Genome-Wide Identification and Expression Analysis of Auxin Response Factor Gene Family in Linum usitatissimum.

Auxin response factors (ARFs) are critical components of the auxin signaling pathway, and are involved in diverse plant biological processes. However, ARF genes have not been investigated in flax (Linum usitatissimum L.), an important oilseed and fiber crop. In this study, we comprehensively analyzed the ARF gene family and identified 33 LuARF genes unevenly distributed on the 13 chromosomes of Longya-10, an oil-use flax variety. Detailed analysis revealed wide variation among the ARF family members and predicted nuclear localization for all proteins. Nineteen LuARFs contained a complete ARF structure, including DBD, MR, and CTD, whereas the other fourteen lacked the CTD. Phylogenetic analysis grouped the LuARFs into four (I-V) clades. Combined with sequence analysis, the LuARFs from the same clade showed structural conservation, implying functional redundancy. Duplication analysis identified twenty-seven whole-genome-duplicated LuARF genes and four tandem-duplicated LuARF genes. These duplicated gene pairs' Ka/Ks ratios suggested a strong purifying selection pressure on the LuARF genes. Collinearity analysis revealed that about half of the LuARF genes had homologs in other species, indicating a relatively conserved nature of the ARFs. The promoter analysis identified numerous hormone- and stress-related elements, and the qRT-PCR experiment revealed that all LuARF genes were responsive to phytohormone (IAA, GA3, and NAA) and stress (PEG, NaCl, cold, and heat) treatments. Finally, expression profiling of LuARF genes in different tissues by qRT-PCR indicated their specific functions in stem or capsule growth. Thus, our findings suggest the potential functions of LuARFs in flax growth and response to an exogenous stimulus, providing a basis for further functional studies on these genes.

Design and Construction of a Focused DNA-Encoded Library for Multivalent Chromatin Reader Proteins.

Chromatin structure and function, and consequently cellular phenotype, is regulated in part by a network of chromatin-modifying enzymes that place post-translational modifications (PTMs) on histone tails. These marks serve as recruitment sites for other chromatin regulatory complexes that 'read' these PTMs. High-quality chemical probes that can block reader functions of proteins involved in chromatin regulation are important tools to improve our understanding of pathways involved in chromatin dynamics. Insight into the intricate system of chromatin PTMs and their context within the epigenome is also therapeutically important as misregulation of this complex system is implicated in numerous human diseases. Using computational methods, along with structure-based knowledge, we have designed and constructed a focused DNA-Encoded Library (DEL) containing approximately 60,000 compounds targeting bi-valent methyl-lysine (Kme) reader domains. Additionally, we have constructed DNA-barcoded control compounds to allow optimization of selection conditions using a model Kme reader domain. We anticipate that this target-class focused approach will serve as a new method for rapid discovery of inhibitors for multivalent chromatin reader domains.

Consensus genetic linkage map construction and QTL mapping for plant height-related traits in linseed flax (Linum usitatissimum L.).

BACKGROUND: Flax is an important field crop that can be used for either oilseed or fiber production. Plant height and technical length are important characters for flax. For linseed flax, plants usually have a short technical length and plant height than those for fiber flax. As an important agronomical character for fiber and linseed flax, plant height is usually a selection target for breeding. However, because of limited technologies and methods available, there has been little research focused on discovering the molecular mechanism controlling plant height. RESULTS: In this study, two related recombinant inbred line (RIL) populations developed from crosses of linseed and fiber parents were developed and phenotyped for plant height and technical length in four environments. A consensus linkage map based on two RIL populations was constructed using SNP markers generated by genotyping by sequencing (GBS) technology. A total of 4497 single nucleotide polymorphism (SNP) markers were included on 15 linkage groups with an average marker density of one marker every 2.71 cM. Quantitative trait locus (QTL) mapping analysis was performed for plant height and technical length using the two populations. A total of 19 QTLs were identified for plant height and technical length. For the MH population, eight plant height QTLs and seven technical length QTLs were identified, five of which were common QTLs for both traits. For the PH population, six plant height and three technical length QTLs were identified. By comparing the QTLs and candidate gene information in the two population, two common QTLs and three candidate genes were discovered. CONCLUSIONS: This study provides a foundation for map-based cloning of QTLs and marker-assisted selection for plant height-related traits in linseed and fiber flax.

Electrospun N-Substituted Polyurethane Membranes with Self-Healing Ability for Self-Cleaning and Oil/Water Separation.

Membranes with special functionalities, such as self-cleaning, especially those for oil/water separation, have attracted much attention due to their wide applications. However, they are difficult to recycle and reuse after being damaged. Herein, we put forward a new N-substituted polyurethane membrane concept with self-healing ability to address this challenge. The membrane obtained by electrospinning has a self-cleaning surface with an excellent self-healing ability. Importantly, by tuning the membrane composition, the membrane exhibits different wettability for effective separation of oil/water mixtures and water-in-oil emulsions, whilst still displaying a self-healing ability and durability against damage. To the best of our knowledge, this is the first report to demonstrate a self-healing membrane for oil/water separation, which provides the fundamental research for the development of advanced oil/water separation materials.

Phosphorus and naphthalene acetic acid increased the seed yield by regulating carbon and nitrogen assimilation of flax.

To evaluate the impact of phosphorus (P) combined with exogenous NAA on flax yield, enhance flax P utilization efficiency and productivity, minimize resource inputs and mitigate negative environmental and human effects. Therefore, it is crucial to comprehend the physiological and biochemical responses of flax to P and naphthylacetic acid (NAA) in order to guide future agronomic management strategies for increasing seed yield. A randomized complete block design trial was conducted under semi-arid conditions in Northwest China, using a factorial split-plot to investigate the effects of three P (0, 67.5, and 135.0 kg P2O5 ha-1) and three exogenous spray NAA levels (0, 20, and 40 mg NAA L-1) on sucrose phosphate synthase (SPS) and diphosphoribulose carboxylase (Rubisco) activities as well as nitrogen (N) and P accumulation and translocation in flax. Results indicated that the SPS and Rubisco activities, N and P accumulation at flowering and maturity along with assimilation and translocation post-flowering, fruiting branches per plant, tillers per plant, capsules per plant, and seed yield were 95, 105, 14, 27, 55, 15, 13, 110, 103, 82, 16, 61, 8, and 13% greater in the P treatments compared to those in the zero P treatment, respectively. Moreover, those characteristics were observed to be greater with exogenous spray NAA treatments compared to that no spray NAA treatment. Additionally, the maximum SPS and Rubisco activities, N and P accumulation, assimilation post-flowering and translocation, capsules per plant, and seed yield were achieved with the application of 67.5 kg P2O5 ha-1 with 20 mg NAA L-1. Therefore, these findings demonstrate that the appropriate combination of P fertilizer and spray NAA is an effective agronomic management strategy for regulating carbon and nitrogen assimilation by maintaining photosynthetic efficiency in plants to increase flax productivity.

Synthesis of betulinic acid derivatives as entry inhibitors against HIV-1 and bevirimat-resistant HIV-1 variants.

Betulinic acid derivatives modified at the C28 position are HIV-1entry inhibitors such as compound A43D; however, modified at the C3 position instead of C28 give HIV-1 maturation inhibitor such as bevirimat. Bevirimat exhibited promising pharmacokinetic profiles in clinical trials, but its effectiveness was compromised by the high baseline drug resistance of HIV-1 variants with polymorphism in the putative drug binding site. In an effort to determine whether the viruses with bevirimat resistant polymorphism also altered their sensitivities to the betulinic acid derivatives that inhibit HIV-1 entry, a series of new betulinic acid entry inhibitors were synthesized and tested for their activities against HIV-1 NL4-3 and NL4-3 variants resistant to bevirimat. The results show that the bevirimat resistant viruses were approximately 5- to10-fold more sensitive to three new glutamine ester derivatives (13, 15 and 38) and A43D in an HIV-1 multi-cycle replication assay. In contrast, the wild type NL4-3 and the bevirimat resistant variants were equally sensitive to the HIV-1 RT inhibitor AZT. In addition, these three new compounds markedly improved microsomal stability compared to A43D.

Overexpression of WRINKLED1 improves the weight and oil content in seeds of flax (Linum usitatissimum L.).

Seeds of flax (Linum usitatissimum L.) are highly rich in both oil and linolenic acid (LIN). It is crucial for flax agricultural production to identify positive regulators of fatty acid biosynthesis. In this study, we find that WRINKLED1 transcription factors play important positive roles during flax seed oil accumulation. Two WRINKLED1 genes, LuWRI1a and LuWRI1b, were cloned from flax, and LuWRI1a was found be expressed predominantly in developing seeds during maturation. Overexpression of LuWRI1a increased seed size, weight, and oil content in Arabidopsis and increased seed storage oil content in transgenic flax without affecting seed production or seed oil quality. The rise in oil content in transgenic flax seeds was primarily attributable to the increase in seed weight, according to a correlational analysis. Furthermore, overexpression or interference of LuWRI1a upregulated the expression of genes in the fatty acid biosynthesis pathway and LAFL genes, and the expression level of WRI1 was highly significantly positively associated between L1L, LEC1, and BCCP2. Our findings give a theoretical scientific foundation for the future application of genetic engineering to enhance the oil content of plant seeds.

Phenotypic analysis of Longya-10 × pale flax hybrid progeny and identification of candidate genes regulating prostrate/erect growth in flax plants.

Flax is a dual-purpose crop that is important for oil and fiber production. The growth habit is one of the crucial targets of selection during flax domestication. Wild hybridization between cultivated flax and wild flax can produce superior germplasms for flax breeding and facilitate the study of the genetic mechanism underlying agronomically important traits. In this study, we used pale flax, Linum grandiflorum, and L. perenne to pollinate Longya-10. Only pale flax interspecific hybrids were obtained, and the trait analysis of the F1 and F2 generations showed that the traits analyzed in this study exhibited disparate genetic characteristics. In the F1 generation, only one trait, i.e., the number of capsules per plant (140) showed significant heterosis, while the characteristics of other traits were closely associated with those of the parents or a decline in hybrid phenotypes. The traits of the F2 generation were widely separated, and the variation coefficient ranged from 9.96% to 146.15%. The quantitative trait locus underlying growth habit was preliminarily found to be situated on chromosome 2 through Bulked-segregant analysis sequencing. Then linkage mapping analysis was performed to fine-map GH2.1 to a 23.5-kb interval containing 4 genes. Among them, L.us.o.m.scaffold22.109 and L.us.o.m.scaffold22.112 contained nonsynonymous SNPs with Δindex=1. Combined with the qRT-PCR results, the two genes might be possible candidate genes for GH2.1. This study will contribute to the development of important germplasms for flax breeding, which would facilitate the elucidation of the genetic mechanisms regulating the growth habit and development of an ideal architecture for the flax plant.

Synthesis and proteasome inhibition of lithocholic acid derivatives.

A new class of proteasome inhibitors was synthesized using lithocholic acid as a scaffold. Modification at the C-3 position of lithocholic acid with a series of acid acyl groups yielded compounds with a range of potency on proteasome inhibition. Among them, the phenylene diacetic acid hemiester derivative (13) displayed the most potent proteasome inhibition with IC(50) = 1.9 µM. Enzyme kinetic analysis indicates that these lithocholic acid derivatives are noncompetitive inhibitors of the proteasome.

Design and Synthesis of Quinolizidine Derivatives as Influenza Virus and HIV-1 Inhibitors.

BACKGROUND: We have previously reported that a quinolizidine natural product, aloperine, and its analogs can inhibit influenza virus and/or HIV-1 at low µM concentrations. OBJECTIVE: The main goal of this study was to further optimize aloperine for improved anti-influenza virus activity. METHODS: Structural modifications have been focused on the N12 position of aloperine scaffold. Conventional chemical synthesis was used to obtain derivatives with improved antiviral activities. The anti-HIV and anti-influenza virus activities of the synthesized compounds were determined using an MT4 cell-based HIV-1 replication assay and an anti- influenza virus infection of MDCK cell assay, respectively. RESULTS: Aloperine derivatives can be classified into three activity groups: those that exhibit anti-HIV activity only, anti-influenza virus only, or activity against both viruses. Aloperine optimized for potent anti-influenza activity often lost anti-HIV-1 activity, and vice versa. Compound 19 inhibited influenza virus PR8 replication with an IC50 of 0.091 µM, which is approximately 160- and 60-fold more potent than aloperine and the previously reported aloperine derivative compound 3, respectively. CONCLUSION: The data suggest that aloperine is a privileged scaffold that can be modified to become a selective antiviral compound with markedly improved potency against influenza virus or HIV-1.

Genomic Comparison and Population Diversity Analysis Provide Insights into the Domestication and Improvement of Flax.

Flax has been cultivated for its oil and fiber for thousands of years. However, it remains unclear how the modifications of agronomic traits occurred on the genetic level during flax cultivation. In this study, we conducted genome-wide variation analyses on multiple accessions of oil-use, fiber-use, landraces, and pale flax to identify the genomic variations during flax cultivation. Our findings indicate that, during flax domestication, genes relevant to flowering, dehiscence, oil production, and plant architecture were preferentially selected. Furthermore, regardless of origins, the improvement of the modern oil-use flax preceded that of the fiber-use flax, although the dual selection on oil-use and fiber-use characteristics might have occurred in the early flax domestication. We also found that the expansion of MYB46/MYB83 genes may have contributed to the unique secondary cell wall biosynthesis in flax and the directional selections on MYB46/MYB83 may have shaped the morphological profile of the current oil-use and fiber-use flax.

Structure Optimization of Aloperine Derivatives as HIV-1 Entry Inhibitors.

As a step toward developing novel anti-HIV agents, we have identified a class of quinolizidines, including aloperine, that inhibit HIV at 1-5 µM by blocking viral entry. In this study, we have optimized the structure of aloperine and derived compounds with markedly improved activity. Our structural optimization has yielded an aloperine derivative 19 with approximately a 15-fold increase in anti-HIV-1 activity. Our mechanism of action study reveals that compound 19 does not inhibit binding of HIV-1 to receptors but arrests the virus from fusion with the membrane. Binding of the compound to HIV-1gp120 might be responsible for its anti-HIV-1 entry activity.

In Situ and Ex Situ pH-Responsive Coatings with Switchable Wettability for Controllable Oil/Water Separation.

The development of stimuli-responsive materials with the ability of controllable oil/water separation is crucial for practical applications. Here, a novel pH responsive nonfluorine-containing copolymer was designed. The copolymer together with silica can be dip-coated on different materials including cotton fabric, filter paper, and polyurethane foam. The coated materials exhibit switchable superhydrophilicity and superhydrophobicity and can be applied in continuous separation of oil/water/oil three phase mixtures, different surfactant stabilized emulsion (oil-in-water, water-in-oil, and oil-in-acidic water) as well as oil uptake and release via in situ and ex situ pH change. We expect that the coatings highlight the practical applications because of the cost-effective preparation process and fluorine-free strategy.

Aloperine and Its Derivatives as a New Class of HIV-1 Entry Inhibitors.

A quinolizidine-type alkaloid aloperine was found to inhibit HIV-1 infection by blocking HIV-1 entry. Aloperine inhibited HIV-1 envelope-mediated cell-cell fusion at low micromolar concentrations. To further improve the antiviral potency, more than 30 aloperine derivatives with a variety of N12-substitutions were synthesized. Among them, 12d with an N-(1-butyl)-4-trifluoromethoxy-benzamide side chain showed the most potent anti-HIV-1 activity with EC50 at 0.69 µM. Aloperine derivatives inhibited both X4 and R5 HIV-1 Env-mediated cell-cell fusions. In addition, both BMS-806, a compound representing a class of HIV-1 gp120-targeting small molecules in clinical trials, and resistant and sensitive HIV-1 Env-mediated cell-cell fusions were equally sensitive to aloperine derivatives. These results suggest that aloperine and its derivatives are a new class of anti-HIV-1 entry inhibitors.

Phenolic diterpenoid derivatives as anti-influenza a virus agents.

A series of diterpenoid derivatives based on podocarpic acid were synthesized and evaluated as anti-influenza A virus agents. Several of the novel podocarpic acid derivatives exhibited nanomolar activities against an H1N1 influenza A virus (A/Puerto Rico/8/34) that was resistant to two anti-influenza drugs, oseltamivir and amantadine. This class of compounds inhibits the influenza virus by targeting the viral hemagglutinin-mediated membrane fusion. These results indicated that podocarpic acid derivatives may serve as potential drug candidates to fight drug-resistant influenza A virus infections.

Identification and synthesis of quinolizidines with anti-influenza a virus activity.

Influenza A virus infection causes a contagious respiratory illness that poses a threat to human health. However, there are limited anti-influenza A therapeutics available, which is further compounded by the emergence of drug resistant viruses. In this study, Sophora quinolizidine alkaloids were identified as a new class of anti-influenza A virus agents. Among the tested Sophora alkaloids, dihydroaloperine exhibited the most potent activity with an EC50 of 11.2 µM. The potency of the quinolizidine alkaloids was improved by approximately 5-fold with chemical modifications on the aloperine molecule. These compounds were effective against an H1N1 influenza A virus that was resistant to the two antiflu drugs oseltamivir and amantadine. The identification of the quinolizidine alkaloids as effective and novel anti-influenza A agents may aid in the development of new therapeutics.

New betulinic acid derivatives for bevirimat-resistant human immunodeficiency virus type-1.

Bevirimat (1, BVM) is an anti-HIV agent that blocks HIV-1 replication by interfering with HIV-1 Gag-SP1 processing at a late stage of viral maturation. However, clinical trials of 1 have revealed a high baseline drug resistance that is attributed to naturally occurring polymorphisms in HIV-1 Gag. To overcome the drug resistance, 28 new derivatives of 1 were synthesized and tested against compound 1-resistant (BVM-R) HIV-1 variants. Among them, compound 6 exhibited much improved activity against several HIV-1 strains carrying BVM-R polymorphisms. Compound 6 was at least 20-fold more potent than 1 against the replication of NL4-3/V370A, which carries the most prevalent clinical BVM-R polymorphism in HIV-1 Gag-SP1. Thus, compound 6 merits further development as a potential anti-AIDS clinical trial candidate.

Synthesis of Lithocholic Acid Derivatives as Proteasome Regulators.

Accumulation of aberrant protein aggregates, such as amyloid beta peptide (Aß), due to decreased proteasome activities might contribute to the neurodegeneration in Alzheimer's disease. In this study, lithocholic acid derivatives 3α-O-pimeloyl-lithocholic acid methyl ester (2) and its isosteric isomer (6) were found to activate the chymotrypsin-like activity of the proteasome at an EC(50) of 7.8 and 4.3 µM, respectively. Replacing the C24 methyl ester in 2 with methylamide resulted in a complete devoid of proteasome activating activity. Epimerizing the C3 substituent from an alpha to beta orientation transformed the activator into a proteasome inhibitor. Unlike the cellular proteasome activator PA28, proteasome activated by 2 was not inhibited by Aß. Furthermore, 2 potently antagonized the inhibitory effect of Aß on the proteasome. In summary, compound 2 represents a novel class of small molecules that not only activates the proteasome but also antagonizes the inhibitory effect of Aß on the proteasome.

Betulinic acid derivatives as human immunodeficiency virus type 2 (HIV-2) inhibitors.

We previously reported that [[N-[3beta-hydroxyllup-20(29)-en-28-oyl]-7-aminoheptyl]carbamoyl]methane (A43D, 4) was a potent HIV-1 entry inhibitor. However, 4 was inactive against HIV-2 virus, suggesting the structural requirements for targeting these two retroviruses are different. In this study, a series of new betulinic acid derivatives were synthesized, and some of them displayed selective anti-HIV-2 activity at nanomolar concentrations. In comparison to compounds with anti-HIV-1 activity, a shorter C-28 side chain is required for optimal anti-HIV-2 activity.

Synthesis and antibacterial activity of novel fluoroquinolones containing substituted piperidines.

The design and synthesis of new fluoroquinolone antibacterial agents having substituted piperidine rings at the C-7 position are described. Most of the new compounds demonstrated high in vitro antibacterial activity. Several of them exhibited significant activities against gram-positive organisms, which were more potent than those of gemifloxacin, Linezolid, and vancomycin.