Targeting EGFR degradation by autophagosome degraders.

Several generations of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors have been developed for the treatment of non-small cell lung cancer (NSCLC) in clinic. However, emerging drug resistance mediated by new EGFR mutations or activations by pass, leads to malignant progression of NSCLC. Proteolysis targeting chimeras (PROTACs) have been utilized to overcome the drug resistance acquired by mutant EGFR, newly potent and selective degraders are still need to be developed for clinical applications. Herein, we developed autophagosome-tethering compounds (ATTECs) in which EGFR can be anchored to microtubule-associated protein-1 light chain-3B (LC3B) on the autophagosome with the assistance of the LC3 ligand GW5074. A series of EGFR-ATTECs have been designed and synthesized. Biological evaluations showed that these compounds could degrade EGFR and exhibited moderate inhibitory effects on certain NSCLC cell lines. The ATTEC 12c potently induced the degradation of EGFR with a DC50 value of 0.98 µM and a Dmax value of 81% in HCC827 cells. Mechanistic exploration revealed that the lysosomal pathway was mainly involved in this degradation. Compound 12c also exhibited promising inhibitory activity, as well as degradation efficiency in vivo. Our study highlights that EGFR-ATTECs could be developed as a new expandable EGFR degradation tool and also reveals a novel potential therapeutic strategy to prevent drug resistance acquired EGFR mutations.

Cotyledonary somatic embryo is one kind of intermediate material similar to callus in the process of in vitro tissue culture from Rosa hybrida 'John F. Kennedy'.

BACKGROUND: Rose is recognized as an important ornamental plant worldwide, and it is also one of the most widely used flowers in gardens. At present, the improvement of rose traits is still difficult and uncertain, and molecular breeding can provide new ideas for the improvement of modern rose varieties. Somatic embryos are quite good receptors for genetic transformation. However, little is known about the molecular mechanisms underlying during the regeneration process of rose somatic embryos. To elucidate the molecular regulation mechanism of somatic embryo plantlet regeneration, the relationship between the differences in traits of the two different regenerated materials and the significantly differentially expressed genes (DEGs) related to phytohormone pathways in the process of regeneration were be investigated. RESULTS: These representative two regenerated samples from single-piece cotyledonary somatic embryo (SPC) culture of Rosa hybrida 'John F. Kennedy', were harvested for transcriptome analysis, with the SPC explants at the initial culture (Day 0) as the control. The differentially expressed genes (DEGs) in the materials from two different types for regeneration approach (SBF type: the regeneration approach type of single bud formed from SPC explants; MBF type: the regeneration approach type of multiple buds formed from SPC explants) were be screened by means of the transcriptome sequencing technology. In this study, a total of about 396.24 million clean reads were obtained, of which 78.95-82.92% were localized to the reference genome, compared with the initial material (CK sample), there were 5594 specific genes in the material of SBF type and 6142 specific genes in the MBF type. The DEGs from the SBF type material were mainly concentrated in the biological processes of GO terms such as phytohormones, substance transport, cell differentiation, and redox reaction. The KEGG enrichment analysis revealed these DEGs were more active in ubiquinone and other terpenoid-quinone biosynthesis, fatty acid elongation, steroid biosynthesis, and glycosphingolipid biosynthesis-globo and isoglobo series. In contrast, the DEGs induced by the MBF type material were mainly associated with the biological processes such as phytohormones, phosphorylation, photosynthesis and signal transduction. According to KEGG analysis, these DEGs of MBF type were significantly enriched in the porphyrin and chlorophyll metabolism, brassinosteroid biosynthesis, carotenoid biosynthesis, and peroxisome. Furthermore, the results from the phytohormone pathways analysis showed that the auxin-responsive factor SAUR and the cell wall modifying enzyme gene XTH were upregulated for expression but the protein phosphatase gene PP2C was downregulated for expression in SBF type; the higher expression of the ethylene receptor ETR, the ethylene transduction genes EBF1/2, the transcription factor EIN3, and the ethylene-responsive transcription factor ERF1/2 were induced by MBF type. CONCLUSIONS: According to the GO and KEGG analysis, it indicated the DEGs between two different regenerated materials from somatic embryos were significantly different which might be causing morphological differences. That was somatic embryos from Rosa hybrida 'John F. Kennedy' could regenerate plantlet via both classic somatic embryogenesis (seed-like germination) and organogenesis, cotyledonary somatic embryos should be considered as one kind of intermediate materials similiar to callus, rather than the indicator materials for somatic embryogenesis.

A New Biocontrol Agent Bacillus velezensis SF334 against Rubber Tree Fungal Leaf Anthracnose and Its Genome Analysis of Versatile Plant Probiotic Traits.

Natural rubber is an important national strategic and industrial raw material. The leaf anthracnose of rubber trees caused by the Colletotrichum species is one of the important factors restricting the yields of natural rubber. In this study, we isolated and identified strain Bacillus velezensis SF334, which exhibited significant antagonistic activity against both C. australisinense and C. siamense, the dominant species of Colletotrichum causing rubber tree leaf anthracnose in the Hainan province of China, from a pool of 223 bacterial strains. The cell suspensions of SF334 had a significant prevention effect for the leaf anthracnose of rubber trees, with an efficacy of 79.67% against C. siamense and 71.8% against C. australisinense. We demonstrated that SF334 can lead to the lysis of C. australisinense and C. siamense mycelia by causing mycelial expansion, resulting in mycelial rupture and subsequent death. B. velezensis SF334 also harbors some plant probiotic traits, such as secreting siderophore, protease, cellulase, pectinase, and the auxin of indole-3-acetic acid (IAA), and it has broad-spectrum antifungal activity against some important plant pathogenic fungi. The genome combined with comparative genomic analyses indicated that SF334 possesses most genes of the central metabolic and gene clusters of secondary metabolites in B. velezensis strains. To our knowledge, this is the first time a Bacillus velezensis strain has been reported as a promising biocontrol agent against the leaf anthracnose of rubber trees caused by C. siamense and C. australisinense. The results suggest that B. velezensis could be a potential candidate agent for the leaf anthracnose of rubber trees.

Development of Sustainable Engineered Cementitious Composites by Incorporating Local Recycled Fine Aggregate.

In this study, sustainable engineered cementitious composites (ECC) exhibiting high tensile strength as well as high tensile strain capacity were successfully developed by incorporating polyethylene (PE) fiber, local recycled fine aggregate (RFA), and limestone calcined clay cement (LC3). The improvement in tensile strength and tensile ductility was attributed to the self-cementing properties of RFA as well as the pozzolanic reaction between calcined clay and cement. Carbonate aluminates were also generated owing to the reaction between calcium carbonate in limestone and the aluminates in both calcined clay and cement. The bond strength between fiber and matrix was also enhanced. At the age of 150 days, the tensile stress-strain curves of ECC containing LC3 and RFA shifted from a bilinear model to a trilinear model, and the hydrophobic PE fiber exhibited hydrophilic bonding performance when embedded in RFA-LC3-ECC matrix, which could be explained by the densified cementitious matrix as well as the refined pore structure of ECC. Moreover, the substitution of ordinary Portland cement (OPC) by LC3 resulted in energy consumption and equivalent CO2 emission reduction ratios of 13.61% and 30.34%, respectively, when the replacement ratio of LC3 is 35%. Therefore, PE fiber-reinforced RFA-LC3-ECC demonstrates excellent mechanical performance as well as considerable environmental benefits.

Antagonistic transcriptome profile reveals potential mechanisms of action on Xanthomonas oryzae pv. oryzicola by the cell-free supernatants of Bacillus velezensis 504, a versatile plant probiotic bacterium.

Bacterial leaf streak (BLS) of rice is a severe disease caused by the bacterial pathogen Xanthomonas oryzae pv. oryzicola (Xoc) that has gradually become the fourth major disease on rice in some rice-growing regions in southern China. Previously, we isolated a Bacillus velezensis strain 504 that exhibited apparent antagonistic activity against the Xoc wild-type strain RS105, and found that B. velezensis 504 was a potential biocontrol agent for BLS. However, the underlying mechanisms of antagonism and biocontrol are not completely understood. Here we mine the genomic data of B. velezensis 504, and the comparative transcriptomic data of Xoc RS105 treated by the cell-free supernatants (CFSs) of B. velezensis 504 to define differentially expressed genes (DEGs). We show that B. velezensis 504 shares over 89% conserved genes with FZB42 and SQR9, two representative model strains of B. velezensis, but 504 is more closely related to FZB42 than SQR9, as well as B. velezensis 504 possesses the secondary metabolite gene clusters encoding the essential anti-Xoc agents difficidin and bacilysin. We conclude that approximately 77% of Xoc RS105 coding sequences are differentially expressed by the CFSs of B. velezensis 504, which significantly downregulates genes involved in signal transduction, oxidative phosphorylation, transmembrane transport, cell motility, cell division, DNA translation, and five physiological metabolisms, as well as depresses an additional set of virulence-associated genes encoding the type III secretion, type II secretion system, type VI secretion system, type IV pilus, lipopolysaccharides and exopolysaccharides. We also show that B. velezensis 504 is a potential biocontrol agent for bacterial blight of rice exhibiting relative control efficiencies over 70% on two susceptible cultivars, and can efficiently antagonize against some important plant pathogenic fungi including Colletotrichum siamense and C. australisinense that are thought to be the two dominant pathogenic species causing leaf anthracnose of rubber tree in Hainan province of China. B. velezensis 504 also harbors some characteristics of plant growth-promoting rhizobacterium such as secreting protease and siderophore, and stimulating plant growth. This study reveals the potential biocontrol mechanisms of B. velezensis against BLS, and also suggests that B. velezensis 504 is a versatile plant probiotic bacterium.

Multiscale Investigation on the Performance of Engineered Cementitious Composites Incorporating PE Fiber and Limstone Calcined Clay Cement (LC3).

Limestone calcined clay cement (LC3) is successfully used to fabricate engineered cementitious composites (ECC) exhibiting tensile strength σtu of 9.55 ± 0.59 MPa or tensile strain capacity εtu of 8.53 ± 0.30%. The high tensile strength of the composites is closely related to the improvement of fiber/matrix interfacial bond strength, and the high ductility is attributed to the enhancement of fiber dispersion homogeneity. For the case of ECC incorporating 50% LC3, the reduction of initial cracking stress σtc that favors the growth of the crack in a controlled manner also contributes to the improvement of strain hardening behavior. The composition analysis indicates that carboaluminates and additional hydration products including C-(A)-S-H and ettringite are generated, which contributes to the densification of the microstructure of the ECC matrix. The pore structure is thus remarkably refined. Besides, when ordinary Portland cement (OPC) is partly replaced by LC3, the consumed energy and equivalent CO2 emission decrease, especially the equivalent CO2 emission with the reduction ratio attaining 40.31%. It is found that ECC using 35% LC3 exhibits the highest mechanical resistance and ECC incorporating 50% LC3 shows the highest ductility from the environmental point of view.

Recycled GFRP Aggregate Concrete Considering Aggregate Grading: Compressive Behavior and Stress-Strain Modeling.

Fiber-reinforced polymer (FRP) composites have been used in various industries, thus a large amount of FRP wastes have been generated due to the out-of-service of FRP products. Recycling FRP wastes into coarse aggregates to replace natural coarse aggregates (NCA) to form the recycled FRP aggregate concrete (RFAC) is a potential approach to dispose of huge quantities of FRP wastes with low environmental impact. In this paper, waste glass FRP (GFRP) bars were cut into particles of 12 sizes to enable the grading of recycled FRP aggregate (RFA) as similar as possible to that of NAC. The influence of different RFA volume replacement ratios (0%, 30%, 50%, 70%, 100%) on the compressive performance of RFAC was investigated based on uniaxial compression tests of 15 standard cylinders. The results showed that the failure mode of RFAC was different from that of NAC. As the RFA replacement ratio increased, the compressive strength and elastic modulus of the RFAC gradually decreased, but its post-peak brittleness was significantly mitigated compared to NAC. The Poisson's ratio of RFAC increased with the increase in the RGFA replacement ratio at the elastic stage and was smaller than that of NCA concrete. Both the existing stress-strain models developed for NAC and recycled aggregate concrete (RAC) were found not fit for the RFAC. Thus, a new stress-strain model that was applicable to RFAC was developed by modifying the classical existing model, and a good agreement between the model predictions and test data was reached.