Structures of Neisseria meningitidis Cas9 Complexes in Catalytically Poised and Anti-CRISPR-Inhibited States.

High-resolution Cas9 structures have yet to reveal catalytic conformations due to HNH nuclease domain positioning away from the cleavage site. Nme1Cas9 and Nme2Cas9 are compact nucleases for in vivo genome editing. Here, we report structures of meningococcal Cas9 homologs in complex with sgRNA, dsDNA, or the AcrIIC3 anti-CRISPR protein. DNA-bound structures represent an early step of target recognition, a later HNH pre-catalytic state, the HNH catalytic state, and a cleaved-target-DNA-bound state. In the HNH catalytic state of Nme1Cas9, the active site is seen poised at the scissile phosphodiester linkage of the target strand, providing a high-resolution view of the active conformation. The HNH active conformation activates the RuvC domain. Our structures explain how Nme1Cas9 and Nme2Cas9 read distinct PAM sequences and how AcrIIC3 inhibits Nme1Cas9 activity. These structures provide insights into Cas9 domain rearrangements, guide-target engagement, cleavage mechanism, and anti-CRISPR inhibition, facilitating the optimization of these genome-editing platforms.

Early change of plasma Epstein-Barr virus DNA load and the viral lytic genome level could positively predict clinical outcome in recurrent or metastatic nasopharyngeal carcinoma receiving anti-programmed cell death 1 monotherapy.

PURPOSE: Patients with recurrent or metastatic nasopharyngeal carcinoma (RM-NPC) have proven benefit from anti-programmed cell death 1 (anti-PD-1) monotherapy. Here, we retrospectively analyze the association of plasma Epstein-Barr virus (EBV) DNA load and tumor viral lytic genome with clinical outcome from 2 registered phase I trials. METHODS: Patients with RM-NPC from Checkmate 077 (nivolumab phase I trial in China) and Camrelizumab phase I trial between March 2016 and January 2018 were enrolled. Baseline EBV DNA titers were tested in 68 patients and EBV assessment was performed in 60 patients who had at least 3 post-baseline timepoints of EBV data and at least 1 post-baseline timepoint of radiographic assessment. We defined "EBV response" as 3 consecutive timepoints of load below 50% of baseline, and "EBV progression" as 3 consecutive timepoints of load above 150% of baseline. Whole-exome sequencing was performed in 60 patients with available tumor samples. RESULTS: We found that the baseline EBV DNA load was positively correlated with tumor size (spearman p < 0.001). Both partial response (PR) and stable disease (SD) patients had significantly lower EBV load than progression disease (PD) patients. EBV assessment was highly consistent with radiographic evaluation. Patients with EBV response had significantly improved overall survival (OS) than patients with EBV progression (log-rank p = 0.004, HR = 0.351 [95% CI: 0.171-0.720], median 22.5 vs. 11.9 months). The median time to initial EBV response and progression were 25 and 36 days prior to initial radiographic response and progression, respectively. Patients with high levels of EBV lytic genomes at baseline, including BKRF2, BKRF3 and BKRF4, had better progression-free survival (PFS) and OS. CONCLUSION: In summary, early clearance of plasma EBV DNA load and high levels of lytic EBV genes were associated with better clinical outcome in patients with RM-NPC receiving anti-PD-1 monotherapy.

The zona pellucida protein piopio regulates the metamorphosis and reproduction in Tribolium castaneum.

The zona pellucida domain protein piopio (Pio) was only reported to mediate the adhesion of the apical epithelial surface and the overlying apical extracellular matrix in Drosophila melanogaster, but the developmental roles of Pio were poorly understood in insects. To address this issue, we comprehensively analyzed the function of Pio in Tribolium castaneum. Phylogenetic analysis indicated that pio exhibited one-to-one orthologous relationship among insects. T. castaneum pio had a 1236-bp ORF and contained eight exons. During development pio was abundantly expressed from larva to adult and lowly expressed at the late stage of embryo and adult, while it had more transcripts in the head, epidermis, and gut but fewer in the fat body of late-stage larvae. Knockdown of pio inhibited the pupation, eclosion, and reproduction of T. castaneum. The expression of vitellogenin 1 (Vg1), Vg2, and Vg receptor (VgR) largely decreased in pio-silenced female adults. Silencing pio increased the 20-hydroxyecdysone titer by upregulating phm and spo expression but decreased the juvenile hormone (JH) titer through downregulating JHAMT3 and promoting JHE, JHEH-r4, and JHDK transcription. These results suggested that Pio might regulate the metamorphosis and reproduction via modulating the ecdysone and JH metabolism in T. castaneum. This study found the novel roles of pio in insect metamorphosis and reproduction, and provided the new insights for analyzing other zona pellucida proteins functions in insects.

Transcriptome analysis indicates the mechanisms of BmNPV resistance in Bombyx mori midgut.

Bombyx mori nucleopolyhedrovirus (BmNPV) caused serious economic losses in sericulture. Analyzing the molecular mechanism of silkworms (B. mori) resistance to BmNPV is of great significance for the prevention and control of silkworm virus diseases and the biological control of agricultural lepidopteran pests. In order to clarify the defense mechanisms of silkworms against BmNPV, we constructed a near isogenic line BC8 with high resistance to BmNPV through the highly BmNPV-resistant strain NB and the highly BmNPV-susceptible strain 306. In this study, RNA-Seq technique was used to analyze the transcriptome level differences in the midgut of BC8 and 306 following BmNPV infection. A total of 1350 DEGs were identified. Clustering analysis showed that these genes could be divided into 8 clusters with different expression patterns. Functional annotations based on GO and KEGG analysis indicated that they were involved in various metabolism pathways. Finally, 32 BmNPV defense responsive genes were screened. They were involved in metabolism, reactive oxygen species (ROS), signal transduction and immune response, and insect hormones. The further verification shows that HSP70 should participate in resistance responses of anti-BmNPV. These findings have paved the way in further functional characterization of candidate genes and subsequently can be used in breeding of BmNPV resistance dominant silkworms.

Mechanofluorochromic behaviors and data security protection properties of salicylaldimine-based difluoroboron complexes with different aryl substituents.

To further explore the relationship between aryl substituents and mechanofluorochromic (MFC) behaviors, four salicylaldimine-based difluoroboron complexes (ts-Ph BF2, ts-Ph-NA BF2, ts-2NA BF2, and ts-triphenylamine [TPA] BF2), including aromatic substituents with different steric hindrance effects, were designed and successfully synthesized. Four complexes with twisted molecular conformation displayed intramolecular charge transfer and aggregation-induced emission properties. Under external mechanical stimuli, the as-synthesized powders of ts-Ph BF2, ts-Ph-NA BF2, and ts-TPA BF2 exhibited redshift fluorescence emission behaviors, and ts-Ph BF2 and ts-TPA BF2 could be recovered to original shifts by fuming, but ts-Ph-NA BF2 displayed irreversible switching. ts-2NA BF2 had no change during the grinding and fuming processes. The results indicated that the MFC behaviors could be attributed to the phase transformation between the well-defined crystalline and disordered amorphous states by X-ray diffraction measurement. Further research illustrated that ts-TPA BF2 with the most significant MFC phenomenon could be applied in data security protection in ink-free rewritable paper.

The efficacy of levonorgestrelintrauterine system, drospirenone & ethinylestradiol tablets (II) and dydrogesterone in preventing the recurrence of endometrial polyps.

OBJECTIVE: To analyze the efficacy of levonorgestrelintrauterine system, Drospirenone & ethinylestradiol tablets (II), and dydrogesterone in preventing the recurrence of endometrial polyps after hysteroscopic endometrial polypectomy. METHODS: One hundred seventy patients who underwent hysteroscopic endometrial polypectomy in the Gynecology Department of Tianmen First People's Hospital in Hubei Province from January 2022 to June 2023 were randomly divided into the levonorgestrelintrauterine system group, Drospirenone & ethinylestradiol tablets (II) group, dydrogesterone group, and a control group. The recurrence rates, endometrial thickness, and menstrual volume changes at 6 and 12 months post-operation were compared among these four groups. RESULTS: The recurrence rates in the levonorgestrelintrauterine system group, Drospirenone & ethinylestradiol tablets (II) group, and dydrogesterone group were lower than the control group, with statistical significance (P < 0.01), with the levonorgestrelintrauterine system group having the lowest recurrence rate. The endometrial thickness at 6 and 12 months post-operation in the levonorgestrelintrauterine system group, Drospirenone & ethinylestradiol tablets (II) group, and dydrogesterone group was thinner than that of the control group and thinner than pre-operation, with statistical significance (P < 0.01). The menstrual volume at 3 months post-operation in the levonorgestrelintrauterine system group, Drospirenone & ethinylestradiol tablets (II) group, and dydrogesterone group was significantly less than the control group, and less than the pre-operation volume. CONCLUSION: Dydrogesterone, drospirenone & ethinylestradiol tablets (II), and levonorgestrelintrauterine system all play a role in preventing the recurrence of endometrial polyps, but levonorgestrelintrauterine system is significantly better than dydrogesterone and Drospirenone & ethinylestradiol tablets (II) in terms of postoperative recurrence rate, endometrial thickness, menstrual changes, and compliance, and is worth promoting in clinical application.

Interferon-γ release assay and mantoux response in infants with tuberculous meningitis in low and intermediate burden countries.

AIM: Until now, the performance of interferon-γ release assay (IGRA) and Mantoux tests remains unclear in infant tuberculous meningitis (TBM). Therefore, a systematic review is performed to evaluate the sensitivity of IGRA and Mantoux tests for the diagnosis of infant TBM in low and intermediate tuberculosis (TB) burden countries, while following PRISMA. METHODS: Several databases, including PubMed, EBSCO, Embase, Scopus, Web of Science, ClinicalTrials.gov, and Cochrane Central Register of Controlled Trials, were searched. Articles describing the results of IGRA or Mantoux tests among infant TBM were included for analysis. Data, such as age, sex, Mantoux test or IGRA, and cerebrospinal fluid (CSF) microbiological examinations (such as acid-fast bacilli (AFB) smear, TB PCR, and TB culture), were extracted from each study. RESULTS: A total of 31 articles were enrolled for further analysis, including 48 cases. The mean age was 9.4 ± 5.8 months and boys accounted for 57.1% of infants (24/42). Mantoux test was positive in 57.4% (27/47) of tested infants and IGRA was positive in 77.8% (7/9) of infants. In addition, among the infants with confirmed TB, 18 (52.9%, 18/34) of them have positive Mantoux responses and 7 (20.0%, 7/35) have positive IGRA results. CONCLUSIONS: In low or intermediate TB burden countries, the Mantoux test has a poor performance for diagnosing TBM among infants, and IGRAs appear to have a moderate sensitivity for the diagnosis of infant TBM.

Research progress on nucleic acid detection and genome editing of CRISPR/Cas12 system.

PURPOSE: This work characterizes the applications of CRISPR/Cas12 system, including nucleic acid detection, animal, plant and microbial genome editing. METHODS: The literature on CRISPR/Cas12 system was collected and reviewed. RESULTS: CRISPR/Cas system is an acquired immune system derived from bacteria and archaea, which has become the most popular technology around the world because of its outstanding contribution in genome editing. Type V CRISPR/Cas systems are distinguished by a single RNA-guided RuvC nuclease domain with single effector molecule. Cas12a, the first reported type V CRISPR/Cas system, targets double-stranded DNA (dsDNA) adjacent to PAM sequences and trans-cleaves single-stranded DNA (ssDNA). We present the applications of CRISPR/Cas12 system for nucleic acid detection and genome editing in animals, plants and microorganisms. Furthermore, this review also summarizes the applications of other Cas12 proteins, such as Cas12b, Cas12c, Cas12d, and so on, which further widen the application prospects of CRISPR/Cas12 system. CONCLUSIONS: Knowledge of the applications of CRISPR/Cas12 system is necessary for improving the understanding of the functional diversity of CRISPR/Cas12 system and also provides significant references for further research and utilization of CRISPR/Cas12 in other new fields.

Mechanofluorochromic behaviours of functionalized carbazole-based difluoroboron compounds with different aryl substituents moieties.

To better understand the relationship between molecular structure and mechanofluorochromic characteristics, three carbazole-based N^O-chelated difluoroboron compounds (Cz-S-BF2 , Cz-PhNp-S-BF2 , and Cz-BNp-S-BF2 ) with different aryl substituents moieties were designed and synthesized. The mechanofluorochromic behaviours of Cz-S-BF2 (luminescence from bluish-green to yellowish-green, emission from 504 to 535 nm) without aryl substitution and Cz-PhNp-S-BF2 (luminescence from green and yellow, emission from 521 to 557 nm) with a phenyl-naphthalene group underwent reversible conversion using the grinding-fuming process. For Cz-BNp-S-BF2 this was not apparent due to the well coplanarity of the binaphthalene moiety. Mechanofluorochromic properties were demonstrated through XRD patterns measurement. We envisage that this study will provide a practicable reference to acquire organic molecules with mechanofluorochromic characteristics.

Comparative transcriptome and proteome reveal synergistic functions of differentially expressed genes and proteins implicated in an over-dominant silkworm heterosis of increased silk yield.

We previously observed an over-dominant silkworm heterosis of increased yield in a cross of Bombyx mori nuclear polyhydrosis virus-resistant strain NB with a susceptible strain 306. In the present study, we found that heterosis also exists in crosses of NB with other susceptible strains, indicating it is a more general phenomenon. We performed comparative transcriptome and proteome and identified 1624 differentially expressed genes (DEGs) and 298 differentially expressed proteins (DEPs) in silk glands between parents and F1 hybrids, of which 24 DEGs/DEPs showed consistent expression at mRNA and protein levels revealed by Venn joint analysis. Their expressions are completely non-additive, mainly transgressive and under low-parent, suggesting recombination of parental genomes may be the major genetic mechanism for the heterosis. GO and KEGG analyses revealed that they may function in generally similar but distinctive aspects of metabolisms and processes with signal transduction and translation being most affected. Notably, they may not only up-regulate biosynthesis and transport of silk proteins but also down-regulate other unrelated processes, synergistically and globally remodelling the silk gland to increase yield and cause the heterosis. Our findings contribute insights into the understanding of silkworm heterosis and silk gland development and provide targets for transgenic manipulation to further increase the silk yield.

MicroRNA-384-5p protects against cardiac hypertrophy via the ALPK3 signaling pathway.

Heart failure is a condition caused by a variety of pathophysiological factors. One important pathological change of chronic heart failure is myocardial hypertrophy. In recent years, several studies have found that dysregulated microRNAs are involved in regulating the pathological process of heart failure. In this study, cardiac hypertrophy models were constructed using isoproterenol (ISO)-/angiotensin-II (Ang-II) to explore the role of miR-384-5p in cardiac hypertrophy and its molecular mechanism in vivo and in vitro. Echocardiography, invasive pressure-volume analysis and hematoxylin-eosin staining were used to explore cardiac structure and function. ALPK3 mRNA and protein expression were detected using quantitative reverse transcription polymerase chain reaction (RT-qPCR) and western blot analysis and miR-384-5p expression were assessed via RT-qPCR. Our findings determined that miR-384-5p was notably decreased in cardiac hypertrophic tissues and cells, and overexpression of miR-384-5p could ameliorate pressure overload. Furthermore, ALPK3 was determined to downregulate the ALPK3 expression to aggravate cardiomyocyte hypertrophy. Our findings provided a potential therapeutic target for the treatment of cardiac hypertrophy.

Connexin 37 Regulates the Kv1.3 Pathway and Promotes the Development of Atherosclerosis.

Objective: To investigate the mechanism of Connexin 37 (Cx37) and Kv1.3 pathways in atherosclerosis (AS). Methods: ApoE-/- mice were given a high-fat diet to establish atherosclerosis (AS) model, and macrophages in mice were isolated and extracted to transfect Cx37 vectors with silencing or overexpressing, and Kv1.3 pathway blockers were used to inhibit the pathway activity. The indexes of body weight, blood glucose, and blood lipid of mice were collected. The protein and mRNA expression levels of Cx37 and Kv1.3 were detected by reverse transcription-PCR (RT-PCR), Western blot, and immunofluorescence technique. Oil red O staining was used to observe plaque area. Masson staining was used to detect collagen content. The concentrations of chemokine CCL7 were quantified using the ELISA kits. CCK8 was used to detect cell proliferation. Results: Cx37 and Kv1.3 were highly expressed in macrophages of AS mice, and the expression of Kv1.3 and CCL7 decreased after Cx37 was silenced, and the proliferation of macrophages was also decreased. Wild-type mice and AS model mice were treated with Cx37 overexpression vectors and Kv1.3 pathway blocking, and it was found that Cx37 overexpression could improve the blood lipid and blood glucose levels and increase the area of AS in AS mice. However, blocking the activity of Kv1.3 pathway can reduce the levels of blood lipid and blood glucose, increase the body weight of mice, and reduce the area of AS mice. Blocking the activity of Kv1.3 pathway can slow down the plaque development of AS mice and make its indexes close to wild-type mice. And the use of Kv1.3 pathway blockers on the basis of overexpression of Cx37 indicated that inhibition of Kv1.3 pathway activity did not affect the expression of Cx37, but could inhibit the collagen content in the plaque area of AS mice, inhibit the expression of chemokine CCL7, and reverse the effect of Cx37 overexpression. Conclusion: Cx37 can improve the activity of macrophages by regulating the expression of chemokines and the activity of Kv1.3 pathway in AS mice, and enrich macrophages in inflammatory tissues and expand the area of plaque formation.

Multiple intestinal hemangioma concurrent with low-grade appendiceal mucinous neoplasm presenting as intussusception-a case report and literature review.

BACKGROUND: Cases with intussusception caused by either intestinal hemangiomas or appendiceal mucinous neoplasms are extremely rare. CASE PRESENTATION: In this study, we reported a 47-year-old male presented with paroxysmal abdominal pain and postprandial bloating for 3 days. CT results indicated a high possibility of secondary intussusception in ascending colon. Histopathology indicated a mixed type of cavernous and capillary hemangioma, combined with low-grade appendiceal mucinous neoplasms (LAMNs) and intestinal obstruction. The patient underwent laparotomy and right hemicolectomy. Finally, the patient was followed up for 4 months with no disease progression. CONCLUSIONS: Rare studies reported the intestine hemangiomas coincided with appendix low-grade mucinous tumor. Its manifestations are not specific, which is a challenge in the preoperative diagnosis. For cases with intussusception that was not observed in time, it may lead to intestinal necrosis and diffuse peritonitis. Additionally, the ruptured mucinous tumor in the appendix may lead to pathogenesis of pseudomyxoma peritonei. Therefore, accurate diagnosis and appropriate surgery-based treatment contribute to the improvement of prognosis and severe outcomes among these patients.

Down-regulation of MANNANASE7 gene in Brassica napus L. enhances silique dehiscence-resistance.

KEY MESSAGE: MANNANASE7 gene in Brassica napus L. encodes a hemicellulose which located at cell wall or extracellular space and dehiscence-resistance can be manipulated by altering the expression of MANNANASE7. Silique dehiscence is an important physiological process in plant reproductive development, but causes heavy yield loss in crops. The lack of dehiscence-resistant germplasm limits the application of mechanized harvesting and greatly restricts the rapeseed (Brassica napus L.) production. Hemicellulases, together with cellulases and pectinases, play important roles in fruit development and maturation. The hemicellulase gene MANNANASE7 (MAN7) was previously shown to be involved in the development and dehiscence of Arabidopsis (Arabidopsis thaliana) siliques. Here, we cloned BnaA07g12590D (BnMAN7A07), an AtMAN7 homolog from rapeseed, and demonstrate its function in the dehiscence of rapeseed siliques. We found that BnMAN7A07 was expressed in both vegetative and reproductive organs and significantly highly expressed in leaves, flowers and siliques where the abscission or dehiscence process occurs. Subcellular localization experiment showed that BnMAN7A07 was localized in the cell wall. The biological activity of the BnMAN7A07 protein isolated and purified through prokaryotic expression system was verified to catalyse the decomposition of xylan into xylose. Phenotypic studies of RNA interference (RNAi) lines revealed that down-regulation of BnMAN7A07 in rapeseed could significantly enhance silique dehiscence-resistance. In addition, the expression of upstream silique development regulators is altered in BnMAN7A07-RNAi plants, suggesting that a possible feedback regulation mechanism exists in the regulation network of silique dehiscence. Our results demonstrate that dehiscence-resistance can be manipulated by altering the expression of hemicellulase gene BnMAN7A07, which could provide an available genetic resource for breeding practice in rapeseed which is beneficial to mechanized harvest.

Tcf3-activated lncRNA Gas5 regulates newborn mouse cardiomyocyte apoptosis in diabetic cardiomyopathy.

Diabetic cardiomyopathy can cause cardiac dysfunction and eventually lead to heart failure and sudden death. Long noncoding RNA (lncRNA) Gas5 has been reported to play a function in cardiomyocyte. Here we studied the function of Gas5 on newborn mouse cardiomyocyte (NMC) apoptosis to detect its molecular mechanism. High-glucose treatment was implemented to induce the apoptosis of NMC in this study. And terminal deoxynucleotidyl transferase dUTP nick-end labeling assay, JC-1 assay, and flow cytometry analysis were conducted to know about the apoptosis of NMC when Gas5 and Tcf3 were silenced. Meanwhile, RNA pull-down assay and luciferase reporter assay were conducted to verify the binding of RNAs. Finally, rescue assay was implemented to evaluate the influence on apoptosis situation affected by competing endogenous RNA pathways. Tcf3 was found to bind to the Gas5 promoter to activate the expression of Gas5. Meanwhile, Gas5 and Tcf3 were both found to promote the apoptosis of NMC. Also, mmu-miR-320-3p could bind to Gas5 and Tcf3. Moreover, the Gas5/miR-320-3p/Tcf3 pathway was found to modulate the apoptosis of NMC. In conclusion, Tcf3-activated lncRNA Gas5 regulates NMC apoptosis in diabetic cardiomyopathy.

Genome-wide identification of the NPR1-like gene family in Brassica napus and functional characterization of BnaNPR1 in resistance to Sclerotinia sclerotiorum.

KEY MESSAGE: The BnaNPR1-like gene family was identified in B. napus, and it was revealed that repression of BnaNPR1 significantly reduces resistance toS. sclerotiorum, intensifies ROS accumulation, and changes the expression of genes associated with SA and JA/ET signaling in response to this pathogen. The NONEXPRESSOR OF PATHOGENESIS-RELATED GENES1 (NPR1) and related NPR1-like genes play an important role in regulating plant defense. Oilseed rape (Brassica napus L.) is an important oilseed crop; however, little is known about the B. napus (Bna) NPR1-like gene family. Here, a total of 19 BnaNPR1-like genes were identified in the B. napus genome, and then named according to their respective best match in Arabidopsis thaliana (At), which led to the determination of B. napus homologs of every AtNPR1-like gene. Analysis of important protein domains and functional motifs indicated the conservation and variation among these homologs. Phylogenetic analysis of these BnaNPR1-like proteins and their Arabidopsis homologs revealed six distinct sub-clades, consequently indicating that their name classification totally conformed to their phylogenetic relationships. Further, B. napus transcriptomic data showed that the expression of three BnaNPR1s was significantly down-regulated in response to infection with Sclerotinia sclerotiorum, the most important pathogen of this crop, whereas BnaNPR2/3/4/5/6s did not show the expression differences in general. Further, we generated B. napus BnaNPR1-RNAi lines to interpret the effect of the down-regulated expression of BnaNPR1s on resistance to S. sclerotiorum. The results showed that BnaNPR1-RNAi significantly decreased this resistance. Further experiments revealed that BnaNPR1-RNAi intensified ROS production and changed defense responses in the interaction of plants with this pathogen. These results indicated that S. sclerotiorum might use BnaNPR1 to regulate specific physiological processes of B. napus, such as ROS production and SA defense response, for the infection.

Pharmacokinetics and bioavailability of ipatasertib in dog plasma using LC/MS/MS.

A rapid, sensitive, and reliable liquid chromatography-tandem mass spectrometric method was developed to quantify ipatasertib in dog plasma. The dog plasma sample was deproteinated by using acetonitrile with ulixertinib as an internal standard followed by separation on a Spursil C18 -EP column with a gradient mobile phase comprising 2 mM ammonium acetate containing 0.1% formic acid and acetonitrile. Positive ion electrospray was used, and multiple reaction monitoring transitions were m/z 458.2 > 387.2 for ipatasertib and m/z 433.1 > 262.1 for the internal standard. The developed method was validated with a linear range of 0.3-1500 ng/mL, and with correlation coefficient greater than 0.9989. The lower limit of quantification was 0.3 ng/mL. The intra- and inter-day precision ranged from 3.58 to 14.32%, whereas the intra- and inter-day accuracy was in the range of -2.50-13.25%. No carry-over and matrix effects were observed under the current conditions. The extraction recovery was demonstrated to be greater than 85.43%. Ipatasertib was stable during the storage, processing, and determination. The validated assay was further successfully applied to a pharmacokinetic study of ipatasertib in dogs after oral and intravenous administrations. The bioavailability of ipatasertib was determined to be 19.3%.

Effects of cadmium-resistant plant growth-promoting rhizobacteria and Funneliformis mosseae on the cadmium tolerance of tomato (Lycopersicon esculentum L.).

Microorganisms are used to alleviate heavy metal stress in plants cultivated in contaminated fields. However, the relevant mechanisms have been rarely explored. The goal of this study was to investigate effects of arbuscular mycorrhizal fungus Funneliformis mosseae and two Cd-resistant bacterial strains (Enterobacter sp. EG16 and Enterobacter ludwigii DJ3) on growth and Cd tolerance of tomato when applied with different inoculation strategies (single or dual) and Cd concentrations (50 and 100 mg kg-1). Better plant growth was observed in mycorrhizal alone or combined treatments. In F. mosseae and EG16 co-inoculation treatment, shoot and root dry weight were 119-154% and 91-173% higher than those of the control, respectively. Higher bacterial and mycorrhizal colonization rate and root Cd concentration were also found in this treatment. However, the decrease of shoot Cd concentration and translocation factor values indicated this treatment was effective in improving Cd tolerance of the host plants. In addition, the increase in soil pH and decline in bioavailable Cd in the rhizosphere might be partly involved in reduction of Cd accumulation in plants. Our results suggest that co-inoculation with suitable microorganisms is important in plant growth and tolerance to Cd in Cd-contaminated soil.

Proteomic analysis of a clavata-like phenotype mutant in Brassica napus.

Rapeseed is one of important oil crops in China. Better understanding of the regulation network of main agronomic traits of rapeseed could improve the yielding of rapeseed. In this study, we obtained an influrescence mutant that showed a fusion phenotype, similar with the Arabidopsis clavata-like phenotype, so we named the mutant as Bnclavata-like (Bnclv-like). Phenotype analysis illustrated that abnormal development of the inflorescence meristem (IM) led to the fused-inflorescence phenotype. At the stage of protein abundance, major regulators in metabolic processes, ROS metabolism, and cytoskeleton formation were seen to be altered in this mutant. These results not only revealed the relationship between biological processes and inflorescence meristem development, but also suggest bioengineering strategies for the improved breeding and production of Brassica napus.

Reciprocal regulation of miR-206 and IL-6/STAT3 pathway mediates IL6-induced gefitinib resistance in EGFR-mutant lung cancer cells.

Persistently activated IL-6/STAT3 pathway promotes acquired resistance to targeted therapy with epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) in non-small-cell lung cancer (NSCLC) treatment. miR-206 has been verified to be dysregulated and plays as a negative regulator in lung cancer. However, whether miR-206 may overcome IL6-induced gefitinib resistance in EGFR-mutant lung cancer remains elusive. In this study, we investigated the role of miR-206 in IL6-induced gefitinib-resistant EGFR-mutated lung cancer cell lines. We showed that forced miR-206 expression restored gefitinib sensitivity in IL6-induced gefitinib-resistant EGFR-mutant lung cancer cells by inhibiting IL6/JAK1/STAT3 pathway. Specifically, mechanistic investigations revealed that miR-206 blocked IL-6/STAT3 signalling via directly targeting the 3'-UTR of intracellular IL-6 messenger RNA. Moreover, IL-6 induced miR-206 down-regulation by reducing the cropping process of primary miR-206 (pri-miR-206) into the Drosha/DGCR8 complex. Taken together, our findings reveal a direct role of miR-206 in regulating IL-6/STAT3 pathway and contrarily activated IL-6/STAT3 signalling mediates the miR-206 maturation process in gefitinib-resistant EGFR-mutant lung cancer cells.