Retrotransposon-mediated evolutionary rewiring of a pathogen response orchestrates a resistance phenotype in an insect host.

Ongoing host-pathogen interactions can trigger a coevolutionary arms race, while genetic diversity within the host can facilitate its adaptation to pathogens. Here, we used the diamondback moth (Plutella xylostella) and its pathogen Bacillus thuringiensis (Bt) as a model for exploring an adaptive evolutionary mechanism. We found that insect host adaptation to the primary Bt virulence factors was tightly associated with a short interspersed nuclear element (SINE - named SE2) insertion into the promoter of the transcriptionally activated MAP4K4 gene. This retrotransposon insertion coopts and potentiates the effect of the transcription factor forkhead box O (FOXO) in inducing a hormone-modulated Mitogen-activated protein kinase (MAPK) signaling cascade, leading to an enhancement of a host defense mechanism against the pathogen. This work demonstrates that reconstructing a cis-trans interaction can escalate a host response mechanism into a more stringent resistance phenotype to resist pathogen infection, providing a new insight into the coevolutionary mechanism of host organisms and their microbial pathogens.

METTL3 is essential for normal progesterone signaling during embryo implantation via m6A-mediated translation control of progesterone receptor.

Embryo implantation, a crucial step in human reproduction, is tightly controlled by estrogen and progesterone (P4) via estrogen receptor alpha and progesterone receptor (PGR), respectively. Here, we report that N6-methyladenosine (m6A), the most abundant mRNA modification in eukaryotes, plays an essential role in embryo implantation through the maintenance of P4 signaling. Conditional deletion of methyltransferase-like 3 (Mettl3), encoding the m6A writer METTL3, in the female reproductive tract using a Cre mouse line with Pgr promoter (Pgr-Cre) resulted in complete implantation failure due to pre-implantation embryo loss and defective uterine receptivity. Moreover, the uterus of Mettl3 null mice failed to respond to artificial decidualization. We further found that Mettl3 deletion was accompanied by a marked decrease in PGR protein expression. Mechanistically, we found that Pgr mRNA is a direct target for METTL3-mediated m6A modification. A luciferase assay revealed that the m6A modification in the 5' untranslated region (5'-UTR) of Pgr mRNA enhances PGR protein translation efficiency in a YTHDF1-dependent manner. Finally, we demonstrated that METTL3 is required for human endometrial stromal cell decidualization in vitro and that the METTL3-PGR axis is conserved between mice and humans. In summary, this study provides evidence that METTL3 is essential for normal P4 signaling during embryo implantation via m6A-mediated translation control of Pgr mRNA.

MAPK-mediated transcription factor GATAd contributes to Cry1Ac resistance in diamondback moth by reducing PxmALP expression.

The benefits of biopesticides and transgenic crops based on the insecticidal Cry-toxins from Bacillus thuringiensis (Bt) are considerably threatened by insect resistance evolution, thus, deciphering the molecular mechanisms underlying insect resistance to Bt products is of great significance to their sustainable utilization. Previously, we have demonstrated that the down-regulation of PxmALP in a strain of Plutella xylostella (L.) highly resistant to the Bt Cry1Ac toxin was due to a hormone-activated MAPK signaling pathway and contributed to the resistance phenotype. However, the underlying transcriptional regulatory mechanism remains enigmatic. Here, we report that the PxGATAd transcription factor (TF) is responsible for the differential expression of PxmALP observed between the Cry1Ac susceptible and resistant strains. We identified that PxGATAd directly activates PxmALP expression via interacting with a non-canonical but specific GATA-like cis-response element (CRE) located in the PxmALP promoter region. A six-nucleotide insertion mutation in this cis-acting element of the PxmALP promoter from the resistant strain resulted in repression of transcriptional activity, affecting the regulatory performance of PxGATAd. Furthermore, silencing of PxGATAd in susceptible larvae reduced the expression of PxmALP and susceptibility to Cry1Ac toxin. Suppressing PxMAP4K4 expression in the resistant larvae transiently recovered both the expression of PxGATAd and PxmALP, indicating that the PxGATAd is a positive responsive factor involved in the activation of PxmALP promoter and negatively regulated by the MAPK signaling pathway. Overall, this study deciphers an intricate regulatory mechanism of PxmALP gene expression and highlights the concurrent involvement of both trans-regulatory factors and cis-acting elements in Cry1Ac resistance development in lepidopteran insects.

Population genomics analysis reveals footprints of selective breeding in a rapid-growth variety of Paulownia fortunei with apical dominance.

Paulownia fortunei is an ecologically and economically valuable tree cultivated for its rapid growth and high-quality timber. To enhance Paulownia germplasm, we have developed the elite variety QingT with patented advantages in growth rate and apical dominance. To illuminate the genetic basis of QingT's superior traits, here we harness comparative population genomics to analyze genomic variation patterns between QingT and common Paulownia. We performed whole-genome re-sequencing of 30 QingT and 30 common samples, detecting 15.6 million SNPs and 2.6 million indels. Phylogeny and population structure analyses robustly partitioned common and QingT into distinct groups which indicate robust genome stabilization. QingT exhibited reduced heterozygosity and linkage disequilibrium decay compared to common Paulownia, reflecting high recombination, indicating hybridizing effects with common white-flowered string is the source of its patented advantages. Genome selection scans uncovered 25 regions of 169 genes with elevated nucleotide diversity, indicating selection sweeps among groups. Functional analysis of sweep genes revealed upregulation of ribosomal, biosynthesis, and growth pathways in QingT, implicating enhanced protein production and developmental processes in its rapid growth phenotype. This study's insights comprehensively chart genomic variation during Paulownia breeding, localizing candidate loci governing agronomic traits, and underpinnings of future molecular breeding efforts to boost productivity.

Development of Phenyl-substituted Isoindolinone- and Benzimidazole-type Cereblon Ligands for Targeted Protein Degradation.

Thalidomide, pomalidomide and lenalidomide, collectively referred to as immunomodulatory imide drugs (IMiDs), are frequently employed in proteolysis-targeting chimeras (PROTACs) as cereblon (CRBN) E3 ligase-recruiting ligands. However, their molecular glue properties that co-opt the CRL4CRBN to degrade its non-natural substrates may lead to undesired off-target effects for the IMiD-based PROTAC degraders. Herein, we reported a small library of potent and cell-permeable CRBN ligands, which exert high selectivity over the well-known CRBN neo-substrates of IMiDs by structure-based design. They were further utilized to construct bromodomain-containing protein 4 (BRD4) degraders, which successfully depleted BRD4 in the tested cells. Overall, we reported a series of functionalized CRBN recruiters that circumvent the promiscuity from traditional IMiDs, and this study is informative to the development of selective CRBN-recruiting PROTACs for many other therapeutic targets.

Recombinant L. lactis vaccine LL-plSAM-WAE targeting four virulence factors provides mucosal immunity against H. pylori infection.

BACKGROUND: Helicobacter pylori (H. pylori) causes chronic gastric disease. An efficient oral vaccine would be mucosa-targeted and offer defense against colonization of invasive infection in the digestive system. Proteolytic enzymes and acidic environment in the gastrointestinal tract (GT) can, however, reduce the effectiveness of oral vaccinations. For the creation of an edible vaccine, L. lactis has been proposed as a means of delivering vaccine antigens. RESULTS: We developed a plSAM (pNZ8148-SAM) that expresses a multiepitope vaccine antigen SAM-WAE containing Urease, HpaA, HSP60, and NAP extracellularly (named LL-plSAM-WAE) to increase the efficacy of oral vaccinations. We then investigated the immunogenicity of LL-plSAM-WAE in Balb/c mice. Mice that received LL-plSAM-WAE or SAM-WAE with adjuvant showed increased levels of antibodies against H. pylori, including IgG and sIgA, and resulted in significant reductions in H. pylori colonization. Furthermore, we show that SAM-WAE and LL-plSAM-WAE improved the capacity to target the vaccine to M cells. CONCLUSIONS: These findings suggest that recombinant L. lactis could be a promising oral mucosa vaccination for preventing H. pylori infection.

Regio- and Stereoselective Iodoamination of Ferrocene-Containing Allenylphosphonates: Synthesis of Multifunctional Tetrasubstituted Allylic Amines and Allylic Azides.

A general and practical methodology for the regio- and stereoselective synthesis of multifunctional tetrasubstituted allylic amines and azides based on iodoamination of ferrocene-containing allenylphosphonates with anilines and sodium azide is described. A tetrasubstituted olefin moiety, as well as an iodine atom, a phosphonate, and a ferrocene group, are installed to the allylic amine motif simultaneously in moderate to good yields.

An M cell-targeting recombinant L. lactis vaccine against four H. pylori adhesins.

The acidic environment and enzyme degradation lead to oral vaccines often having little immune effect. Therefore, it is an attractive strategy to study an effective and safe oral vaccine delivery system that can promote gastrointestinal mucosal immune responses and inhibit antigen degradation. Moreover, the antigens uptake by microfold cells (M cells) is the determining step in initiating efficient immune responses. Therefore, M cell-targeting is one promising approach for enhancing oral vaccine potency. In the present study, an M cell-targeting L. lactis surface display system (plSAM) was built to favor the multivalent epitope vaccine antigen (FAdE) to achieve effective gastrointestinal mucosal immunity against Helicobacter pylori. Therefore, a recombinant Lactococcus lactic acid vaccine (LL-plSAM-FAdE) was successfully prepared, and its immunological properties and protective efficacy were analyzed. The results showed that LL-plSAM-FAdE can secretively express the recombinant proteins SAM-FAdE and display the SAM-FAdE on the bacterial cell surface. More importantly, LL-plSAM-FAdE effectively promoted the phagocytosis and transport of vaccine antigen by M cells in the gastrointestinal tract of mice, and simulated high levels of cellular and humoral immune responses against four key H. pylori adhesins (Urease, CagL, HpaA, and Lpp20) in the gastrointestinal tract, thus enabling effective prevention of H. pylori infection and to some extent eliminating H. pylori already present in the gastrointestinal tract. KEY POINTS: • M-cell-targeting L. lactis surface display system LL- plSAM was designed • This system displays H. pylori vaccine-promoted phagocytosis and transport of M cell • A promising vaccine candidate for controlling H. pylori infection was verified.

Assessment of the Diagnostic Performance of Clinical Examinations and High-Frequency Ultrasound in Patients With Pigmented Skin Tumors.

OBJECTIVES: To investigate whether the integration of high-frequency ultrasound (HFUS) to routine clinical examinations could improve diagnostic performance and management decision for pigmented skin tumors. METHODS: Three general practitioners trained previously and a dermatologist independently assessed pigmented skin tumors and rendered management decision based on clinical examinations alone or clinical examinations integrating HFUS. RESULTS: After integrating HFUS, the diagnostic area under the curve (AUC) (0.658-0.693 versus 0.848, all P < .05) and specificity (46.6-58.6% versus 89.7%, all P < .05) for pigmented skin malignancies were improved for general practitioners, meanwhile unnecessary biopsy rate reduced (42.9-53.6% versus 10.7%, P < .001). To the dermatologist, the diagnostic AUC (0.822 versus 0.949, P < .001), sensitivity (81.7% versus 96.7%, P = .012) and specificity (0.828 versus 0.931, P = .031) improved significantly, meanwhile both missed biopsy rate (14.5% versus 4.8%, P = .031) and unnecessary biopsy rate (19.6% versus 7.1%, P = .016) decreased. Additionally, the diagnostic performance of the general practitioner with integrating HFUS could be comparable with the dermatologist based on clinical examinations alone (all P > .05). CONCLUSIONS: As a complementary tool of clinical examinations, HFUS could help physicians differentiate pigmented skin malignancies and manage decision.

Two Undescribed Germacrane-Type Sesquiterpenoids from Salvia cavaleriei var. simplicifolia Stib. and Their Anti-Alzheimer's Disease Activity.

Two undescribed germacrane-type sesquiterpenoids, salcasins A (1) and B (2), together with three known compounds (3-5) were isolated and identified from the whole plant of Salvia cavaleriei var. simplicifolia Stib. The structures of the undescribed compounds were elucidated on the basis of spectroscopic methods, such as HR-ESI-MS, 1D and 2D NMR data. The relative configurations of 1 and 2 were established by analyzing their NOESY spectra as well as by 13C NMR calculations with DP4+ probability analyses. The absolute configurations of 1 and 2 were determined by comparing experimental and calculated ECD spectra. Furthermore, the in vivo anti-Alzheimer's disease activities of 1-5 were evaluated using Caenorhabditis elegans AD pathological model. Among all isolated compounds, salcasin A (1) significantly delayed AD-like symptoms of worm paralysis, which may be a potential anti-AD candidate agent.

A Modular Chemistry Platform for the Development of a Cereblon E3 Ligase-Based Partial PROTAC Library.

Proteolysis targeting chimeras (PROTACs) are a promising therapeutic strategy to selectively promote the degradation of protein targets by exploiting the ubiquitin-proteasome system. Among the limited number of E3 ligase ligands discovered for the PROTAC technology, ligands of cereblon (CRBN) E3 ligase, such as pomalidomide, thalidomide, or lenalidomide, are the most frequently used for the development of PROTACs. Our group previously reported that a phenyl group could be tolerated on the C4-position of lenalidomide as the ligand of CRBN to develop PROTACs. Herein, we report a modular chemistry platform for the efficient attachment of various ortho-, meta-, and para-substituted phenyls to the C4-position of the lenalidomide via Suzuki cross-coupling reaction, which allows the systematic investigation of the linker effect for the development of PROTACs against any target. We examined the substrate scope by preparing twelve lenalidomide-derived CRBN E3 ligase ligands with different linkers.

Overexpression of POLA2 in hepatocellular carcinoma is involved in immune infiltration and predicts a poor prognosis.

BACKGROUND: Hepatocellular carcinoma (HCC) is the second malignancy worldwide. POLA2 initiates DNA replication, regulates cell cycle and gene repair that promote tumorigenesis and disease progression. However, the prognostic and biological function roles of POLA2 in HCC had not been conclusively determined. METHODS: The expression levels and prognosis role of POLA1 and POLA2 in HCC were analyzed based on TCGA-LIHC database and recruited 24 HCC patients. Gene mutations were analyzed using "maftools" package. POLA2 and immune cells correlations were analyzed by TIMER. POLA2 co-expressed genes functional enrichment were evaluated using Metascape. The mRNA and protein level of POLA2 was detected in HCC cells and tissues. Cell migration, invasion, proliferation, cell cycle and HCC cell lines derived xenograft model were performed to investigate POLA2 biological function. RESULTS: POLA2 was significantly high expressed in HCC than in normal liver tissue in both TCGA-LIHC and our collected HCC samples. In validation cohort, POLA2 significantly related to tumor differentiation, tumor size and Ki-67 (p < 0.05). In TCGA-LIHC cohort, overexpression of POLA2 predicted a low OS and associated with different clinical stages. Multivariate Cox regression showed overexpression of POLA2 effectively distinguished the prognosis at different T, N, M, stages and grades of HCC. POLA2 expression correlated with mutation burden, immune cells infiltration and immune-associated genes expression of HCC. Functional enrichment revealed that POLA2 co-expressed genes were linked to cellular activity, plasma membrane protein complex and leukocyte activity, immune response-regulated cell surface receptor signaling pathway, and immune response-regulated signaling pathway. Moreover, POLA2 was also positively co-expressed with some immune checkpoints (CD274, CTL-4, HAVCR2, PDCD1, PDCD1LG2, TIGIT, and LAG3) (p < 0.001). Gene knockdown revealed that POLA2 promoted proliferation, migration, invasion, and cell cycle of SMMC-7721 and HepG2. The HCC xenograft tumor model also demonstrated remarkably tumor size inhibition, tumor proliferation inhibtion and tumor necrosis promotion when POLA2 knockdown. CONCLUSIONS: POLA2 influenced immune microenvironment and tumor progression of HCC indicated that it might be a potential molecular marker for prognostic evaluation or a therapeutic target for HCC.

Value of conventional ultrasound and shear wave elastography in the assessment of muscle mass and function in elderly people with type 2 diabetes.

OBJECTIVES: We assessed muscle mass and function using ultrasound (US) and shear wave elastography (SWE) for sarcopenia in elderly patients with type 2 diabetes. METHODS: There were 84 patients with type 2 diabetes enrolled in this study; of these, 30 had sarcopenia and 54 did not. We measured appendicular skeletal muscle mass index (ASMI), handgrip strength, calf circumference, 6-m walking speed, and 5-time chair stand test. All patients were in the supine position with their knees in straight and bent poses in turn. The US-derived thickness (Tstraight, Tbent), cross-sectional area (CSAstraight, CSAbent), and SWE (SWEstraight, SWEbent) of the rectus femoris muscle (RFM) were measured and the differences (ΔT, ΔCSA, ΔSWE) were calculated. We assessed the correlations of clinical indicators with US and SWE features. We then compared the clinical indicators and US and SWE features between patients with and without sarcopenia to determine independent predictors. Diagnostic models were established based on these independent predictors. RESULTS: The ASMI was correlated with Tbent (r = 0.57, p < 0.001) and CSAbent (r = 0.50, p < 0.001). Handgrip strength was correlated with Tbent (r = 0.53, p < 0.001) and CSAbent (r = 0.51, p < 0.001). Between patients with and without sarcopenia, the indicators of age, ΔCSA, and ΔSWE were statically different (all p ≤ 0.001). Based on these results, a diagnostic model for sarcopenia was established with 83.3% sensitivity, 83.3% specificity, and 83.3% accuracy. CONCLUSIONS: In elderly people with type 2 diabetes, sarcopenia patients had smaller muscle CSA and less stiffness than non-sarcopenia patients. US and SWE might be useful to screen them. KEY POINTS: • Sarcopenia is common in elderly people with type 2 diabetes. • Ultrasound and shear wave elastography might be useful methods for quantitatively assessing muscle mass and strength. • Ultrasound and shear wave elastography might be useful methods for screening sarcopenia in elderly patients with type 2 diabetes.

The role of ultrasound in screening subclinical psoriatic arthritis in patients with moderate to severe psoriasis.

OBJECTIVE: To identify patients in the subclinical psoriatic arthritis (Sub-PsA) phase by ultrasound (US) and provide a solution to screen them. METHODS: A total of 490 participants with moderate-to-severe psoriasis were evaluated. Among them, 384 participants without arthritis symptoms were enrolled into the silent psoriasis group and 106 participants with arthritis symptoms, called prodromal/active PsA phase, were enrolled into the clinical PsA group. Another 80 non-psoriasis participants were enrolled into the control group. Each participant received clinical assessments and US examinations of 60 joints, 38 tendons, and 40 entheses. We compared the incidences of synovio-enthesitis, synovitis, tenosynovitis, erosion, and dactylitis detected on US among the three groups. Subsequently, on the basis of significant US findings, we distinguished Sub-PsA from psoriasis alone (PsO) in the silent psoriasis group and analyzed the clinical characteristics, mainly including basic clinical characteristics, body surface area (BSA), and Psoriasis Area and Severity Index (PASI) score. RESULTS: Only synovio-enthesitis significantly differed between the control group and the silent psoriasis group (1.3% vs. 16.1%, p < 0.001). The knee was the most commonly involved site of synovio-enthesitis (79.0%). Taking synovio-enthesitis as the standard, 16.1% of silent psoriasis participants and 12.7% of all psoriasis participants were in the Sub-PsA phase. Furthermore, there were no differences in BSA and PASI among the three phases of PsO, Sub-PsA, and prodromal/active PsA. CONCLUSIONS: Since the psoriasis patients in Sub-PsA phase was as high as 12.7% in all patients with moderate-to-severe psoriasis, US-detected synovio-enthesitis was recommended routinely for screening them regardless of arthritis symptoms, especially in the lower limbs. KEY POINTS: • Synovio-enthesitis on ultrasound was significantly associated with subclinical psoriatic arthritis, especially in the lower limbs. • Routine ultrasound evaluation could help screen psoriasis patients in the subclinical psoriatic arthritis phase, which was as high as 12.7% in all psoriasis patients.

The Value of Ultra-High-Frequency Ultrasound for the Differentiation between Superficial Basal Cell Carcinoma and Bowen's Disease.

BACKGROUND: The similar visual appearance of superficial basal cell carcinoma (sBCC) and Bowen's disease (BD) may cause confusion for diagnosis. OBJECTIVE: The aim of the study was to investigate the value of ultra-high-frequency ultrasound (uHFUS) in differentiating sBCC from BD. MATERIALS AND METHODS: This prospective study included a pilot cohort of 110 patients (73 BDs and 37 sBCCs) from November 2016 to October 2020 and a validation cohort of 42 patients (30 BDs and 12 sBCCs) from July 2021 to December 2021. Clinical and uHFUS features of pathologically confirmed sBCC and BD were assessed. A predictive model was developed based on the uHFUS features of the pilot cohort. Subsequently, the model was validated and compared with clinical diagnosis in the validation cohort. RESULTS: uHFUS features with significant differences between sBCC and BD included lesion surface, skin layer involvement, hyperkeratosis, and hyperechoic spots (all p < 0.05). A prediction model based on the above features was established to identify sBCC and BD in the pilot and validation cohorts with areas under the curve (AUC) of 0.908 and 0.923, sensitivity of 82.3% and 83.3%, specificity of 91.9% and 91.7%, and accuracy of 85.5% and 85.7%, respectively, which were significantly higher than those obtained by clinical diagnosis based on photographic pictures of lesions, with the AUC of 0.692, sensitivity of 63.3%, specificity of 75.3%, and accuracy of 66.7% (all p < 0.05). CONCLUSION: uHFUS provides detailed internal features of sBCC and BD, which facilitates the differentiation between sBCC and BD, and its diagnostic performance is superior to clinical diagnosis.

Diagnostic value of high-frequency ultrasound (HFUS) in evaluation of subcutaneous lesions.

BACKGROUND: It is unknown whether high-frequency ultrasound (HFUS) can evaluate invisible subcutaneous lesions. We aimed to investigate the diagnostic value of HFUS in invisible subcutaneous lesions. METHOD: Patients with invisible subcutaneous lesions were prospectively recruited from two centres. Before undergoing biopsy or surgery, each lesion was independently evaluated by two clinicians. One provides a clinical diagnosis by only clinical examination and the other provides an integrated diagnosis by combining clinical examination and HFUS information. Diagnoses were classified as correct, wrong, and indeterminate. A total of 391 lesions from 355 patients were enrolled, including 225 epidermoid cysts, 77 lipomas, 25 pilomatrixomas, 21 haemangiomas, 19 dermatofibromas, 11 dermatofibrosarcoma protuberans (DFSP), 7 neurofibromas, and 6 leiomyomas. Using pathological results as the gold standard, diagnostic performance was compared. RESULTS: The number of correct diagnoses increased from 185 (47.3%) by clinical examination alone to 316 (80.8%) after the addition of HFUS (P < 0.05). Meanwhile, the indeterminate diagnosis rate decreased from 143 (36.6%) to 10 (2.6%). Using HFUS, the accuracy improved significantly for epidermoid cysts (59.6% vs. 86.7%), lipomas (50.6% vs. 94.8%), pilomatrixomas (0% vs. 48.0%), haemangiomas (23.8% vs. 57.1%), and DFSPs (0% vs. 81.8%) (all p < 0.05). However, HFUS did not significantly improve the diagnostic accuracy of dermatofibromas (15.8% vs. 21.1%, p > 0.999), neurofibromas (42.9% vs. 71.4%, p = 0.625), or leiomyomas (16.7% vs. 100%, p = 0.063). CONCLUSION: Combining HFUS and clinical examination can generally improve the diagnostic accuracy and decrease the indeterminacy of invisible subcutaneous lesions, especially epidermoid cysts, lipomas, pilomatrixomas, haemangiomas, and DFSPs. However, for some rare lesions, HFUS cannot provide useful information.

The Value of Ultrasound for Differentiating Trichilemmal Cysts From Epidermoid Cysts.

OBJECTIVES: This study was aimed to evaluate the diagnostic performance of ultrasound (US) in differentiating trichilemmal cysts (TCs) from epidermoid cysts (ECs). METHODS: Based on clinical and ultrasound features, a prediction model was established and validated. 164 cysts in the pilot cohort and another 69 in the validation cohort diagnosed with TCs or ECs histopathologically were evaluated. The same radiologist performed all ultrasound examinations. RESULTS: For clinic features, TCs tended to occur in females compared with ECs (66.7 vs 28.5%; P < .001). In addition, TCs were prone to occur in the hairy area compared with ECs (77.8 vs 13.1%; P < .001). For ultrasound features, the internal hyperechogenicity and cystic change were more likely to appear in TCs in comparison with ECs (92.6 vs 25.5%; P < .001; 70.4 vs 23.4%; P < .001, respectively). Upon the features mentioned above, a prediction model was established with the areas under the receiver operating characteristic curves of 0.936 and 0.864 in the pilot and validation cohorts, respectively. CONCLUSIONS: US is promising for differentiating TCs from ECs and is valuable for their clinical management.

A versatile contribution of both aminopeptidases N and ABC transporters to Bt Cry1Ac toxicity in the diamondback moth.

BACKGROUND: Biopesticides and transgenic crops based on Bacillus thuringiensis (Bt) toxins are extensively used to control insect pests, but the rapid evolution of insect resistance seriously threatens their effectiveness. Bt resistance is often polygenic and complex. Mutations that confer resistance occur in midgut proteins that act as cell surface receptors for the toxin, and it is thought they facilitate its assembly as a membrane-damaging pore. However, the mechanistic details of the action of Bt toxins remain controversial. RESULTS: We have examined the contribution of two paralogous ABC transporters and two aminopeptidases N to Bt Cry1Ac toxicity in the diamondback moth, Plutella xylostella, using CRISPR/Cas9 to generate a series of homozygous polygenic knockout strains. A double-gene knockout strain, in which the two paralogous ABC transporters ABCC2 and ABCC3 were deleted, exhibited 4482-fold resistance to Cry1A toxin, significantly greater than that previously reported for single-gene knockouts and confirming the mutual functional redundancy of these ABC transporters in acting as toxin receptors in P. xylostella. A double-gene knockout strain in which APN1 and APN3a were deleted exhibited 1425-fold resistance to Cry1Ac toxin, providing the most direct evidence to date for these APN proteins acting as Cry1Ac toxin receptors, while also indicating their functional redundancy. Genetic crosses of the two double-gene knockouts yielded a hybrid strain in which all four receptor genes were deleted and this resulted in a > 34,000-fold resistance, indicating that while both types of receptor need to be present for the toxin to be fully effective, there is a level of functional redundancy between them. The highly resistant quadruple knockout strain was less fit than wild-type moths, but no fitness cost was detected in the double knockout strains. CONCLUSION: Our results provide direct evidence that APN1 and APN3a are important for Cry1Ac toxicity. They support our overarching hypothesis of a versatile mode of action of Bt toxins, which can compensate for the absence of individual receptors, and are consistent with an interplay among diverse midgut receptors in the toxins' mechanism of action in a super pest.

Construction and validation of a novel Ferroptosis-related gene signature predictive model in rectal Cancer.

BACKGROUND: Rectal cancer (RC) is one of the most common malignant tumors. Ferroptosis is an iron-dependent form of cell death, which plays an important role in various cancers. However, the correlation between ferroptosis-related genes (FRGs) and prognosis in RC remains unclear. METHODS: Gene expression data from The Cancer Genome Atlas Rectum adenocarcinoma (TCGA-READ) and GSE87211 were downloaded. Clustering and functional enrichment were evaluated. A FRGs risk score was established based on the univariate Cox analysis and the Least absolute shrinkage and selection operator (LASSO) analysis. K-M analysis and ROC analysis were conducted to determine prognostic values. qRT-PCR was performed to validate levels of mRNA expression. Multivariate Cox analysis was used to build a prognostic prediction model based on the risk score. RESULTS: Based on FRGs, RC patients were grouped into two clusters. In the functional enrichment of differentially expressed genes between the two clusters, immune-related pathways dominated. A novel FRGs signature with 14 genes related to the overall survival (OS) of RC was established. qRT-PCR of the 14 genes identified TP63, ISCU, PLIN4, MAP3K5, OXSR, FANCD2 and ATM were overexpressed in RC tissue; HSPB1, MAPK1, ABCC1, PANX1, MAPK9 and ATG7 were underexpressed; TUBE1 had no difference. The high-risk group had a significantly lower OS than the low-risk group (P < 0.001), and ROC curve analysis confirmed the signature's predictive capacity. Multivariate analysis demonstrated that the risk score and age were independent prognostic factors. CONCLUSION: A novel FRGs model can be used to predict the prognosis in RC, as well as to guide individual treatment.

Development of selective FGFR1 degraders using a Rapid synthesis of proteolysis targeting Chimera (Rapid-TAC) platform.

Proteolysis Targeting Chimera (PROTAC) has emerged as a novel therapeutic strategy. The major bottleneck for the development of PROTACs is the need to screen multiple parameters to create an effective degrader. It often involves the synthesis of dozens to hundreds of compounds one by one through a tedious process. We have developed a two-stage approach that allows for the rapid synthesis of PROTACs (Rapid-TAC) using preassembled building blocks to screen multiple parameters simultaneously. We herein report the application of this method to the development of PROTACs for FGFR, a challenging membrane protein target. In the first stage, we prepared 24 potential PROTACs quickly from a hydrazide-containing FGFR inhibitor and our previously reported VHL and CRBN ligand library bearing various linkers and an aldehyde functional group. These 24 PROTACs were then directly used for screening in cellular assay for protein degradation. Multiple hits were identified from the initial screening. We then prepared the corresponding stable analogues by replacing the hydrolytic labile acylhydrazone motif with an amide in the second stage. Among them, PROTAC LG1188 was identified as a potent and selective FGFR1 degrader.