Dynamic cytological and transcriptomic analyses provide novel insights into the mechanisms of sex determination in Castanea henryi.

Castanea henryi is a monoecious woody food tree species whose yield and industrialization potential are limited by its low female-to-male flower ratio. Here, the male flowers on the male inflorescence of C. henryi were converted to female flowers by triple applications of exogenous cytokinin (CK) (N-(2-chloro-4-pyridyl)-N'-phenylurea, CPPU). To study the role of exogenous CK in flower sex determination, cytological and transcriptomic analyses were performed on samples from the five stages after CK treatment. Cytological analysis showed that stage 3 (nine days after the last CK treatment) was the critical stage in the differential development of the pistil primordium and stamen primordium. On this basis, one key module and two modules with significant positive correlations with stage 3 were identified by weighted gene co-expression network analysis (WGCNA), combined with transcriptome data. The CK and GA biosynthesis- and signaling-related genes, three transcription factor (TF) families, and 11 floral organ identity genes were identified in the related modules. In particular, the TFs WRKY47, ERF021, and MYB4, and floral organ identity genes AGL11/15, DEF, and SEP1 with large differences are considered to be critical regulators of sex determination in C. henryi. Based on these results, a genetic regulatory network for exogenous CK in the sex determination of flowers in C. henryi is proposed. This study contributes to the understanding of the role of CK in the sex regulation of flowers and provides new insights into the regulatory network of sex determination in C. henryi.

Discovery of Linvencorvir (RG7907), a Hepatitis B Virus Core Protein Allosteric Modulator, for the Treatment of Chronic HBV Infection.

Described herein is the first-time disclosure of Linvencorvir (RG7907), a clinical compound and a hepatitis B virus (HBV) core protein allosteric modulator, for the treatment of chronic HBV infection. Built upon the core structure of hetero aryl dihydropyrimidine, RG7907 was rationally designed by combining all the drug-like features of low CYP3A4 induction, potent anti-HBV activity, high metabolic stability, low hERG liability, and favorable animal pharmacokinetic (PK) profiles. In particular, the chemistry strategy to mitigate CYP3A4 induction through introducing a large, rigid, and polar substituent at the position that has less interaction with the therapeutic biological target (HBV core proteins herein) is of general interest to the medicinal chemistry community. RG7907 demonstrated favorable animal PK, pharmacodynamics, and safety profiles with sufficient safety margins supporting its clinical development in healthy volunteers and HBV-infected patients.

Comparative analysis of the transcriptome during single-seed formation of Castanea henryi: regulation of starch metabolism and endogenous hormones.

BACKGROUND: In seed plants, the ovule is the precursor to the seed. The process of ovule development and differentiation is regulated by multiple factors, including starch metabolism and endogenous hormones. Castanea henryi produces nuts with high nutritional value. However, the high proportion of empty buds restricts the commercial use of the tree. Previous studies have shown that the empty bud phenotype is closely related to ovule abortion. If none of the ovules in the ovary expand rapidly and develop in 7-8 weeks after pollination, an empty bud will form. Therefore, we studied the development and molecular mechanisms underlying single seed formation in C. henryi. RESULTS: We found that 49 days after pollination (DAP) is a critical period for the formation of fertile and abortive ovules. The morphology and starch distribution of the fertile and abortive ovules differed significantly at 49 DAP. The fertile ovules were smooth and round in appearance, with a large amount of starch. In contrast, abortive ovules were smaller with only a small amount of starch. The embryo sac of the abortive ovule proceeded to develop abnormally, and the entire ovule lacked starch. We identified 37 candidate genes involved in metabolism with potential roles in the regulation of starch levels. Three ADP-glucose pyrophosphorylase (AGPase) genes, one granule-bound starch synthase (GBSS) gene, and two beta-amylase genes could affect starch accumulation. The levels of auxin, cytokinins, gibberellins, and jasmonic acid in fertile ovules were higher than those in abortive ovules. In addition, the levels of endogenous abscisic acid and salicylic acid in abortive ovules were higher than those in fertile ovules of the same age, consistent with the expression patterns of genes related to the synthesis of abscisic and salicylic acid and signal transduction. We identified and mapped the differentially expressed genes associated with hormone synthesis and signal transduction. CONCLUSIONS: These results improve our general understanding of the molecular mechanisms underlying single seed development in C. henryi and the phenomenon of empty buds, providing directions for future research.

Transcriptome Analysis Reveals the Regulatory Networks of Cytokinin in Promoting Floral Feminization in Castanea henryi.

Castanea henryi is a monoecious plant with a low female-to-male ratio, which limits its yield. The phytohormone cytokinin (CK) plays a crucial role in flower development, especially gynoecium development. Here, the feminizing effect of CK on the development of C. henryi was confirmed by the exogenous spraying of N-(2-chloro-4-pyridyl)-N'-phenylurea (CPPU). Spraying CPPU at 125 mg·L-1 thrice changed the male catkin into a pure female catkin, whereas at 5 mg·L-1 and 25 mg·L-1, only a part of the male catkin was transformed into a female catkin. A comparative transcriptome analysis of male catkins subjected to CPPU was performed to study the mechanism of the role of CKs in sex differentiation. Using Pearson's correlation analysis between hormone content and hormone synthesis gene expression, four key genes, LOG1, LOG3, LOG7 and KO, were identified in the CK and GA synthesis pathways. Moreover, a hub gene in the crosstalk between JA and the other hormone signaling pathways, MYC2, was identified, and 15 flowering-related genes were significantly differentially expressed after CPPU treatment. These results suggest that CK interacts with other phytohormones to determine the sex of C. henryi, and CK may directly target floral organ recognition genes to control flower sex.

[Effect of electronic moxibustion on memory function in patients with amnestic mild cognitive impairment].

OBJECTIVE: To observe the effect of electronic moxibustion on memory function in the patients with amnestic mild cognitive impairment (aMCI). METHODS: A total of 59 aMCI patients were randomized into an electronic moxibustion group (30 cases) and a placebo moxibustion group (29 cases). In the electronic moxibustion group, the electronic moxibustion was applied to Baihui (GV 20), Dazhui (GV 14), Mingmen (GV 4) and Taixi (KI 3), 45 ℃ in temperature, 20 min each time. The treatment was given once a day, 5 times a week. The treatment for 4 weeks was as one course and 2 courses were required totally. In the placebo moxibustion group, the moxa-free patch was used, 38 ℃ in temperature. The acupoint selection and the treatment frequency were same as the electronic moxibustion group. Before and after treatment, Rivermead behavior memory test (RBMT) was adopted to evaluate the global memory function of the patients in the two groups and the N-back task test was adopted to evaluate working memory function separately. Additionally, the mini-mental state examination (MMSE) and its immediate memory, Montreal cognitive assessment (MoCA) and its delay recall were adopted to evaluate the global cognitive function and memory function. RESULTS: In the electronic moxibustion group, after treatment, RBMT score, N-back accuracy rates, MMSE and MoCA scores and the scores of immediate memory and delay recall were improved significantly as compared with those before treatment (P<0.01). In the placebo moxibustion group, the accuracy rates of 1-back and 2-back task and the scores of immediate memory and delay recall were improved obviously as compared with those before treatment (P<0.05, P<0.01). After treatment, the improvements of RBMT score, the accuracy rates of N-back task and MMSE and MoCA scores in the electronic moxibustion group were higher than those in the placebo moxibustion group (P<0.05). CONCLUSION: Electronic moxibustion improves memory function in the patients with amnestic mild cognitive impairment.

Discovery of (aza)indole derivatives as novel respiratory syncytial virus fusion inhibitors.

A new class of indole derivatives (3) have been identified as potent RSV fusion inhibitors. SAR exploration revealed that 5-Cl and the sulfonyl side chain of the indole scaffold are crucial for anti-RSV activity. Further optimization led to the discovery of a cyclic sulfone (8i) with 2 nM anti-RSV activity and a much improved PK profile compared to the non-cyclic sulfone counterpart.

Measurement Method for Nonlinearity in Heterodyne Laser Interferometers Based on Double-Channel Quadrature Demodulation.

The phase quadrature measurement method is capable of measuring nonlinearity in heterodyne laser interferometers with picometer accuracy whereas it cannot be applied in the new kind of heterodyne interferometers with bidirectional Doppler frequency shift especially in the condition of non-uniform motion of the target. To solve this problem, a novel measurement method of nonlinearity is proposed in this paper. By employing double-channel quadrature demodulation and substituting the external reference signal with internal ones, this method is free from the type of heterodyne laser interferometer and the motion state of the target. For phase demodulation, the phase differential algorithm is utilized to improve the computing efficiency. Experimental verification is carried out and the results indicate that the proposed measurement method achieves accuracy better than 2 pm.

Highly thermal-stable heterodyne interferometer with minimized periodic nonlinearity.

Heterodyne interferometers suffer from thermal drift of optics (TDO), which may introduce error even up to several micrometers. In this paper, we propose a symmetric heterodyne interferometer with spatially separated beams, which realizes completely balanced optical paths between probe and reference beams and simultaneously avoids the optical mixing, and thereby is theoretically capable of eliminating TDO and the periodic nonlinearity (PNL). To validate the performance of the proposed interferometer, first a special experimental setup is constructed, with which the TDO test can be conducted in vacuum, and the result shows that a thermal coefficient of 1.2 nm/°C is achieved. Next, the PNL of the proposed interferometer is measured by both the frequency domain method and the phase quadrature method, which demonstrates an undetectable PNL at a noise level of 13 pm.

Discovery and Pre-Clinical Characterization of Third-Generation 4-H Heteroaryldihydropyrimidine (HAP) Analogues as Hepatitis B Virus (HBV) Capsid Inhibitors.

Described herein are the discovery and structure-activity relationship (SAR) studies of the third-generation 4-H heteroaryldihydropyrimidines (4-H HAPs) featuring the introduction of a C6 carboxyl group as novel HBV capsid inhibitors. This new series of 4-H HAPs showed improved anti-HBV activity and better drug-like properties compared to the first- and second-generation 4-H HAPs. X-ray crystallographic study of analogue 12 (HAP_R01) with Cp149 Y132A mutant hexamer clearly elucidated the role of C6 carboxyl group played for the increased binding affinity, which formed strong hydrogen bonding interactions with capsid protein and coordinated waters. The representative analogue 10 (HAP_R10) was extensively characterized in vitro (ADMET) and in vivo (mouse PK and PD) and subsequently selected for further development as oral anti-HBV infection agent.

Design and Synthesis of Orally Bioavailable 4-Methyl Heteroaryldihydropyrimidine Based Hepatitis B Virus (HBV) Capsid Inhibitors.

Targeting the capsid protein of hepatitis B virus (HBV) and thus interrupting normal capsid formation have been an attractive approach to block the replication of HBV viruses. We carried out multidimensional structural optimizations based on the heteroaryldihydropyrimidine (HAP) analogue Bay41-4109 (1) and identified a novel series of HBV capsid inhibitors that demonstrated promising cellular selectivity indexes, metabolic stabilities, and in vitro safety profiles. Herein we disclose the design, synthesis, structure-activity relationship (SAR), cocrystal structure in complex with HBV capsid proteins and in vivo pharmacological study of the 4-methyl HAP analogues. In particular, the (2S,4S)-4,4-difluoroproline substituted analogue 34a demonstrated high oral bioavailability and liver exposure and achieved over 2 log viral load reduction in a hydrodynamic injected (HDI) HBV mouse model.

Toward a nonlinearity model for a heterodyne interferometer: not based on double-frequency mixing.

Residual periodic errors detected in picometer-level heterodyne interferometers cannot be explained by the model based on double-frequency mixing. A new model is established and proposed in this paper for analysis of these errors. The multi-order Doppler frequency shift ghost beams from measurement beam itself are involved in final interference leading to multi-order periodic errors, whether or not frequency-mixing originating from the two incident beams occurs. For model validation, a novel setup free from double-frequency mixing is constructed. The analyzed measurement signal shows that phase mixing of measurement beam itself can lead to multi-order periodic errors ranging from tens of picometers to one nanometer.