Molecular regulatory mechanisms of staminate strobilus development and dehiscence in Torreya grandis.

Gymnosperms are mostly dioecious, and their staminate strobili undergo a longer developmental period than those of angiosperms. However, the underlying molecular mechanisms remain unclear. This study aimed to identify key genes and pathways involved in staminate strobilus development and dehiscence in Torreya grandis. Through weighted gene co-expression network analysis (WGCNA), we identified fast elongation-related genes enriched in carbon metabolism and auxin signal transduction, whereas dehiscence-related genes were abundant in alpha-linolenic acid metabolism and the phenylpropanoid pathway. Based on WGCNA, we also identified PHYTOCHROME-INTERACTING FACTOR4 (TgPIF4) as a potential regulator for fast elongation of staminate strobilus and 2 WRKY proteins (TgWRKY3 and TgWRKY31) as potential regulators for staminate strobilus dehiscence. Multiple protein-DNA interaction analyses showed that TgPIF4 directly activates the expression of TRANSPORT INHIBITOR RESPONSE2 (TgTIR2) and NADP-MALIC ENZYME (TgNADP-ME). Overexpression of TgPIF4 significantly promoted staminate strobilus elongation by elevating auxin signal transduction and pyruvate content. TgWRKY3 and TgWRKY31 bind to the promoters of the lignin biosynthesis gene PHENYLALANINE AMMONIA-LYASE (TgPAL) and jasmonic acid metabolism gene JASMONATE O-METHYLTRANSFERASE (TgJMT), respectively, and directly activate their transcription. Overexpression of TgWRKY3 and TgWRKY31 in the staminate strobilus led to early dehiscence, accompanied by increased lignin and methyl jasmonate levels, respectively. Collectively, our findings offer a perspective for understanding the growth of staminate strobili in gymnosperms.

A LncRNA gene polymorphism (rs1814343) is associated with the risk of coronary artery lesions in southern Chinese Kawasaki disease patients.

BACKGROUND: Kawasaki disease (KD) is a multisystemic angiitis, and its most disastrous complication is coronary artery lesions (CALs). Recently, the role of long non-coding RNAs (lncRNAs) in KD has been reported. rs1814343 is a lncRNA, but the relationship between the lncRNA rs1814343 polymorphism and KD risk remains elusive. METHODS: We enrolled 1625 Kawasaki disease patients (583 patients with CAL and 1042 without CAL) and 1000 healthy controls from a southern Chinese population. We genotyped the rs1814343 C > T polymorphism in KD and control patients using the TaqMan method. The odds ratio (OR) and 95% confidence interval (CI) were used to estimate the strength of the association. RESULTS: There was no significant association between the lncRNA rs1814343 C > T polymorphism and KD susceptibility. However, we stratified patients in this study by CAL and sex. First, compared with the control groups, we found that the rs1814343 genotype increased risk for KD patients with CAL (TT vs. CC + CT: OR = 1.36, 95% CI = 1.08-1.71, p = 0.009). Moreover, when KD patients were stratified by CAL, the TT genotypes of this lncRNA polymorphism contributed to a relatively higher occurrence of KD with CAL than that was found in the CC/CT genotype patients (TT vs. CC + CT: OR = 1.35, 95% CI = 1.07-1.69, p = 0.011). In addition, our research suggested that the TT variant genotype in the lncRNA rs1814343 had an obvious risk of KD with CAL susceptibility in male children. CONCLUSION: The lncRNA rs1814343 C > T polymorphism was related to higher susceptibility of KD with CAL.

High-entropy-rate broadband chaos generation by using short-resonant-cavity DFB semiconductor laser with optical feedback.

Semiconductor lasers with delayed optical feedback are a promising source of optical chaos for practical applications, owing to simple configurations that are easy to integrate and synchronize. However, for traditional semiconductor lasers, the chaos bandwidth is limited by the relaxation frequency to several gigahertz. Here, we propose and experimentally demonstrate that a short-resonant-cavity distributed-feedback (SC-DFB) laser can generate broadband chaos only with simple feedback from an external mirror. The short distributed-feedback resonant cavity not only enhances laser relaxation frequency but also makes the laser mode more susceptible to external feedback. Experiments obtained a laser chaos with 33.6 GHz bandwidth and a spectral flatness of 4.5 dB. The corresponding entropy rate is estimated as more than 33.3 Gbit/s. It is believed that the SC-DFB lasers will promote development of chaos-based secure communication and physical key distribution.

Lysine demethylase LSD1 delivered via small extracellular vesicles promotes gastric cancer cell stemness.

Several studies have examined the functions of nucleic acids in small extracellular vesicles (sEVs). However, much less is known about the protein cargos of sEVs and their functions in recipient cells. This study demonstrates the presence of lysine-specific demethylase 1 (LSD1), which is the first identified histone demethylase, in the culture medium of gastric cancer cells. We show that sEVs derived from gastric cancer cells and the plasma of patients with gastric cancer harbor LSD1. The shuttling of LSD1-containing sEVs from donor cells to recipient gastric cancer cells promotes cancer cell stemness by positively regulating the expression of Nanog, OCT4, SOX2, and CD44. Additionally, sEV-delivered LSD1 suppresses oxaliplatin response of recipient cells in vitro and in vivo, whereas LSD1-depleted sEVs do not. Taken together, we demonstrate that LSD1-loaded sEVs can promote stemness and chemoresistance to oxaliplatin. These findings suggest that the LSD1 content of sEV could serve as a biomarker to predict oxaliplatin response in gastric cancer patients.

Enantioselective Synthesis of 2,3,3a,8a-Tetrahydrofuro[2,3-b]benzofuran Scaffolds Enabled by Cu(II)/SPDO-Catalyzed [3+2] Cycloaddition of 2,3-Dihydrofuran and Quinone Esters.

An asymmetric [3+2] cycloaddition of quinone esters with 2,3-dihydrofuran has been realized via a newly developed Cu(II)/SPDO complex. It provides straightforward access to 2,3,3a,8a-tetrahydrofuro[2,3-b]benzofurans (TFB) with high enantioselectivity (up to 97.5:2.5 er) and diastereoselectivity (all >20:1 dr). The resulting adducts contain two adjacent stereocenters and a continuously functionalized benzene ring. Additionally, this transformation could be easily performed on a gram scale, allowing for expedient synthesis of natural dihydroaflatoxin D2 and aflatoxin B2.

Multiplexed CRISPR/Cas9 editing of the long-chain acyl-CoA synthetase family in the diatom Phaeodactylum tricornutum reveals that mitochondrial ptACSL3 is involved in the synthesis of storage lipids.

Long-chain acyl-CoA synthetases (LACS) play diverse and fundamentally important roles in lipid metabolism. While their functions have been well established in bacteria, yeast and plants, the mechanisms by which LACS isozymes regulate lipid metabolism in unicellular oil-producing microalgae, including the diatom Phaeodactylum tricornutum, remain largely unknown. In P. tricornutum, a family of five genes (ptACSL1-ptACSL5) encodes LACS activities. We generated single lacs knockout/knockdown mutants using multiplexed CRISPR/Cas9 method, and determined their substrate specificities towards different fatty acids (FAs) and subcellular localisations. ptACSL3 is localised in the mitochondria and its disruption led to compromised growth and reduced triacylglycerol (TAG) content when cells were bubbled with air. The ptACSL3 mutants showed altered FA profiles in two galactoglycerolipids and phosphatidylcholine (PC) with significantly reduced distribution of 16:0 and 16:1. ptACSL5 is localised in the peroxisome and its knockdown resulted in reduced growth rate and altered molecular species of PC and TAG, indicating a role in controlling the composition of acyl-CoAs for lipid synthesis. Our work demonstrates the potential of generating gene knockout mutants with the mutation of large fragment deletion using multiplexed CRISPR/Cas9 and provides insight into the functions of LACS isozymes in lipid metabolism in the oleaginous microalgae.

Ultraviolet C Irradiation-Induced Dehybridization of Double-Stranded Oligonucleotides: Mechanism Investigation and Label-Free Measurement of the Photodamage Level.

Elucidating the mechanism and estimating the extent of conformation change of double-stranded DNA (dsDNA) upon ultraviolet (UV) exposure are of vital importance for understanding the DNA photodamage process. The existing research was mainly focused on the lesions of single-stranded DNA (ssDNA) and involved off-site measurement of the photodamage level. In this work, short-wavelength UV (UVC) (254 nm) irradiation was demonstrated to induce the dehybridization of dsDNA due to the loss of paring capacity of photodamaged pyrimidine nucleobases. The intrinsic programmability of dsDNA enabled researchers to rationally design the on-demand dehybridization sites. The spatial conformation switch of dsDNA caused by UVC irradiation could be evolved into a label-free sensing platform for the on-site measurement of the DNA photodamage level.

First-principles investigation of copper diffusion barrier performance in defective 2D layered materials.

This paper investigates the diffusion barrier performance of 2D layered materials with pre-existing vacancy defects using first-principles density functional theory. Vacancy defects in 2D materials may give rise to a large amount of Cu accumulation, and consequently, the defect becomes a diffusion path for Cu. Five 2D layered structures are investigated as diffusion barriers for Cu, i.e. graphene with C vacancy, hBN with B/N vacancy, and MoS2with Mo/2S vacancy. The calculated energy barriers using climbing image-nudged elastic band show that MoS2-V2Shas the highest diffusion energy barrier among other 2D layers, followed by hBN-VNand graphene. The obtained energy barrier of Cu on defected layer is found to be proportional to the length of the diffusion path. Moreover, the diffusion of Cu through vacancy defects is found to modulate the electronic structures and magnetic properties of the 2D layer. The charge density difference shows that there exists a considerable charge transfer between Cu and barrier layer as quantified by Bader charge. Given the current need for an ultra-thin diffusion barrier layer, the obtained results contribute to the field of application of 2D materials as Cu diffusion barrier in the presence of mono-vacancy defects.

Comparative proteomic analysis of tomato (Solanum lycopersicum L.) shoots reveals crosstalk between strigolactone and auxin.

As one of the main vegetable crops cultivated in the world, the tomato has advantages of high yield and economic benefits, and plays an important role in promoting farmers' income and social and economic growth. However, lateral branches during the growth process of tomato consume considerable nutrients and reduce the yield of tomato. Phytohormones such as strigolactone and auxin can inhibit the formation of lateral branches. However, the mechanism of their interaction is not particularly clear. To better understand the effects of exogenous strigolactone and auxin on tomato, proteome analyses of tomato shoots treated with exogenous GR24 and indole acetic acid were performed using an integrated approach involving tandem mass tag (TMT) labeling and liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). We identified 6685 proteins, of which 5822 contained quantitative information. Many differentially expressed proteins (DEPs) were found in different comparisons, including 415, 148, and 130 DEPs in GR24 vs mock, IAA vs mock, and GR24 + IAA vs mock comparisons, respectively. The Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that 'photosynthesis - antenna proteins' were significantly enriched in three treatments. Our data can help reveal the interaction between strigolactone and auxin in tomato seedlings.

Lycium barbarum Polysaccharides and Capsaicin Inhibit Oxidative Stress, Inflammatory Responses, and Pain Signaling in Rats with Dextran Sulfate Sodium-Induced Colitis.

Ulcerative colitis (UC) is an inflammatory disease with chronic relapsing symptoms. This study investigated the effects of Lycium barbarum polysaccharides (LBP) and capsaicin (CAP) in dextran sulfate sodium (DSS)-induced UC rats. Rats were divided into normal, DSS-induced UC, and UC treated with 100 mg LBP/kg bw, 12 mg CAP/kg bw, or 50 mg LBP/kg bw and 6 mg CAP/kg bw. Rats were fed LBP or CAP orally by gavage for 4 weeks, and UC model was established by feeding 5% DSS in drinking water for 6 days during week 3. Oral CAP and mixture significantly reduced disease activity index. Oral LBP significantly decreased serum malondialdehyde, interleukin (IL)-6, colonic tumor necrosis factor (TNF)-α levels, and protein expression of transient receptor potential cation channel V1 (TRPV1) and transient receptor potential ankyrin 1 (TRPA1), but increased serum catalase activity. Oral CAP significantly suppressed serum IL-6, colonic TRPV1 and TRPA1 protein expression, but elevated IL-10 levels, serum superoxide dismutase and catalase activities. The mixture of LBP and CAP significantly reduced serum IL-6, colonic TNF-α and TRPA1 protein. In conclusion, administration of LBP and/or CAP attenuate DSS-induced UC symptoms through inhibiting oxidative stress, proinflammatory cytokines, and protein expression of TRPV1 and TRPA1.

Functional Plasmon-Activated Water Increases Akkermansia muciniphila Abundance in Gut Microbiota to Ameliorate Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD) is associated with dysbiosis and intestinal barrier dysfunction, as indicated by epithelial hyperpermeability and high levels of mucosal-associated bacteria. Changes in gut microbiota may be correlated with IBD pathogenesis. Additionally, microbe-based treatments could mitigate clinical IBD symptoms. Plasmon-activated water (PAW) is known to have an anti-inflammatory potential. In this work, we studied the association between the anti-inflammatory ability of PAW and intestinal microbes, thereby improving IBD treatment. We examined the PAW-induced changes in the colonic immune activity and microbiota of mice by immunohistochemistry and next generation sequencing, determined whether drinking PAW can mitigate IBD induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS) and dysbiosis through mice animal models. The effects of specific probiotic species on mice with TNBS-induced IBD were also investigated. Experimental results indicated that PAW could change the local inflammation in the intestinal microenvironment. Moreover, the abundance of Akkermansia spp. was degraded in the TNBS-treated mice but elevated in the PAW-drinking mice. Daily rectal injection of Akkermansia muciniphila, a potential probiotic species in Akkermansia spp., also improved the health of the mice. Correspondingly, both PAW consumption and increasing the intestinal abundance of Akkermansia muciniphila can mitigate IBD in mice. These findings indicate that increasing the abundance of Akkermansia muciniphila in the gut through PAW consumption or other methods may mitigate IBD in mice with clinically significant IBD.

CDC42EP3 is a key promoter involved in the development and progression of gastric cancer.

Gastric cancer (GC) is one of the most prevalent cancers and severely endangers human health. Due to the low rate of diagnosis, most patients with GC are diagnosed as advanced. CDC42 effector protein 3 (CDC42EP3) has been revealed to be involved in several types of human cancers' development and progression. However, the function of CDC42EP3 in GC is not yet clear. CDC42EP3 expression was detected by immunohistochemistry, quantitative real-time PCR and Western blot assay in tumor tissues and cell lines of GC. CDC42EP3 knockdown cell models were constructed by lentivirus transfection. Cell proliferation was evaluated by the 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The wound-healing assay and the transwell assay were utilized to assess the cell migration. Also, the cell apoptosis and the cell cycle were evaluated by flow cytometry. Moreover, the mechanism was investigated by Human Apoptosis Antibody Array. The in vivo experiments were conducted to verify the effects of CDC42EP3 knockdown on the tumor growth of GC. The expression level of CDC42EP3 was up-regulated in tumor tissues. High CDC42EP3 expression was positively related to more advanced tumor grade. CDC42EP3 knockdown inhibited cell proliferation and migration, promoted cell apoptosis and suppressed the tumor growth. On the other hand, it was also found that the silencing of CDC42EP3 inhibited HSP27 and IGF-1sR expression as well as promoted Caspase3, p53, TNF-α, TNF-ß, TRAILR-1 and TRAILR-2 expression. CDC42EP3 was revealed to work as a tumor promoter in the development and progression of GC, which could be a promising therapeutic target for the therapy of GC.

CCNI2 promotes the progression of human gastric cancer through HDGF.

BACKGROUND: Gastric cancer is a highly aggressive malignant tumor with heterogeneity and is still a global health problem. The present study aimed to investigate the role of Cyclin I-like (CCNI2) in the regulation of phenotype and tumorigenesis, as well as its underlying mechanisms. METHOD: The expression profile of CCNI2 in gastric cancer was determined based on The Cancer Genome Atlas (TCGA) database and immunohistochemical staining. The effects of altered CCNI2 expression on the biological phenotypes such as proliferation, clone formation, apoptosis and migration of gastric cancer cell lines BGC-823 and SGC-7901 were investigated. Mice xenograft models were established to reveal the role of CCNI2 knockdown on tumorigenesis. The potential mechanism of CCNI2 regulating gastric cancer was preliminarily determined by RNA sequencing. RESULT: CCNI2 was abundantly expressed in gastric cancer and was positively correlated with pathological stage. Knockdown of CCNI2 slowed down the malignant progression of gastric cancer by inhibiting tumor cell proliferation, increasing the susceptibility to apoptosis and suppressing migration. Moreover, downregulation of CCNI2 attenuated the ability of gastric cancer cells to form tumors in mice. Additionally, there was an interaction between CCNI2 and transcription factor hepatoma-derived growth factor (HDGF) in SGC-7901 cells. Knockdown of CCNI2 alleviated the promoting effects of HDGF overexpression in gastric cancer cells. CONCLUSIONS: CCNI2 promoted the progression of human gastric cancer through HDGF, which drew further interest regarding its clinical application as a potential therapeutic target.

A genomic-clinicopathologic Nomogram for the preoperative prediction of lymph node metastasis in gastric cancer.

BACKGROUND: Preoperative evaluation of lymph node (LN) state is of pivotal significance for informing therapeutic decisions in gastric cancer (GC) patients. However, there are no non-invasive methods that can be used to preoperatively identify such status. We aimed at developing a genomic biosignature based model to predict the possibility of LN metastasis in GC patients. METHODS: We used the RNA profile retrieving strategy and performed RNA expression profiling in a large GC cohort (GSE62254, n = 300) from Gene Expression Ominus (GEO). In the exploratory stage, 300 GC patients from GSE62254 were involved and the differentially expressed RNAs (DERs) for LN-status were determined using the R software. GC samples in GSE62254 were randomly allocated into a learning set (n = 210) and a verification set (n = 90). By using the Least absolute shrinkage and selection operator (LASSO) regression approach, a set of 23-RNA signatures were established and the signature based nomogram was subsequently built for distinguishing LN condition. The diagnostic efficiency, as well as the clinical performance of this model were assessed using the decision curve analysis (DCA). Metascape was used for bioinformatic analysis of the DERs. RESULTS: Based on the genomic signature, we established a nomogram that robustly distinguished LN status in the learning (AUC = 0.916, 95% CI 0.833-0.999) and verification sets (AUC = 0.775, 95% CI 0.647-0.903). DCA demonstrated the clinical value of this nomogram. Functional enrichment analysis of the DERs was performed using bioinformatics methods which revealed that these DERs were involved in several lymphangiogenesis-correlated cascades. CONCLUSIONS: In this study, we present a genomic signature based nomogram that integrates the 23-RNA biosignature based scores and Lauren classification. This model can be utilized to estimate the probability of LN metastasis with good performance in GC. The functional analysis of the DERs reveals the prospective biogenesis of LN metastasis in GC.

Gut transit time, using radiological contrast imaging, to predict early signs of necrotizing enterocolitis.

BACKGROUND: Immature gut motility in preterm neonates may be a risk factor for necrotizing enterocolitis (NEC). Using preterm pigs as a model for infants, we hypothesized that intestinal dysmotility precedes NEC development. METHODS: Eighty-five preterm pigs were fed increasing amounts of milk diets to induce NEC lesions, as detected at autopsy on day 5. Gut transit time was determined on day 4 by x-ray imaging after oral intake of contrast solution. RESULTS: No clinical or radiological signs of NEC were detected on day 4, but macroscopic NEC lesions were recorded in 59% of pigs (n = 50) on day 5. Relative to pigs without NEC (noNEC, n = 35), pigs with small intestinal lesions (siNEC, n = 18) showed delayed stomach emptying time (StEmpty) and time for contrast to reach cecum (ToCecum) already on day 4. Pigs with lesions only in colon (coNEC, n = 20) showed more diarrhea, shorter ToCecum time, but longer small intestinal emptying time (SiEmpty). ToCecum time predicted siNEC and coNEC lesions with a receiver-operator characteristic area under the curve of 78-81%. CONCLUSIONS: Region-dependent changes in gut transit time is associated with early NEC development in preterm pigs. How gut dysmotility is related to NEC in preterm infants requires further investigations. IMPACT: Using preterm pigs as a model for preterm infants, we show that gut transit time, using serial x-ray contrast imaging, was changed in individuals with NEC-like lesions before they showed the typical radiological signs of NEC. Thus prolonged transit time across the entire gut was recorded when NEC lesions appeared in the small intestine but not when lesions were detected only in the colon. Until now, recordings of food transit have mainly investigated changes in the upper gut. Using serial x-rays, this study describes food transit across the entire gut and documents a region-dependent effect of NEC lesions on gut transit changes in preterm individuals. The findings provide proof of concept for use of x-ray contrast imaging as a tool to monitor gut transit in preterm pigs as models for infants. Delayed passage across the entire gut may be an early sign of small intestinal NEC, at least in pigs. More studies are needed to confirm relations in infants. In the future, it might be possible to use x-ray contrast imaging in preterm infants to better understand gut motility in relation to early NEC progression and need for medical NEC treatment.

Enantioselective Addition-Alkylation of α,ß-Unsaturated Carbonyls via Bisguanidinium Silicate Ion Pair Catalysis.

Silicon hydrides, alkynylsilanes, and alkoxylsilanes were activated by fluoride in the presence of bisguanidinium catalyst to form hypervalent silicate ion pairs. These activated silicates undergo 1,4-additions with chromones, coumarins, and α-cyanocinnamic esters generating enolsilicate intermediates, for a consequent stereoselective alkylation reaction. The reduction-alkylation reaction proceeded under mild conditions using polymethylhydrosiloxane, a cheap and environmentally friendly hydride source. The addition-alkylation reactions with alkynylsilanes and alkoxylsilanes resulted in the construction of two vicinal chiral carbon centers with excellent enantioselectivities and diastereoselectivities (up to 99% ee, >99:1 dr). Density functional theory calculations and experimental NMR studies revealed that penta-coordinated silicates are crucial intermediates.

A novel single-base mutation in CaBRI1 confers dwarf phenotype and brassinosteroid accumulation in pepper.

Dwarfing is the development trend of pepper breeding. It is of great practical and scientific value to generate new dwarf germplasms, and identify new genes or alleles conferring dwarf traits in pepper. In our previous study, a weakly BR-insensitive dwarf mutant, E29, was obtained by EMS mutagenesis of the pepper inbred line 6421. It can be used as a good parent material for breeding new dwarf varieties. Here, we found that this dwarf phenotype was controlled by a single recessive gene. Whole-genome resequencing, dCAPs analysis, and VIGs validation revealed that this mutation was caused by a nonsynonymous single-nucleotide mutation (C to T) in CaBRI1. An enzyme activity assay, transcriptome sequencing, and BL content determination further revealed that an amino-acid change (Pro1157Ser) in the serine/threonine protein kinase and catalytic (S_TKc) domain of CaBRI1 impaired its kinase activity and caused the transcript levels of two important genes (CaDWF4 and CaROT3) participating in BR biosynthesis to increase dramatically in the E29 mutant, accompanied by significantly increased accumulation of brassinolide (BL). Therefore, we concluded that the novel single-base mutation in CaBRI1 conferred the dwarf phenotype and resulted in brassinosteroid (BR) accumulation in pepper. This study provides a new allelic variant of the height-regulating gene CaBRI1 that has theoretical and practical values for the breeding of the plants suitable for the facility cultivation and mechanized harvesting of pepper varieties.

The Phospholipid:Diacylglycerol Acyltransferase-Mediated Acyl-Coenzyme A-Independent Pathway Efficiently Diverts Fatty Acid Flux from Phospholipid into Triacylglycerol in Escherichia coli.

Researchers have long endeavored to accumulate triacylglycerols (TAGs) or their derivatives in easily managed microbes. The attempted production of TAGs in Escherichia coli has revealed barriers to the broad applications of this technology, including low TAG productivity and slow cell growth. We have demonstrated that an acyl-CoA-independent pathway can divert phospholipid flux into TAG formation in E. coli mediated by Chlamydomonas reinhardtii phospholipid:diacylglycerol acyltransferase (CrPDAT) without interfering with membrane functions. We then showed the synergistic effect on TAG accumulation via the acyl-CoA-independent pathway mediated by PDAT and the acyl-CoA-dependent pathway mediated by wax ester synthase/acyl-CoA:diacylglycerol acyltransferase (WS/DGAT). Furthermore, CrPDAT led to synchronous TAG accumulation during cell growth, and this could be enhanced by supplementation of arbutin. We also showed that rationally mutated CrPDAT was capable of decreasing TAG lipase activity without impairing PDAT activity. Finally, ScPDAT from Saccharomyces cerevisiae exhibited similar activities as CrPDAT in E. coli Our results suggest that the improvement in accumulation of TAGs and their derivatives can be achieved by fine-tuning of phospholipid metabolism in E. coli Understanding the roles of PDAT in the conversion of phospholipids into TAGs during the logarithmic growth phase may enable a novel strategy for the production of microbial oils.IMPORTANCE Although phospholipid:diacylglycerol acyltransferase (PDAT) activity is presumed to exist in prokaryotic oleaginous bacteria, the corresponding gene has not been identified yet. In this article, we have demonstrated that an acyl-CoA-independent pathway can divert phospholipid flux into TAG formation in Escherichia coli mediated by exogenous CrPDAT from Chlamydomonas reinhardtii without interfering with membrane functions. In addition, the acyl-CoA-independent pathway and the acyl-CoA-dependent pathway had the synergistic effect on TAG accumulation. Overexpression of CrPDAT led to synchronous TAG accumulation during cell growth. In particular, CrPDAT possessed multiple catalytic activities, and the rational mutation of CrPDAT led to the decrease of TAG lipase activity without impairing acyltransferase activity. The present findings suggested that applying PDAT in E. coli or other prokaryotic microbes may be a promising strategy for accumulation of TAGs and their derivatives.

STAT3 Mediated miR-30a-5p Inhibition Enhances Proliferation and Inhibits Apoptosis in Colorectal Cancer Cells.

Signal transducer and activator of transcription 3 (STAT3), a transcriptional factor involved in tumorigenesis and cancer stemness formation, contributes to drug resistance in cancer therapies. STAT3 not only mediates gene transcription but also participates in microRNA suppression. This study identified a STAT3-downstream micro RNA (miRNA) involved in drug resistance against regorafenib in colorectal cancer stem-like tumorspheres. Small RNAseq was used to investigate differential microRNAs in colorectal cancer cell-derived tumorspheres and in a STAT3-knockdown strain. The miRNA-mediated genes were identified by comparing RNAseq data with gene targets predicted using TargetScan. Assays for detecting cell viability and apoptosis were used to validate findings. The formation of colorectal cancer stem-like tumorspheres was inhibited by BBI608, a STAT3 inhibitor, but not by regorafenib. Additional investigations for microRNA expression demonstrated an increase in 10 miRNAs and a decrease in 13 miRNAs in HT29-derived tumorspheres. A comparison of small RNAseq results between tumorspheres and HT29shSTAT3 cells revealed the presence of four STAT3-mediated miRNAs in HT29-derived tumorspheres: hsa-miR-215-5p, hsa-miR-4521, and hsa-miR-215-3p were upregulated, whereas miR-30a-5p was downregulated. Furthermore, hsa-miR-4521 was associated with poor overall survival probability, and miR-30a-5p was associated with better overall survival probability in patients with rectum cancer. Comparisons of RNAseq findings between HCT116- and HT29-derived tumorspheres revealed that HSPA5 were mediated by the STAT3-miR-30a-5p axis, which is overexpressed in colorectal tumorspheres associating to anti-apoptosis. In addition, the transfection of miR-30a-5p and inhibition of HSPA5 by HA15 significantly reduced cell viability and increased apoptosis in HT29 cells. In conclusion, a STAT3-miR-30a-5p-HSPA5 axis was observed against regorafenib-mediated apoptosis in colorectal cancer tumorspheres. The expression of miR-30a-5p was repressed by STAT3; in addition, HSPA5 was identified as the target gene of miR-30a-5p and contributed to both tumorsphere formation and anti-apoptosis.

Comprehensive Phosphoproteomic Analysis of Pepper Fruit Development Provides Insight into Plant Signaling Transduction.

Limited knowledge is available for phosphorylation modifications in pepper (Capsicum annuum L.), especially in pepper fruit development. In this study, we conducted the first comprehensive phosphoproteomic analysis of pepper fruit at four development stage by Tandem Mass Tag proteomic approaches. A total of 2639 unique phosphopeptides spanning 1566 proteins with 4150 nonredundant sites of phosphorylation were identified, among which 2327 peptides in 1413 proteins were accurately quantified at four different stages. Mature Green (MG) to breaker stage showed the largest number of differentially expressed phosphoproteins and the number of downregulated phosphoproteins was significantly higher than that of upregulated after MG stage. Twenty seven phosphorylation motifs, including 22 pSer motifs and five pThr motifs and 85 kinase including 28 serine/threonine kinases, 14 receptor protein kinases, six mitogen-activated protein kinases, seven calcium-dependent protein kinases, two casein kinases, and some other kinases were quantified. Then the dynamic changes of phosphorylated proteins in ethylene and abscisic acid signaling transduction pathways during fruit development were analyzed. Our results provide a cascade of phosphoproteins and a regulatory network of phosphorylation signals, which help to further understand the mechanism of phosphorylation in pepper fruit development.