polyGBLUP: a modified genomic best linear unbiased prediction improved the genomic prediction efficiency for autopolyploid species.

Given the universality of autopolyploid species in nature, it is crucial to develop genomic selection methods that consider different allele dosages for autopolyploid breeding. However, no method has been developed to deal with autopolyploid data regardless of the ploidy level. In this study, we developed a modified genomic best linear unbiased prediction (GBLUP) model (polyGBLUP) through constructing additive and dominant genomic relationship matrices based on different allele dosages. polyGBLUP could carry out genomic prediction for autopolyploid species regardless of the ploidy level. Through comprehensive simulations and analysis of real data of autotetraploid blueberry and guinea grass and autohexaploid sweet potato, the results showed that polyGBLUP achieved higher prediction accuracy than GBLUP and its superiority was more obvious when the ploidy level of autopolyploids is high. Furthermore, when the dominant effect was added to polyGBLUP (polyGDBLUP), the greater the dominance degree, the more obvious the advantages of polyGDBLUP over the diploid models in terms of prediction accuracy, bias, mean squared error and mean absolute error. For real data, the superiority of polyGBLUP over GBLUP appeared in blueberry and sweet potato populations and a part of the traits in guinea grass population due to the high correlation coefficients between diploid and polyploidy genomic relationship matrices. In addition, polyGDBLUP did not produce higher prediction accuracy than polyGBLUP for most traits of real data as dominant genetic variance was not captured for these traits. Our study will be a significant promising method for genomic prediction of autopolyploid species.

Pyruvate kinase M2 is a PHD3-stimulated coactivator for hypoxia-inducible factor 1.

The pyruvate kinase isoforms PKM1 and PKM2 are alternatively spliced products of the PKM2 gene. PKM2, but not PKM1, alters glucose metabolism in cancer cells and contributes to tumorigenesis by mechanisms that are not explained by its known biochemical activity. We show that PKM2 gene transcription is activated by hypoxia-inducible factor 1 (HIF-1). PKM2 interacts directly with the HIF-1α subunit and promotes transactivation of HIF-1 target genes by enhancing HIF-1 binding and p300 recruitment to hypoxia response elements, whereas PKM1 fails to regulate HIF-1 activity. Interaction of PKM2 with prolyl hydroxylase 3 (PHD3) enhances PKM2 binding to HIF-1α and PKM2 coactivator function. Mass spectrometry and anti-hydroxyproline antibody assays demonstrate PKM2 hydroxylation on proline-403/408. PHD3 knockdown inhibits PKM2 coactivator function, reduces glucose uptake and lactate production, and increases O(2) consumption in cancer cells. Thus, PKM2 participates in a positive feedback loop that promotes HIF-1 transactivation and reprograms glucose metabolism in cancer cells.

Cost-effective genomic prediction of critical economic traits in sturgeons through low-coverage sequencing.

Low-coverage whole-genome sequencing (LCS) offers a cost-effective alternative for sturgeon breeding, especially given the lack of SNP chips and the high costs associated with whole-genome sequencing. In this study, the efficiency of LCS for genotype imputation and genomic prediction was assessed in 643 sequenced Russian sturgeons (∼13.68×). The results showed that using BaseVar+STITCH at a sequencing depth of 2× with a sample size larger than 300 resulted in the highest genotyping accuracy. In addition, when the sequencing depth reached 0.5× and SNP density was reduced to 50 K through linkage disequilibrium pruning, the prediction accuracy was comparable to that of whole sequencing depth. Furthermore, an incremental feature selection method has the potential to improve prediction accuracy. This study suggests that the combination of LCS and imputation can be a cost-effective strategy, contributing to the genetic improvement of economic traits and promoting genetic gains in aquaculture species.

The synergistic impact of probiotics and dietary fiber on constipation management in chlorpromazine-induced schizophrenia patients: exploring the modulation of intestinal microbiota and neurotrophic factors.

OBJECTIVE: To explore the improvement effect of probiotics combined with dietary fiber on constipation in patients with schizophrenia. METHODS: To compare the improvement scores of constipation, constipation symptoms, quality of life, neurotrophic factors-related indicators, and clinical efficacy between the two groups. RESULTS: There was no statistically significant difference in Cleveland Constipation Scoring System (CCS) scores in the control group before and after treatment (p > 0.05), while the CCS scores in the observation group decreased significantly after treatment (p < 0.05); Patient Assessment of Constipation Symptoms scores significantly decreased in the observation group compared to the control group (p < 0.05), with no significant difference in Patient Assessment of Constipation Quality of Life scores between the two groups pre- and post-treatment; Neuron-specific enolase levels decreased significantly in both groups post-treatment, while brain-derived neurotrophic factor, neuregulin-1, and nerve growth factor levels increased significantly, with a more pronounced rise in the observation group (p < 0.05). Additionally, the total effective rate of clinical treatment in the observation group was higher than that in the control group (p < 0.05). CONCLUSION: Probiotics combined with dietary fiber can improve constipation symptoms in patients with schizophrenia accompanied by constipation, effectively maintain the balance of intestinal microbiota, and improve the quality of life of patients. Additionally, levels of neurotrophic factors associated with bowel function and neurological health increased significantly, with a higher total effective rate of clinical treatment observed in the probiotics and dietary fiber group. These findings suggest the potential efficacy of probiotics and dietary fiber in managing constipation in this patient population.

Reproductive Toxicity and Teratogenicity of Fluorene-9-bisphenol on Chinese Medaka (Oryzias sinensis): A Study from Laboratory to Field.

Fluorene-9-bisphenol (BHPF), a bisphenol A (BPA) substitute, has been increasingly used as a material in syntheses of polymers that are widely used in road markings, artificial tracks, coating floors, building paints, etc., increasing the likelihood of BHPF contamination in the aquatic environment due to its release from the products. However, to date, it is unknown whether it may have actual impacts on fish in real environments. In this study, a 105-day exposure experiment of BHPF at various concentrations (0.01, 0.1, 1, and 10 µg/L) on Chinese medaka (Oryzias sinensis) was performed under laboratory conditions and found decreased fecundity, such as lower egg qualities and quantities, retarded oogenesis, and atretic follicles in the fish and deformed eyes and bodies in its F1 generation. Toxico-transcriptome analyses showed that estrogen-responsive genes were significantly suppressed by BHPF, indicating that antagonist properties of BHPF on estrogen receptors might be causes for the decreased fecundity. Field investigations (Beijing) demonstrated that BHPF was detectable in 60% surface waters, with a mean concentration of 10.49 ± 6.33 ng/L, by gas chromatography-mass spectrometry, and similar effects in wild Chinese medaka were also observed, some of which the parameters were found to be obviously correlated with the BHPF levels in corresponding waters.

The immune response of Locusta migratoria manilensis at different times of infection with Paranosema locustae.

Paranosema locustae is an entomopathogenic microsporidia with promising potential for controlling agricultural pests, including Locusta migratoria manilensis. However, it has the disadvantage of having a slow insecticidal rate, and how P. locustae infection impacts the host immune response is currently unknown. The present study investigated the effect of P. locustae on the natural immune response of L. migratoria and the activities of enzymes that protect against oxidative stress. Infection with P. locustae increased the hemocytes and nodulation number of L. migratoria at the initial stage of infection. The hemocyte-mediated modulation of immune response was also affected by a decrease in the number of hemocytes 12 days postinfection. Superoxide dismutase activity in locusts increased in the early stages of infection but decreased in the later stages, whereas the activities of peroxidase (POD) and catalase (CAT) showed opposite trends may be due to their different mechanisms of action. Furthermore, the transcription levels of mRNA of antimicrobial peptide-related genes and phenoloxidase activity in hemolymph in L. migratoria were suppressed within 15 days of P. locustae infection. Overall, our data suggest that P. locustae create a conducive environment for its own proliferation in the host by disrupting the immune defense against it. These findings provide useful information for the potential application of P. locustae as a biocontrol agent.

Molecular and biochemical changes in Locusta migratoria (Orthoptera: Acrididae) infected with Paranosema locustae.

Microsporidia are a group of eukaryotic intracellular parasitic organisms that infect almost all vertebrates and invertebrates. Paranosema locustae are specialized parasites of Orthoptera that are often used as biological controls of locusts, with slow effects of action. In this study, we found that after infection with P. locustae, changes in energy metabolism in male and female Locusta migratoria as were consistent, with no gender differences. During the first 8 days of infection, L. migratoria used sugar as a source of energy. After 8 days, lipids and proteins were consumed to provide energy when the spore load was considerably heavy, and energy supply was insufficient. With increasing infection concentration and time, energy conversion from sugar, fats, and proteins was improved, which may explain why high mortality did not occur until about 15 days after P. locustae infection. The tandem mass tag-based quantitative proteomics analysis revealed that most altered metabolism-related proteins were upregulated (27 of 29 in the metabolic pathway). This result suggests that P. locustae infection accelerated metabolism in L. migratoria, which facilitated the pathogen's life cycle, inhibiting the growth and development of the locusts and eventually killing them. Our findings will be useful to better understand of the chronic pathogenic mechanisms of P. locustae and inform on applications of P. locustae to control locusts.

Presence of GnRH3 in sturgeon and the roles of GnRH1 and GnRH2 on the regulation of LH/FSH in mature female sterlet Acipenser ruthenus in vitro.

Three forms of gonadotropin-releasing hormones (GnRHs), ArGnRH1, ArGnRH2, and ArGnRH3, were identified in sterlet. Compared with their orthologue, ArGnRH1 and ArGnRH2 have conserved core decapeptide but show low identity in the signal peptide and the rest of the sequences. The existence of the GnRH3 paralogue of sturgeon was predicted for the first time with TBLASTN by using the amino acid sequences of catshark and whale shark GnRH3 precursor as queries against the whole genome and transcript data of sterlet. The predicted ArGnRH3 cDNA sequence was composed of three exons containing all the elements of the GnRH family. The successful molecular cloning of GnRH3 from sterlets verified its expression in the brain of sturgeons. The analysis of the ArGnRH3 amino acid sequence revealed a completely conserved decapeptide sequence that shows 100% identity with the sequence of teleosts and differs in one amino acid with that of the cartilaginous fish (catshark and whale shark) at the 5th position. The structure of the phylogenetic tree showed that a total of 52 vertebrate GnRH sequences were clustered into three main clades corresponding to GnRH1, GnRH2, and GnRH3. The ArGnRH3 sequence is the oldest GnRH3 identified in teleosts. The tissue distribution analysis showed that ArGnRH1 was expressed in all the 13 examined tissues of females and in most of the tested tissues of male fish, with the highest expression in the pituitary and hypothalamus. ArGnRH2 is only expressed in the pituitary, hypothalamus, and gonads of both female and male sterlets. ArGnRH3 mRNA could be detected in the pituitary, hypothalamus, and gonad in both female and male fish. It is also present in the spleen, head kidney, and gill in female fish and in kidney and heart in male fish. However, the ArGnRH3 only showed weak expression in all the positive tissues. ArGnRH1 and ArGnRH2 active decapeptides were synthesized to investigate their roles on the regulation of LH/FSH using a mixed brain cell line from a sexually mature female sterlet. The results showed that ArGnRH1 and ArGnRH2 exerted different effects on the gene expression and release of gonadotropins. ArGnRH1 promoted the expression of fshß significantly around 48 h, and the expression was suppressed when the treatment time was extended to 72 h. ArGnRH1 had no significant effects on the level of either mRNA or secreted lh in any of the tested treatment length or concentrations. Moreover, ArGnRH1 did not stimulate the activity of gonadotropins in the maturation stage of female sturgeons. ArGnRH2 promoted the expression of fshß at 24 h and 48 h and increased mRNA level of lhß at 6 h and 48 h, accompanied by the significant secretion of LH at 72 h, although the high mRNA level of fsh did not correlate with the secretion of FSH in ArGnRH2-treated groups. In conclusion, ArGnRH2 plays an important role in the maturation stage of female sterlets. Therefore, ArGnRH2 has the potential to induce ovulation and spermiation in sturgeons.

Molecular Variation and Genomic Function of Citrus Vein Enation Virus.

In this study, we identified a new citrus vein enation virus (CVEV) isolate (named CVEV-DT1) through sRNA high-throughput sequencing and traditional sequencing. Phylogenetic analysis based on whole genome sequences of all known CVEV isolates revealed that CVEV-DT1 was in an evolutionary branch with other isolates from China. Molecular variation analysis showed that the single nucleotide variability along CVEV full-length sequences was less than 8%, with more transitions (60.55%) than transversions (39.43%), indicating a genetically homogeneous CVEV population. In addition, non-synonymous nucleotide mutations mainly occurred in ORF1 and ORF2. Based on disorder analysis of all encoded ORF by CVEV-DT1, we identified that the CVEV-DT1 coat protein (CP) formed spherical granules, mainly in the cell nucleus and partly throughout the cytoplasm, with liquid properties through subcellular localization and photobleaching assay. Furthermore, we also confirmed that the CVEV P0 protein has weak post-transcriptional RNA-silencing suppressor activity and could elicit a strong hypersensitive response (HR) in tobacco plants. Collectively, to the best of our knowledge, our study was the first to profile the genomic variation in all the reported CVEV isolates and reveal the functions of CVEV-DT1-encoded proteins.

Lipase is associated with deltamethrin resistance in Culex pipiens pallens.

The wide application of pyrethroids has led to the rapid development of insecticide resistance in mosquitoes, leading to a rise in mosquito-borne diseases. We previously identified five differentially expressed lipase family genes upon evaluating the transcriptomes of deltamethrin-resistant and deltamethrin-susceptible strains of Culex pipiens pallens. Herein, the gene expression levels were verified by quantitative real-time PCR, and two lipase family genes, lipase A and pancreatic triacylglycerol lipase A, were chosen for further investigations. Using cell viability assays and Centers for Disease Control and Prevention bottle bioassays, lipase A was found to increase the resistance of mosquitoes against deltamethrin both in vitro and in vivo. Our findings indicate that lipase A is involved in conferring deltamethrin resistance in Cx. pipiens pallens.

Molecular characterization of manganese superoxide dismutase (MnSOD) from sterlet Acipenser ruthenus and its responses to Aeromonas hydrophila challenge and hypoxia stress.

A novel gene encoding the mitochondrial manganese superoxide dismutase from sterlet Acipenser ruthenus (Ar-MnSOD) was cloned. The full-length cDNA of MnSOD was of 1040 bp with a 672 bp open reading frame encoding 224 amino acids and the deduced amino acid sequence was located in mitochondria. Sequence comparison analysis showed that Ar-MnSOD was highly similar to MnSODs of invertebrates and vertebrates, especially those of freshwater Cyprinidae fishes and mammals. Phylogenetic analysis revealed that Ar-MnSOD was distant from MnSODs of other fishes and belonged to the family of mitochondrial MnSODs (mMnSOD). Consistently, Ar-MnSOD was located in mitochondria. The 3D structure of Ar-MnSOD was predicted and the overall structure was similar to that of MnSODs of humans and the bay scallop Argopecten irradians. In addition, mRNA of Ar-MnSOD was detected to extensively express in all tissues, with the highest level in brain and liver. Spleen and head kidney inoculation of Aeromonas hydrophila led to a significant up-regulation of Ar-MnSOD transcript levels. Also, hypoxia induced a transient increase in transcription of Ar-MnSOD in the gills, but not in the heart and brain, suggesting metabolic depression in these vital organs. The results also implied the anti-hypoxia properties of Ar-MnSOD in the related tissues and proved that Ar-MnSOD was involved in the stress response and (anti) oxidative processes triggered by hypoxia. The results indicated that Ar-MnSOD is induced upon A. hydrophila infection and hypoxia, consistent with its role in host immune and stress-induced anti-oxidative responses.

Selection of Reference Genes for Normalization of Real-Time PCR Data in Calliptamus italicus (Orthoptera: Acrididae) Under Different Temperature Conditions.

Global warming has dominated worldwide climate change trends, and adaptability to high temperatures is the main factor underlying the spread of the pest Calliptamus italicus in Xinjiang Province, China. However, knowledge about the molecular mechanisms responsible for this adaptability and other related biological properties of C. italicus remain relatively unclear. Real-time quantitative polymerase chain reaction (RT-qPCR) is a key tool for gene expression analysis associated with various biological processes. Reference genes are necessary for normalizing gene expression levels across samples taken from specific experimental conditions. In this study, transcript level of five genes (GAPDH, 18S, TUB, ACT, and EF1α), commonly used as reference genes, were evaluated under nine different temperatures (27, 30, 33, 36, 39, 42, 45, 48, and 51°C) to assess their expression stability and further select the most suitable to be used on normalization of target gene expression data. Gene expression profiles were analyzed using geNorm, NormFinder, and BestKeeper software packages. The combined results demonstrated that the best-ranked reference genes for C. italicus are EF1α, GAPDH, and ACT under different thermal stress conditions. This is the first study that assesses gene expression analysis across a range of temperatures to select the most appropriate reference genes for RT-qPCR data normalization in C. italicus. These results should assist target gene expression analysis associated with heat stress in C. italicus.

Molecular Features and Expression Patterns of Vitellogenin Receptor in Calliptamus italicus (Orthoptera: Acrididae).

Vitellogenin receptor (VgR) mediates the intake of vitellin via oocytes, thus exerting an important role in vitellogenesis. In this study, reverse transcription-polymerase chain reaction (RT-PCR) and rapid-amplification of cDNA ends techniques were adopted to clone the CiVgR gene, namely the VgR gene of Calliptamus italicus, i.e., Orthopteran. The full length of CiVgR was 5,589 bp, and the open reading frame was estimated to be 5,265 bp, which encoded 1,754 amino acids (aa). Sequence alignment analysis showed that CiVgR belonged to the superfamily of low-density lipoprotein receptor genes, which contained several conserved domains, including ligand-binding domains, epidermal growth factor precursor homology domains, transmembrane domains, and cytoplasmic domains. However, no O-linked sugar domain was identified. Phylogenetic analysis showed that CiVgR had the closest genetic relationship to Blattarias. RT-PCR showed that CiVgR was only specifically expressed in the ovarian tissue of females. quantitative real time polymerase chain reaction showed that the transcription of CiVgR already appeared in the fourth-instar nymph of C. italicus, which gradually increased after adult emergence, peaked at the previtellogenesis stage, and then started to decrease. The expression pattern of CiVgR was closely associated with vitellogenesis. The findings of this study further our understanding of the molecular mechanisms involved in the reproduction of C. italicus, and provide new ideas to control this insect.

Chemotherapy triggers HIF-1-dependent glutathione synthesis and copper chelation that induces the breast cancer stem cell phenotype.

Triple negative breast cancer (TNBC) accounts for 10-15% of all breast cancer but is responsible for a disproportionate share of morbidity and mortality because of its aggressive characteristics and lack of targeted therapies. Chemotherapy induces enrichment of breast cancer stem cells (BCSCs), which are responsible for tumor recurrence and metastasis. Here, we demonstrate that chemotherapy induces the expression of the cystine transporter xCT and the regulatory subunit of glutamate-cysteine ligase (GCLM) in a hypoxia-inducible factor (HIF)-1-dependent manner, leading to increased intracellular glutathione levels, which inhibit mitogen-activated protein kinase kinase (MEK) activity through copper chelation. Loss of MEK-ERK signaling causes FoxO3 nuclear translocation and transcriptional activation of the gene encoding the pluripotency factor Nanog, which is required for enrichment of BCSCs. Inhibition of xCT, GCLM, FoxO3, or Nanog blocks chemotherapy-induced enrichment of BCSCs and impairs tumor initiation. These results suggest that, in combination with chemotherapy, targeting BCSCs by inhibiting HIF-1-regulated glutathione synthesis may improve outcome in TNBC.

Peritoneal bacterial infection repressed the expression of IL17D in Siberia sturgeon a chondrostean fish in the early immune response.

IL17s are pro-inflammatory cytokines that play important roles in host fighting against extracellular bacteria and auto-immune and allergic diseases. IL17D is believed to be the most ancient IL17 member and its functions are far from clarity. Although it has been found in invertebrates, jawless fish, teleosts, and tetrapods, it has not been described in chondrostean fish. Moreover, there are discrepancies concerning its expression pattern in these animals. In this study, we cloned and characterized the cDNA of il17d in Siberia sturgeon (Acipenser baerii), a chondrostean fish and commercially important species in aquaculture. The sturgeon il17d cDNA encodes a deduced protein of 210aa. The classical characteristics of IL17, such as IL17 domain, cysteine and serine residues importantly for cystine-knot formation, and signal peptide, were observed in sturgeon IL17D. Phylogenetic analysis and multiple alignment suggest it is a counterpart of mammalian IL17D. However, in vivo studies demonstrated that the expression pattern of sturgeon il17d mRNA is different from that of other teleosts and jawless fish, and in most cases its expression was down-regulated at the early time points and gradually increasing at late time points when sturgeon were challenged with bacteria (Aernomas hydrophila or Staphylococcus aureus). The In vitro study by using primary spleen cells stimulated with polyI:C revealed a similar expression pattern to that in vivo studies, while the stimulation with ß-glucan or LPS, which normally induced expression of il17d mRNA in target cells in vitro in other animals, did not show apparent changes in the expression of il17d mRNA. The results of present study indicated sturgeon IL17D may possess some different characteristics from its counterparts of other fish and invertebrates in the immune response, and may contribute to the understanding of IL17D functions in evolution as well as the potential use in sturgeon aquaculture.

Cyclin-dependent kinases regulate lysosomal degradation of hypoxia-inducible factor 1α to promote cell-cycle progression.

Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that mediates adaptive responses to oxygen deprivation. In addition, the HIF-1α subunit has a nontranscriptional role as a negative regulator of DNA replication through effects on minichromosome maintenance helicase loading and activation. However, some cell types continue to replicate under hypoxic conditions. The mechanism by which these cells maintain proliferation in the presence of elevated HIF-1α levels is unclear. Here we report that HIF-1α physically and functionally interacts with cyclin-dependent kinase 1 (Cdk1) and Cdk2. Cdk1 activity blocks lysosomal degradation of HIF-1α and increases HIF-1α protein stability and transcriptional activity. By contrast, Cdk2 activity promotes lysosomal degradation of HIF-1α at the G1/S phase transition. Blocking lysosomal degradation by genetic or pharmacological means leads to HIF-1α-dependent cell-cycle arrest, demonstrating that lysosomal degradation of HIF-1α is an essential step for the maintenance of cell-cycle progression under hypoxic conditions.

Dimorphic expression of sex-related genes in different gonadal development stages of sterlet, Acipenser ruthenus, a primitive fish species.

Molecular mechanism of sex determination and differentiation of sturgeon, a primitive fish species, is extraordinarily important due to the valuable caviar; however, it is still poorly known. The present work aimed to identify the major genes involved in regulating gonadal development of sterlet, a small species of sturgeon, from 13 candidate genes which have been shown to relate to gonadal differentiation and development in other teleost fish. The sex and gonadal development of sterlets were determined by histological observation and levels of sex steroids testosterone (T), 11-ketotestosterone (11-KT), and 17ß-estradiol (E2) in serum. Sexually dimorphic gene expressions were investigated. The results revealed that gonadal development were asynchronous in 2-year-old male and female sterlets with the testes in early or mid-spermatogenesis and the ovaries in chromatin nucleolus stage or perinucleolus stage, respectively. The levels of T and E2 were not significantly different between sexes or different gonadal development stages while 11-KT had the higher level in mid-spermatogenesis testis stage. In all the investigated gonadal development stages, gene dmrt1 and hsd11b2 were expressed higher in male whereas foxl2 and cyp19a1 were expressed higher in female. Thus, these genes provided the promising markers for sex identification of sterlet. It was unexpected that dkk1 and dax1 had significantly higher expression in ovarian perinucleolus stage than in ovarian chromatin nucleolus stage and in the testis, suggesting that these two genes had more correlation with ovarian development than with the testis, contrary to the previous reports in other vertebrates. Testicular development-related genes (gsdf and amh) and estrogen receptor genes (era and erb) differentially expressed at different testis or ovary development stages, but their expressions were not absolutely significantly different in male and female, depending on the gonadal development stage. Expression of androgen receptor gene ar or rspo, which was supposed to be related to ovarian development, presented no difference between gonadal development stages investigated in this study whenever in male or female.

MiR-285 targets P450 (CYP6N23) to regulate pyrethroid resistance in Culex pipiens pallens.

MicroRNAs play critical roles in post-transcriptional regulation of gene expression, which participate in the modulation of almost all of the cellular processes. Although emerging evidence indicates that microRNAs are related with antineoplastic drugs resistance, whether microRNAs are responsible for insecticide resistance in mosquitos is poorly understood. In this paper, we found that miR-285 was significantly upregulated in the deltamethrin-resistant strain of Culex pipiens pallens, and overexpression miR-285 through microinjection increased mosquito survival rate against deltamethrin treatement. Using bioinformatic software, quantitative reverse transcription PCR, luciferase reporter assay and microinjection approaches, we conformed that CYP6N23 was the target of miR-285. Lower expression of CYP6N23 was observed in the deltamethrin-resistant strain. While, mosquito mortality rate was decreased after downregulating expression of CYP6N23 by dsRNA against CYP6N23 or miR-285 mimic microinjection. These findings revealed that miR-285 could target CYP6N23 to regulate pyrethroid resistance, providing new insights into mosquito insecticide resistance surveillance and control.

Hypoxia-inducible factors regulate human and rat cystathionine ß-synthase gene expression.

Increased catalytic activity of CBS (cystathionine ß-synthase) was recently shown to mediate vasodilation of the cerebral microcirculation, which is initiated within minutes of the onset of acute hypoxia. To test whether chronic hypoxia was a stimulus for increased CBS expression, U87-MG human glioblastoma and PC12 rat phaeochromocytoma cells were exposed to 1% or 20% O2 for 24-72 h. CBS mRNA and protein expression were increased in hypoxic cells. Hypoxic induction of CBS expression was abrogated in cells transfected with vector encoding shRNA targeting HIF (hypoxia-inducible factor) 1α or 2α. Exposure of rats to hypobaric hypoxia (0.35 atm; 1 atm=101.325 kPa) for 3 days induced increased CBS mRNA, protein and catalytic activity in the cerebral cortex and cerebellum, which was blocked by administration of the HIF inhibitor digoxin. HIF-binding sites, located 0.8 and 1.2 kb 5' to the transcription start site of the human CBS and rat Cbs genes respectively, were identified by ChIP assays. A 49-bp human sequence, which encompassed an inverted repeat of the core HIF-binding site, functioned as a hypoxia-response element in luciferase reporter transcription assays. Thus HIFs mediate tissue-specific CBS expression, which may augment cerebral vasodilation as an adaptive response to chronic hypoxia.