Lp-PLA2 (Lipoprotein-Associated Phospholipase A2) Deficiency Lowers Cholesterol Levels and Protects Against Atherosclerosis in Rabbits.

BACKGROUND: Elevated plasma Lp-PLA2 (lipoprotein-associated phospholipase A2) activity is closely associated with an increased risk of cardiovascular events. However, whether and how Lp-PLA2 is directly involved in the pathogenesis of atherosclerosis is still unclear. To examine the hypothesis that Lp-PLA2 could be a potential preventative target of atherosclerosis, we generated Lp-PLA2 knockout rabbits and investigated the pathophysiological functions of Lp-PLA2. METHODS: Lp-PLA2 knockout rabbits were generated using CRISPR/Cas9 system to assess the role of Lp-PLA2 in plasma lipids regulation and identify its underlying molecular mechanisms. Homozygous knockout rabbits along with wild-type rabbits were fed a cholesterol-rich diet for up to 14 weeks and their atherosclerotic lesions were compared. Moreover, the effects of Lp-PLA2 deficiency on the key cellular behaviors in atherosclerosis were assessed in vitro. RESULTS: When rabbits were fed a standard diet, Lp-PLA2 deficiency led to a significant reduction in plasma lipids. The decreased protein levels of SREBP2 (sterol regulatory element-binding protein 2) and HMGCR (3-hydroxy-3-methylglutaryl coenzyme A reductase) in livers of homozygous knockout rabbits indicated that the cholesterol biosynthetic pathway was impaired with Lp-PLA2 deficiency. In vitro experiments further demonstrated that intracellular Lp-PLA2 efficiently enhanced SREBP2-related cholesterol biosynthesis signaling independently of INSIGs (insulin-induced genes). When fed a cholesterol-rich diet, homozygous knockout rabbits exhibited consistently lower level of hypercholesterolemia, and their aortic atherosclerosis lesions were significantly reduced by 60.2% compared with those of wild-type rabbits. The lesions of homozygous knockout rabbits were characterized by reduced macrophages and the expression of inflammatory cytokines. Macrophages of homozygous knockout rabbits were insensitive to M1 polarization and showed reduced DiI-labeled lipoprotein uptake capacity compared with wild-type macrophages. Lp-PLA2 deficiency also inhibited the adhesion between monocytes and endothelial cells. CONCLUSIONS: These results demonstrate that Lp-PLA2 plays a causal role in regulating blood lipid homeostasis and Lp-PLA2 deficiency protects against dietary cholesterol-induced atherosclerosis in rabbits. Lp-PLA2 could be a potential target for the prevention of atherosclerosis.

Specification of female germline by microRNA orchestrated auxin signaling in Arabidopsis.

Germline determination is essential for species survival and evolution in multicellular organisms. In most flowering plants, formation of the female germline is initiated with specification of one megaspore mother cell (MMC) in each ovule; however, the molecular mechanism underlying this key event remains unclear. Here we report that spatially restricted auxin signaling promotes MMC fate in Arabidopsis. Our results show that the microRNA160 (miR160) targeted gene ARF17 (AUXIN RESPONSE FACTOR17) is required for promoting MMC specification by genetically interacting with the SPL/NZZ (SPOROCYTELESS/NOZZLE) gene. Alterations of auxin signaling cause formation of supernumerary MMCs in an ARF17- and SPL/NZZ-dependent manner. Furthermore, miR160 and ARF17 are indispensable for attaining a normal auxin maximum at the ovule apex via modulating the expression domain of PIN1 (PIN-FORMED1) auxin transporter. Our findings elucidate the mechanism by which auxin signaling promotes the acquisition of female germline cell fate in plants.

Microbial spore genetic marker technology, a potential technology for traditional Chinese medicine traceability system.

Traditional Chinese medicine (TCM) has a long history, rich clinical experience, and unique advantages in the prevention and treatment of diseases. The quality and safety of Chinese medicinal materials (CMMs) directly affect the clinical efficacy and development of the TCM industry. However, confused provenance, counterfeiting and adulteration of CMMs hinder the acceptance of its therapeutic benefits in modern society. Therefore, the establishment and improvement of a TCM traceability system would be conducive to the transparency of the CMMs production, distribution, and circulation, thereby improving drug safety and promoting industry development. This review discusses the challenges faced in the development of TCM traceability system, the technologies currently available for tracing CMMs, and the potential application of Barcoded Microbial Spores (BMS) to improve CMMs origin traceability and TCM traceability systems.

Epigenetic Regulation of Heat Stress in Plant Male Reproduction.

In flowering plants, male reproductive development is highly susceptible to heat stress. In this mini-review, we summarized different anomalies in tapetum, microspores, and pollen grains during anther development under heat stress. We then discussed how epigenetic control, particularly DNA methylation, is employed to cope with heat stress in male reproduction. Further understanding of epigenetic mechanisms by which plants manage heat stress during male reproduction will provide new genetic engineering and molecular breeding tools for generating heat-resistant crops.

BRF Negatively Regulates Thermotolerance Defect of fes1a in Arabidopsis.

FES1A is a heat shock protein 70 binding protein. Mutation of FES1A leads to a defect in thermotolerance of Arabidopsis; however, independent fes1a mutants exhibit a range in the extent of thermosensitivity. Here, we found that BRF2, a gene adjacent to FES1A and encoding a component of transcription factor IIIB, affects the thermosensitivity of fes1a mutants. Knockout of BRF2 suppressed fes1a thermosensitivity, while overexpression of BRF2 increased thermosensitivity of fes1a. BRF2 in fes1a mutants regulates the transcriptional strength of RNA Polymerase II and accumulation of heat shock proteins and eventually affects the thermotolerance of fes1a. There is a cross-talking between RNA Pol III and Pol II. The cross-talking is initiated by BRF, magnified by the mutation of FES1A, and finally has an effect on thermotolerance.

Arabidopsis ZINC FINGER PROTEIN1 Acts Downstream of GL2 to Repress Root Hair Initiation and Elongation by Directly Suppressing bHLH Genes.

Cys2His2-like fold group (C2H2)-type zinc finger proteins promote root hair growth and development by regulating their target genes. However, little is known about their potential negative roles in root hair initiation and elongation. Here, we show that the C2H2-type zinc finger protein named ZINC FINGER PROTEIN1 (AtZP1), which contains an ERF-associated amphiphilic repression (EAR) motif, negatively regulates Arabidopsis (Arabidopsis thaliana) root hair initiation and elongation. Our results demonstrate that AtZP1 is highly expressed in root hairs and that AtZP1 inhibits transcriptional activity during root hair development. Plants overexpressing AtZP1 lacked root hairs, while loss-of-function mutants had longer and more numerous root hairs than the wild type. Transcriptome analysis indicated that AtZP1 downregulates genes encoding basic helix-loop-helix (bHLH) transcription factors associated with root hair cell differentiation and elongation. Mutation or deletion of the EAR motif substantially reduced the inhibitory activity of AtZP1. Chromatin immunoprecipitation assays, AtZP1:glucocorticoid receptor (GR) induction experiments, electrophoretic mobility shift assays, and yeast one-hybrid assays showed that AtZP1 directly targets the promoters of bHLH transcription factor genes, including the key root hair initiation gene ROOT HAIR DEFECTIVE6 (RHD6) and root hair elongation genes ROOT HAIR DEFECTIVE 6-LIKE 2 (RSL2) and RSL4, and suppresses root hair development. Our findings suggest that AtZP1 functions downstream of GL2 and negatively regulates root hair initiation and elongation, by suppressing RHD6, RSL4, and RSL2 transcription via the GL2/ZP1/RSL pathway.

Abnormality of hepatic triglyceride metabolism in ApcMin/+ mice with colon cancer cachexia.

AIMS: Colorectal cancer syndrome has been one of the greatest concerns in the world. Although several epidemiological studies have shown that hepatic low lipoprotein lipase (LPL) mRNA expression may be associated with dyslipidemia and tumor progression, it is still not known whether the liver plays an essential role in hyperlipidemia of ApcMin/+ mice. MAIN METHODS: We measured the expression of metabolic enzymes that involved fatty acid uptake, de novo lipogenesis (DNL), ß-oxidation and investigated hepatic triglyceride production in the liver of wild-type and ApcMin/+ mice. KEY FINDINGS: We found that hepatic fatty acid uptake and DNL decreased, but there was no significant difference in fatty acid ß-oxidation. Interestingly, the production of hepatic very low-density lipoprotein-triglyceride (VLDL-TG) decreased at 20 weeks of age, but marked steatosis was observed in the livers of the ApcMin/+ mouse. To further explore hypertriglyceridemia, we assessed the function of hepatic glycosylphosphatidylinositol-anchored high-density lipoprotein binding protein 1 (GPIHBP1) for the first time. GPIHBP1 is governed by the transcription factor octamer-binding transcription factor-1 (Oct-1) which are involved in the nuclear factor-κB (NF-κB) signaling pathway in the liver of ApcMin/+ mice. Importantly, it was also confirmed that sn50 (100 µg/mL, an inhibitor of the NF-κB) reversed the tumor necrosis factor α (TNFα)-induced Oct-1 and GPIHBP1 reduction in HepG2 cells. SIGNIFICANCE: Altogether, these findings highlighted a novel role of GPIHBP1 that might be responsible for hypertriglyceridemia in ApcMin/+ mice. Hypertriglyceridemia in these mice may be associated with their hepatic lipid metabolism development.

A comparative analysis of infection in patients with malignant cancer: A clinical pharmacist consultation study.

BACKGROUND: Infection analysisamongst malignant cancer patients remains elusive. The objective of this study is to investigate the characteristics of both infection and anti-infection treatments in patients group with malignant cancer. METHODS: We retrospectively studied the clinical data of 148 patients with malignant cancer and 171 benign patients enrolled in the pharmacist consultation from April 2015 to April 2017. Statistical analysis was performed by chi-square test to compare the classification of primary disease, sites of infection, composition of pathogenic bacteria, and the effectiveness of drug treatment. P value <0.05 was considered statistically significant. RESULTS: A total of 102 pathogen strains were detected in the patients with malignant cancer and 129 pathogen strains were noted in the benign patient group, respectively. Statistics indicated that more abdominal infections were observed in malignant cancer patients rather than in non-cancer patients. Additionally, more pseudomonas aeruginosa infection was found in the malignant cancer patient group while more Klebsiella pneumonia infection was noted in the benign group. These findings were supported by statistical evidence. There were fewer extended-spectrum ß-lactamases (ESBL) that produced Escherichia coli, which was commonly found in a gastrointestinal cancer patient group compared to patients under other types of cancer; it accounted for 51.3% of all malignant cases involved in the current study. CONCLUSIONS: Patients with malignant cancer are more likely to suffer from an infection containing pathogenic bacteria in comparison to benign patients. There have been considerable differences in the composition of pathogenic bacteria and its resistance to drugs. Overall, evaluating pathogens plays an essential role in the anti-infection treatment of patients with malignant cancer.

Cyclodextrin Ameliorates the Progression of Atherosclerosis via Increasing High-Density Lipoprotein Cholesterol Plasma Levels and Anti-inflammatory Effects in Rabbits.

To investigate the therapeutic effects of cyclodextrin on the development of atherosclerosis in rabbits, we evaluated the effects of (2-hydroxypropyl)-ß-cyclodextrin (HPßCD) therapy on the organ coefficient, lipid profiles, inflammatory cytokines, and atherosclerotic plaques in rabbits fed a high-fat diet. Our results demonstrated that HPßCD therapy reduced plasma triglyceride levels and inflammatory cytokine levels but increased plasma high-density lipoprotein cholesterol levels. HPßCD therapy produced a significant decrease in the atherosclerotic lesion area and reduced macrophage and collagen content in the lesions. The expression levels of inflammatory genes in aortic plaques were significantly reduced by HPßCD treatment, but the expression of ATP-binding cassette (ABC) transporters A1 (ABCA1) and G1 (ABCG1) in aortic plaques and livers increased significantly. HPßCD therapy may produce additional antiatherosclerotic benefits likely via increasing high-density lipoprotein cholesterol levels.

Abnormality of intestinal cholesterol absorption in ApcMin/+ mice with colon cancer cachexia.

Colorectal cancer syndrome has been one of the greatest concerns in the world, particularly in developed countries. Several epidemiological studies have shown that dyslipidemia may be associated with the progression of intestinal cachexia, but there is little research on the function of the small intestine, which is involved in blood lipid metabolism, in dyslipidemia. In the present study, we aimed to explore the function of intestinal cholesterol absorption in the ApcMin/+ mouse model using an intestinal lipid absorption test. We found that both triglyceride (TG) and total cholesterol (TC) uptake were inhibited in the intestine of ApcMin/+ mice with age and the intestinal peroxisome proliferator-activated receptor α (PPARα) downregulated the processes of ß-oxidation, oxidative stress response, and cholesterol absorption in APC-deficient mice. In addition, reduced expression levels of farnesoid X receptor (FXR) and apical sodium-dependent bile acid transporter (ASBT) indicated that bile acid metabolism might be associated with intestinal cholesterol absorption in ApcMin/+ mice. Thus, our data suggested that the intestine plays an essential role in cholesterol uptake and that bile acid metabolism seems to cause a decrease in intestinal cholesterol uptake in ApcMin/+ mice.

The Arabidopsis catalase triple mutant reveals important roles of catalases and peroxisome-derived signaling in plant development.

Hydrogen peroxide (H2 O2 ) is generated in many metabolic processes. As a signaling molecule, H2 O2 plays important roles in plant growth and development, as well as environmental stress response. In Arabidopsis, there are three catalase genes, CAT1, CAT2, and CAT3. The encoded catalases are predominately peroxisomal proteins and are critical for scavenging H2 O2 . Since CAT1 and CAT3 are linked on chromosome 1, it has been almost impossible to generate cat1/3 and cat1/2/3 mutants by traditional genetic tools. In this study, we constructed cat1/3 double mutants and cat1/2/3 triple mutants by CRISPR/Cas9 to investigate the role of catalases. The cat1/2/3 triple mutants displayed severe redox disturbance and growth defects under physiological conditions compared with wild-type and the cat2/3 double mutants. Transcriptome analysis showed a more profound transcriptional response in the cat1/2/3 triple mutants compared to the cat2/3 mutants. These differentially expressed genes are involved in plant growth regulation as well as abiotic and biotic stress responses. In addition, expression of OXI1 (OXIDATIVE SIGNAL INDUCIBLE 1) and several MAPK cascade genes were changed dramatically in the catalase triple mutant, suggesting that H2 O2 produced in peroxisomes could serve as a peroxisomal retrograde signal.

Carbonic Anhydrases Function in Anther Cell Differentiation Downstream of the Receptor-Like Kinase EMS1.

Plants extensively employ leucine-rich repeat receptor-like kinases (LRR-RLKs), the largest family of RLKs, to control a wide range of growth and developmental processes as well as defense responses. To date, only a few direct downstream effectors for LRR-RLKs have been identified. We previously showed that the LRR-RLK EMS1 (EXCESS MICROSPOROCYTES1) and its ligand TPD1 (TAPETUM DETERMINANT1) are required for the differentiation of somatic tapetal cells and reproductive microsporocytes during early anther development in Arabidopsis thaliana Here, we report the identification of ß-carbonic anhydrases (ßCAs) as the direct downstream targets of EMS1. EMS1 biochemically interacts with ßCA proteins. Loss of function of ßCA genes caused defective tapetal cell differentiation, while overexpression of ßCA1 led to the formation of extra tapetal cells. EMS1 phosphorylates ßCA1 at four sites, resulting in increased ßCA1 activity. Furthermore, phosphorylation-blocking mutations impaired the function of ßCA1 in tapetal cell differentiation; however, a phosphorylation mimic mutation promoted the formation of tapetal cells. ßCAs are also involved in pH regulation in tapetal cells. Our findings highlight the role of ßCA in controlling cell differentiation and provide insights into the posttranslational modification of carbonic anhydrases via receptor-like kinase-mediated phosphorylation.

Morphological Characterization of a New and Easily Recognizable Nuclear Male Sterile Mutant of Sorghum (Sorghum bicolor).

Sorghum (Sorghum bicolor L. Moench) is one of the most important grain crops in the world. The nuclear male sterility (NMS) trait, which is caused by mutations on the nuclear gene, is valuable for hybrid breeding and genetic studies. Several NMS mutants have been reported previously, but none of them were well characterized. Here, we present our detailed morphological characterization of a new and easily recognizable NMS sorghum mutant male sterile 8 (ms8) isolated from an elite inbred BTx623 mutagenized by ethyl methane sulfonate (EMS). Our results show that the ms8 mutant phenotype was caused by a mutation on a single recessive nuclear gene that is different from all available NMS loci reported in sorghum. In fertile sorghum plants, yellow anthers appeared first during anthesis, while in the ms8 mutant, white hairy stigma emerged first and only small white anthers were observed, making ms8 plants easily recognizable when flowering. The ovary development and seed production after manual pollination are normal in the ms8 mutant, indicating it is female fertile and male sterile only. We found that ms8 anthers did not produce pollen grains. Further analysis revealed that ms8 anthers were defective in tapetum development, which led to the arrest of pollen formation. As a stable male sterile mutant across different environments, greenhouses, and fields in different locations, the ms8 mutant could be a useful breeding tool. Moreover, ms8 might be an important for elucidating male gametophyte development in sorghum and other plants.

Two SERK Receptor-Like Kinases Interact with EMS1 to Control Anther Cell Fate Determination.

Cell signaling pathways mediated by leucine-rich repeat receptor-like kinases (LRR-RLKs) are essential for plant growth, development, and defense. The EMS1 (EXCESS MICROSPOROCYTES1) LRR-RLK and its small protein ligand TPD1 (TAPETUM DETERMINANT1) play a fundamental role in somatic and reproductive cell differentiation during early anther development in Arabidopsis (Arabidopsis thaliana). However, it is unclear whether other cell surface molecules serve as coregulators of EMS1. Here, we show that SERK1 (SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE1) and SERK2 LRR-RLKs act redundantly as coregulatory and physical partners of EMS1. The SERK1/2 genes function in the same genetic pathway as EMS1 in anther development. Bimolecular fluorescence complementation, Förster resonance energy transfer, and coimmunoprecipitation approaches revealed that SERK1 interacted biochemically with EMS1. Transphosphorylation of EMS1 by SERK1 enhances EMS1 kinase activity. Among 12 in vitro autophosphorylation and transphosphorylation sites identified by tandem mass spectrometry, seven of them were found to be critical for EMS1 autophosphorylation activity. Furthermore, complementation test results suggest that phosphorylation of EMS1 is required for its function in anther development. Collectively, these data provide genetic and biochemical evidence of the interaction and phosphorylation between SERK1/2 and EMS1 in anther development.

Rhizobium sp. IRBG74 Alters Arabidopsis Root Development by Affecting Auxin Signaling.

Rhizobium sp. IRBG74 not only nodulates Sesbania cannabina but also can enhance rice growth; however, the underlying molecular mechanisms are not clear. Here, we show that Rhizobium sp. IRBG74 colonizes the roots of Arabidopsis thaliana, which leads to inhibition in the growth of main root but enhancement in the formation of lateral roots. The promotion of lateral root formation by Rhizobium sp. IRBG74 in the fls2-1 mutant, which is insensitive to flagellin, is similar to the wild-type plant, while the auxin response deficient mutant tir1-1 is significantly less sensitive to Rhizobium sp. IRBG74 than the wild type in terms of the inhibition of main root elongation and the promotion of lateral root formation. Further transcriptome analysis of Arabidopsis roots inoculated with Rhizobium sp. IRBG74 revealed differential expression of 50 and 211 genes at 24 and 48 h, respectively, and a majority of these genes are involved in auxin signaling. Consistent with the transcriptome analysis results, Rhizobium sp. IRBG74 treatment induces expression of the auxin responsive reporter DR5:GUS in roots. Our results suggest that in Arabidopsis Rhizobium sp. IRBG74 colonizes roots and promotes the lateral root formation likely through modulating auxin signaling. Our work provides insight into the molecular mechanisms of interactions between legume-nodulating rhizobia and non-legume plants.

Sterility Caused by Floral Organ Degeneration and Abiotic Stresses in Arabidopsis and Cereal Grains.

Natural floral organ degeneration or abortion results in unisexual or fully sterile flowers, while abiotic stresses lead to sterility after initiation of floral reproductive organs. Since normal flower development is essential for plant sexual reproduction and crop yield, it is imperative to have a better understanding of plant sterility under regular and stress conditions. Here, we review the functions of ABC genes together with their downstream genes in floral organ degeneration and the formation of unisexual flowers in Arabidopsis and several agriculturally significant cereal grains. We further explore the roles of hormones, including auxin, brassinosteroids, jasmonic acid, gibberellic acid, and ethylene, in floral organ formation and fertility. We show that alterations in genes affecting hormone biosynthesis, hormone transport and perception cause loss of stamens/carpels, abnormal floral organ development, poor pollen production, which consequently result in unisexual flowers and male/female sterility. Moreover, abiotic stresses, such as heat, cold, and drought, commonly affect floral organ development and fertility. Sterility is induced by abiotic stresses mostly in male floral organ development, particularly during meiosis, tapetum development, anthesis, dehiscence, and fertilization. A variety of genes including those involved in heat shock, hormone signaling, cold tolerance, metabolisms of starch and sucrose, meiosis, and tapetum development are essential for plants to maintain normal fertility under abiotic stress conditions. Further elucidation of cellular, biochemical, and molecular mechanisms about regulation of fertility will improve yield and quality for many agriculturally valuable crops.

Control of Anther Cell Differentiation by the Small Protein Ligand TPD1 and Its Receptor EMS1 in Arabidopsis.

A fundamental feature of sexual reproduction in plants and animals is the specification of reproductive cells that conduct meiosis to form gametes, and the associated somatic cells that provide nutrition and developmental cues to ensure successful gamete production. The anther, which is the male reproductive organ in seed plants, produces reproductive microsporocytes (pollen mother cells) and surrounding somatic cells. The microsporocytes yield pollen via meiosis, and the somatic cells, particularly the tapetum, are required for the normal development of pollen. It is not known how the reproductive cells affect the differentiation of these somatic cells, and vice versa. Here, we use molecular genetics, cell biological, and biochemical approaches to demonstrate that TPD1 (TAPETUM DETERMINANT1) is a small secreted cysteine-rich protein ligand that interacts with the LRR (Leucine-Rich Repeat) domain of the EMS1 (EXCESS MICROSPOROCYTES1) receptor kinase at two sites. Analyses of the expressions and localizations of TPD1 and EMS1, ectopic expression of TPD1, experimental missorting of TPD1, and ablation of microsporocytes yielded results suggesting that the precursors of microsporocyte/microsporocyte-derived TPD1 and pre-tapetal-cell-localized EMS1 initially promote the periclinal division of secondary parietal cells and then determine one of the two daughter cells as a functional tapetal cell. Our results also indicate that tapetal cells suppress microsporocyte proliferation. Collectively, our findings show that tapetal cell differentiation requires reproductive-cell-secreted TPD1, illuminating a novel mechanism whereby signals from reproductive cells determine somatic cell fate in plant sexual reproduction.

Deregulation of the OsmiR160 Target Gene OsARF18 Causes Growth and Developmental Defects with an Alteration of Auxin Signaling in Rice.

MicroRNAs (miRNAs) control gene expression as key negative regulators at the post-transcriptional level. MiR160 plays a pivotal role in Arabidopsis growth and development through repressing expression of its target AUXIN RESPONSE FACTOR (ARF) genes; however, the function of miR160 in monocots remains elusive. In this study, we found that the mature rice miR160 (OsmiR160) was mainly derived from OsMIR160a and OsMIR160b genes. Among four potential OsmiR160 target OsARF genes, the OsARF18 transcript was cleaved at the OsmiR160 target site. Rice transgenic plants (named mOsARF18) expressing an OsmiR160-resistant version of OsARF18 exhibited pleiotropic defects in growth and development, including dwarf stature, rolled leaves, and small seeds. mOsARF18 leaves were abnormal in bulliform cell differentiation and epidermal cell division. Starch accumulation in mOsARF18 seeds was also reduced. Moreover, auxin induced expression of OsMIR160a, OsMIR160b, and OsARF18, whereas expression of OsMIR160a and OsMIR160b as well as genes involved in auxin signaling was altered in mOsARF18 plants. Our results show that negative regulation of OsARF18 expression by OsmiR160 is critical for rice growth and development via affecting auxin signaling, which will advance future studies on the molecular mechanism by which miR160 fine-tunes auxin signaling in plants.