When Fe2+ and H2O2 are added to the system, the Fenton reaction produces hydroxyl radicals (˙OH). The generated ˙OH reacts with IR-808 and the structure of IR-808 is destroyed. As a result, the ET process is suppressed, causing recovery of the luminescence of the UCNPs, which is reflected as an increase in the number of luminescent particles. Accurate quantification of Fe2+ is achieved by statistically counting the target concentration-dependent luminescent particles. Under the optimal conditions, the linear detection range of Fe2+ is 5-10 000 nM, which is much wider than the ensemble luminescence spectra measurements in bulk solution. Moreover, this strategy can be applied to detection in serum samples with satisfactory results.

Turn-on detection of glutathione S-transferase based on luminescence resonance energy transfer between near-infrared to near-infrared core-shell upconversion nanoparticles and organic dye.

Glutathione S-transferase (GST) is a detoxification enzyme of the liver and kidney. Based on the toxicological effect of GST, it is of great significance to develop a rapid and sensitive detection method for GST. In this work, a new luminescence resonance energy transfer (LRET) system has been designed to detect glutathione S-transferase in the near-infrared (NIR) region by utilizing NaGdF4:Yb3+,Tm3+@NaYF4 upconversion nanoparticles (UCNPs) as the donor and NIR dye-806@Glutathione (IR806@GSH) as the acceptor. NaGdF4:Yb3+,Tm3+@NaYF4 UCNPs were synthesized by a coprecipitation method and surface modification of NOBF4. The donor (positively charged) interacted with the acceptor (negatively charged) via electrostatic interactions to bring them into close proximity; then, LRET occurred and the luminescence was quenched. In the presence of GST, GST can specifically interact with the GSH of IR806@GSH molecule, making IR806@GSH far away from the donor surface, inhibiting the LRET, and restoring the luminescence of the UCNPs. There was a good linear relationship between the luminescence recovery intensity of UCNPs and GST concentration, ranging from 0.11 to 14.19 nM, and the detection of limit was 0.06 nM. The method has been used in the detection of GST in human serum samples and is expected to have potential applications in the biological field. Graphical abstract A luminescence resonance energy transfer system was developed for determination of glutathione S-transferase in the near-infrared region by utilizing NaGdF4:Yb3+,Tm3+@NaYF4 upconversion nanoparticles as the donor and NIR dye-806@Glutathione as the acceptor.

Gut microbiota dysbiosis signature is associated with the colorectal carcinogenesis sequence and improves the diagnosis of colorectal lesions.

BACKGROUND AND AIM: The gut microbiota is associated with colorectal lesions in cases of precancer and colorectal cancer (CRC). However, there are apparent differences in studies on the gut microbiota in the pathogenic sequence from precancer to cancer. Here, we characterize the gut microbiota signatures of colorectal precancer and cancer and test their utility in detecting colorectal lesions in two independent Chinese cohorts. METHODS: Stool samples collected from patients with precancer and CRC were subjected to 16S ribosomal RNA gene sequencing and metagenomic shotgun sequencing analyses, which revealed the microbial signatures of the two disease stages. RESULTS: In comparison with healthy controls, lower microbial richness and diversity were observed in precancer and intensive interbacterial associations were found in CRC. We identified 41 bacteria that showed gradual increases while 12 bacteria showed gradual decreases at the genus level gradually during the development of CRC. Novel CRC-associated pathogenetic species were identified. Species units that contributed to altered microbial functions were identified in CRC patients and healthy controls. The microbial panel showed a comparable ability to fecal immunochemical test (FIT) in detecting CRC. However, the combination of microbes and FIT significantly improved the detection ability and sensitivity of colon lesions based on 18 genera. Microbial network analysis revealed a significant positive correlation among beneficial microbes and a negative correlation in detrimental phenotypes. CONCLUSIONS: Microbial dysbiosis was revealed in colorectal lesions. The combination of microbial markers and FIT improved the CRC detection ability, which might assist in the early diagnosis of CRC.

Development of Robust Yeast Strains for Lignocellulosic Biorefineries Based on Genome-Wide Studies.

Lignocellulosic biomass has been widely studied as the renewable feedstock for the production of biofuels and biochemicals. Budding yeast Saccharomyces cerevisiae is commonly used as a cell factory for bioconversion of lignocellulosic biomass. However, economic bioproduction using fermentable sugars released from lignocellulosic feedstocks is still challenging. Due to impaired cell viability and fermentation performance by various inhibitors that are present in the cellulosic hydrolysates, robust yeast strains resistant to various stress environments are highly desired. Here, we summarize recent progress on yeast strain development for the production of biofuels and biochemical using lignocellulosic biomass. Genome-wide studies which have contributed to the elucidation of mechanisms of yeast stress tolerance are reviewed. Key gene targets recently identified based on multiomics analysis such as transcriptomic, proteomic, and metabolomics studies are summarized. Physiological genomic studies based on zinc sulfate supplementation are highlighted, and novel zinc-responsive genes involved in yeast stress tolerance are focused. The dependence of host genetic background of yeast stress tolerance and roles of histones and their modifications are emphasized. The development of robust yeast strains based on multiomics analysis benefits economic bioconversion of lignocellulosic biomass.

CFTR activation suppresses glioblastoma cell proliferation, migration and invasion.

The aim of this study was to investigate the function of Cystic fibrosis transmembrane conductance regulator (CFTR) in human glioblastoma (GBM) cells. Data dining results of the Human Protein Atlas showed that low CFTR expression was associated with poor prognosis for GBM patients. We found that CFTR protein expression was lower in U87 and U251 GBM cells than that in normal humane astrocyte cells. CFTR activation significantly reduced GBM cell proliferation. In addition, CFTR activation significantly abrogated migration and invasion of GBM cells. Besides, CFTR activator Forskolin treatment markedly reduced MMP-2 protein expression. These effects of CFTR activation were significantly inhibited by CFTR inhibitor CFTRinh-172 pretreatment. Our findings suggested that JAK2/STAT3 signaling was involved in the anti-glioblastoma effects of CFTR activation. Moreover, CFTR overexpression in combination with Forskolin induced a synergistic anti-proliferative response in U87 cells. Overall, our findings demonstrated that CFTR activation suppressed GBM cell proliferation, migration and invasion likely through the inhibition of JAK2/STAT3 signaling.

Improvement of inhibitor tolerance in Saccharomyces cerevisiae by overexpression of the quinone oxidoreductase family gene YCR102C.

Budding yeast Saccharomyces cerevisiae is widely used for lignocellulosic biorefinery. However, its fermentation efficiency is challenged by various inhibitors (e.g. weak acids, furfural) in the lignocellulosic hydrolysate, and acetic acid is commonly present as a major inhibitor. The effects of oxidoreductases on the inhibitor tolerance of S. cerevisiae have mainly focused on furfural and vanillin, whereas the influence of quinone oxidoreductase on acetic acid tolerance is still unknown. In this study, we show that overexpression of a quinone oxidoreductase-encoding gene, YCR102C, in S. cerevisiae, significantly enhanced ethanol production under acetic acid stress as well as in the inhibitor mixture, and also improved resistance to simultaneous stress of 40°C and 3.6 g/L acetic acid. Increased catalase activities, NADH/NAD+ ratio and contents of several metals, especially potassium, were observed by YCR102C overexpression under acetic acid stress. To our knowledge, this is the first report that the quinone oxidoreductase family protein is related to acid stress tolerance. Our study provides a novel strategy to increase lignocellulosic biorefinery efficiency using yeast cell factory.

Core-shell upconversion nanoparticles of type NaGdF4:Yb,Er@NaGdF4:Nd,Yb and sensitized with a NIR dye are a viable probe for luminescence determination of the fraction of water in organic solvents.

Lanthanide-doped core-shell upconversion nanoparticles (UCNPs) of type NaGdF4:Yb,Er@NaGdF4:Yb,Nd were prepared by the co-precipitation method. The luminescence intensity was further enhanced by adding the sensitizer dye IR-808. If water is added to organic solvents [such as N,N-dimethylformamide (DMF), dimethyl sulfoxide, methanol, acetone, acetonitrile, and ethanol] containing the probe, its luminescence intensity peaking at 545 nm is reduced. The decrease is linearly related to the percentage of water in the respective organic solvent. Water fractions ranging from 0.05% to 10% (volume %) can be sensitively detected, and the detection limit is 0.018% of water in DMF. The detection scheme is mainly attributed to the fact that the transfer of energy from the near-infrared light (NIR) dye to the UCNPs is strongly reduced in the presence of traces of water. Graphical abstract The near infrared dye (IR-808) transfer efficiency to NaGdF4:Yb, Er@NaGdF4:Yb, Nd upconversion nanoparticles in water is far less than that in organic phase. Several methods for determination of trace water in organic solvents were developed by using this effect.

Redox luminescence switch based on Mn2+ -doped NaYF4 :Yb,Er upconversion nanorods.

An redox luminescence switch was developed for the sensing of glutathione (GSH), l-cysteine (Cys) or l-ascorbic acid (AA) based on redox reaction. The Mn2+ -doped NaYF4 :Yb,Er upconversion nanorods (UCNRs) with an emission peak located in the red region were synthesized. The luminescence intensity of the UCNRs could be quenched due to the Mn2+ could be oxidized to MnO2 by KMnO4 . Subsequently, when the AA, GSH or Cys was added into the MnO2 modified upconversion nanosystem, which could reduced MnO2 to Mn2+ and the luminescence intensity was recovered. The concentration ranges of the nanosystem are 0.500-3.375 mM (R2  = 0.99) for AA, 0.6250-11.88 mM (R2  = 0.99) for GSH and 0.6250-9.375 mM (R2  = 0.99) for Cys, respectively.

Near-infrared-emitting NaYF4:Yb,Tm/Mn upconverting nanoparticle/gold nanorod electrochemiluminescence resonance energy transfer system for sensitive prostate-specific antigen detection.

An electrochemiluminescence resonance energy transfer (ECL-RET) system that detects prostate-specific antigen (PSA) was created. Near-infrared (NIR)-emitting NaYF4:Yb,Tm/Mn upconverting nanoparticles (UCNPs) are used as donors, and gold nanorods (GNRs) are used as acceptors. The ECL was enhanced nearly threefold by Mn2+ doping, with an emission peak appearing at an NIR wavelength of 808 nm. Anti-PSA 1 (Ab1) was bound to the surfaces of UCNPs after being modified with poly(acrylic acid) (PAA). As for acceptors, cetyltrimethylammonium bromide (CTAB)-capped GNRs were treated with 11-mercaptoundecanoic acid (MUDA) and then conjugated with Anti-PSA 2 (Ab2). When PSA was added, donors and acceptors were brought in close proximity through specific interactions of antibodies and antigens, resulting in high quenching efficiency levels. Under optimal conditions, the linear range of detection was 3.75-938 pg/mL for PSA (R = 0.999), with a detection limit as low as 3.16 pg/mL. This method can be applied to detect PSA in human serums with satisfactory results. Graphical abstract An electrochemiluminescence resonance energy transfer system was developed for determination of prostate-specific antigen using near-infrared-emitting NaYF4:Yb,Tm/Mn upconverting nanoparticles as donors and gold nanorods as acceptors.

Knockdown of SLC39A7 inhibits cell growth and induces apoptosis in human colorectal cancer cells.

SLC39A7 (zip7) is a zinc transporter that plays a key role in intestinal epithelial self-renewal. However, little is known about SLC39A7 in colorectal cancer. To assess the biological function of SLC39A7 in colorectal cancer, the expression of SLC39A7 in human colorectal tumors and five colorectal cancer cell lines were evaluated by Oncomine Cancer Microarray Database and western blot analysis. In addition, short hairpin RNAs specifically targeting SLC39A7 were transfected into HCT116 and SW1116 cells to knockdown SLC39A7 expression. Then, the effects of SLC39A7 knockdown on colorectal cancer cells were detected by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide, colony-forming assay and flow cytometry. Our results showed that colorectal tumors have higher expression levels of SLC39A7 than normal colon tissues. Knockdown of SLC39A7 exhibited a significant decrease in cell viability and proliferation of colorectal cancer cells. It was also shown that knockdown of SLC39A7 interfered with cell cycle progression and induced G2/M cell cycle arrest, as well as boosted early and late apoptosis in colorectal cancer cells. Furthermore, downregulation of SLC39A7 promoted the cleavage of PARP and enhanced the expression of Bad, Caspase-9, and cleaved-Caspase-3, as well as suppressed Bcl-2 expression. In conclusion, our results suggest that SLC39A7 plays a crucial role in the proliferation and survival of colorectal cancer cells, which associates with colorectal tumorigenesis.

Novel evidence for an oncogenic role of microRNA-21 in colitis-associated colorectal cancer.

OBJECTIVE: miR-21 was found to be overexpressed in the colon tissues and serum of patients with UC and colorectal cancer (CRC); however, the exact roles of miR-21 in colitis-associated CRC remain unclear. The aim of our study was to investigate the biological mechanisms of miR-21 in colitis-associated colon cancer (CAC). DESIGN: miR-21 expression was examined in the tumours of 62 patients with CRC from China and 37 colitis-associated neoplastic tissues from Japan and Austria. The biological functions of miR-21 were studied using a series of in vitro, in vivo and clinical approaches. RESULTS: miR-21 levels were markedly upregulated in the tumours of 62 patients with CRC, 22 patients with CAC, and in a mouse model of CAC. Following azoxymethane and dextran sulfate sodium intervention, miR-21-knockout mice showed reduced expression of proinflammatory and procarcinogenic cytokines (interleukin (IL) 6, IL-23, IL-17A and IL-21) and a decrease in the size and number of tumours compared with the control mouse group. The absence of miR-21 resulted in the reduced expression of Ki67 and the attenuated proliferation of tumour cells with a simultaneous increase in E-cadherin and decrease in ß-catenin and SOX9 in the tumours of CAC mice. Furthermore, the absence of miR-21 increased the expression of its target gene PDCD4 and subsequently modulated nuclear factor (NF)-κB activation. Meanwhile, miR-21 loss reduced STAT3 and Bcl-2 activation, causing an increase in the apoptosis of tumour cells in CAC mice. CONCLUSIONS: These observations provide novel evidence for miR-21 blockade to be a key strategy in reducing CAC.

OsMAPK6, a mitogen-activated protein kinase, influences rice grain size and biomass production.

Grain size is an important agronomic trait in determining grain yield. However, the molecular mechanisms that determine the final grain size are not well understood. Here, we report the functional analysis of a rice (Oryza sativa L.) mutant, dwarf and small grain1 (dsg1), which displays pleiotropic phenotypes, including small grains, dwarfism and erect leaves. Cytological observations revealed that the small grain and dwarfism of dsg1 were mainly caused by the inhibition of cell proliferation. Map-based cloning revealed that DSG1 encoded a mitogen-activated protein kinase (MAPK), OsMAPK6. OsMAPK6 was mainly located in the nucleus and cytoplasm, and was ubiquitously distributed in various organs, predominately in spikelets and spikelet hulls, consistent with its role in grain size and biomass production. As a functional kinase, OsMAPK6 interacts strongly with OsMKK4, indicating that OsMKK4 is likely to be the upstream MAPK kinase of OsMAPK6 in rice. In addition, hormone sensitivity tests indicated that the dsg1 mutant was less sensitive to brassinosteroids (BRs). The endogenous BR levels were reduced in dsg1, and the expression of several BR signaling pathway genes and feedback-inhibited genes was altered in the dsg1 mutant, with or without exogenous BRs, indicating that OsMAPK6 may contribute to influence BR homeostasis and signaling. Thus, OsMAPK6, a MAPK, plays a pivotal role in grain size in rice, via cell proliferation, and BR signaling and homeostasis.

Rice TUTOU1 Encodes a Suppressor of cAMP Receptor-Like Protein That Is Important for Actin Organization and Panicle Development.

Panicle development, a key event in rice (Oryza sativa) reproduction and a critical determinant of grain yield, forms a branched structure containing multiple spikelets. Genetic and environmental factors can perturb panicle development, causing panicles to degenerate and producing characteristic whitish, small spikelets with severely reduced fertility and yield; however, little is known about the molecular basis of the formation of degenerating panicles in rice. Here, we report the identification and characterization of the rice panicle degenerative mutant tutou1 (tut1), which shows severe defects in panicle development. The tut1 also shows a pleiotropic phenotype, characterized by short roots, reduced plant height, and abnormal development of anthers and pollen grains. Molecular genetic studies revealed that TUT1 encodes a suppressor of cAMP receptor/Wiskott-Aldrich syndrome protein family verprolin-homologous (SCAR/WAVE)-like protein. We found that TUT1 contains conserved functional domains found in eukaryotic SCAR/WAVE proteins, and was able to activate Actin-related protein2/3 to promote actin nucleation and polymerization in vitro. Consistently, tut1 mutants show defects in the arrangement of actin filaments in trichome. These results indicate that TUT1 is a functional SCAR/WAVE protein and plays an important role in panicle development.