CRISPR-Cas9/Cas12a systems for efficient genome editing and large genomic fragment deletions in Aspergillus niger.

CRISPR technology has revolutionized fungal genetic engineering by accelerating the pace and expanding the feasible scope of experiments in this field. Among various CRISPR-Cas systems, Cas9 and Cas12a are widely used in genetic and metabolic engineering. In filamentous fungi, both Cas9 and Cas12a have been utilized as CRISPR nucleases. In this work we first compared efficacies and types of genetic edits for CRISPR-Cas9 and -Cas12a systems at the polyketide synthase (albA) gene locus in Aspergillus niger. By employing a tRNA-based gRNA polycistronic cassette, both Cas9 and Cas12a have demonstrated equally remarkable editing efficacy. Cas12a showed potential superiority over Cas9 protein when one gRNA was used for targeting, achieving an editing efficiency of 86.5% compared to 31.7% for Cas9. Moreover, when employing two gRNAs for targeting, both systems achieved up to 100% editing efficiency for single gene editing. In addition, the CRISPR-Cas9 system has been reported to induce large genomic deletions in various species. However, its use for engineering large chromosomal segments deletions in filamentous fungi still requires optimization. Here, we engineered Cas9 and -Cas12a-induced large genomic fragment deletions by targeting various genomic regions of A. niger ranging from 3.5 kb to 40 kb. Our findings demonstrate that targeted engineering of large chromosomal segments can be achieved, with deletions of up to 69.1% efficiency. Furthermore, by targeting a secondary metabolite gene cluster, we show that fragments over 100 kb can be efficiently and specifically deleted using the CRISPR-Cas9 or -Cas12a system. Overall, in this paper, we present an efficient multi-gRNA genome editing system utilizing Cas9 or Cas12a that enables highly efficient targeted editing of genes and large chromosomal regions in A. niger.

Efficient cadmium-resistant plant growth-promoting bacteria loaded on pig bone biochar has higher efficiency in reducing cadmium phytoavailability and improving maize performance than on rice husk biochar.

Green agriculture faced challenges due to the shortage of efficient cadmium (Cd)-resistant plant growth-promoting bacteria (CdR-PGPB) and their low survival rate and activity during application. In this study, a diverse range of efficient CdR-PGPB were isolated from the rhizosphere soil of Desmodium elegans, especially those with high phosphate-solubilizing capabilities (272.87-450.45 mg L-1). Two highly efficient CdR-PGPB namely, XH1 and XH3 were loaded on to rice husk biochar (RHB) and pig bone biochar (PBB), labelled as RHBM and PBBM respectively. This study aimed to explore their effectiveness and mechanisms in promoting maize growth in a Cd-contaminated planting system. Results showed that PBBM performed best among all treatments. It significantly decreased soil phytoavailable Cd by 53.19 % and Cd content in maize shoot by 85.89 %. It also increased soil available phosphorus by 145.72 %, soil alkaline phosphatase activity by 76.34 %, maize shoot/root biomass by 47.06 %/67.98 %, Chlorophyll (a/b) content by 66.80 %/134.13 % and peroxidase activity by 171.96 %. These results were achieved through the synergistic action of efficient CdR-PGPB and PBB. Therefore, PBBM proved to be a promising and innovative application technique for sustainable agricultural development in Cd-contaminated farmland ecosystems.

KHSRP Stabilizes m6A-Modified Transcripts to Activate FAK Signaling and Promote Pancreatic Ductal Adenocarcinoma Progression.

N6-methyladenosine (m6A) is the most prevalent RNA modification and is associated with various biological processes. Proteins that function as readers and writers of m6A modifications have been shown to play critical roles in human malignancies. Here, we identified KH-type splicing regulatory protein (KHSRP) as an m6A binding protein that contributes to the progression of pancreatic ductal adenocarcinoma (PDAC). High KHSRP levels were detected in PDAC and predicted poor patient survival. KHSRP deficiency suppressed PDAC growth and metastasis in vivo. Mechanistically, KHSRP recognized and stabilized FAK pathway mRNAs, including MET, ITGAV and ITGB1, in an m6A-dependent manner, which led to activation of downstream FAK signaling that promoted PDAC progression. Targeting KHSRP with a PROTAC showed promising tumor suppressive effects in mouse models, leading to prolonged survival. Together, these findings indicate that KHSRP mediates FAK pathway activation in an m6A-dependent manner to support PDAC growth and metastasis, highlighting the potential of KHSRP as a therapeutic target in pancreatic cancer.

Clinical diagnostic value of methylated SEPT9 combined with NLR, PLR and LMR in colorectal cancer.

PURPOSE: This study aimed to investigate clinical diagnostic values of mSEPT9 combined with NLR, PLR and LMR in CRC. METHODS: 329 subjects composed of 120 CRC patients, 105 polyps patients and 104 healthy participants were prospectively recruited. Clinicopathologic features were collected and analyzed. Plasma samples were collected for mSEPT9, NLR, PLR and LMR test. The sensitivity, specificity and AUC of each biomarker separately or in combination were estimated by the ROC curve. RESULTS: The levels of NLR, PLR and the PDR of mSEPT9 in CRC patients were significantly higher than those in non-CRC subjects, while LMR was the opposite. The PDR of mSEPT9 in CRC patients was significantly correlated with age, tumor size, tumor stage and M stage. ROC curve analysis demonstrated moderate diagnostic values of mSEPT9, NLR, PLR and LMR in CRC patients with AUC of 0.78 (Se = 0.68, and Sp = 0.89), 0.78 (Se = 0.68, and Sp = 0.83), 0.80 (Se = 0.68, and Sp = 0.81), and 0.77 (Se = 0.72, and Sp = 0.73), respectively. Moreover, combination of these four biomarkers dramatically enhanced the diagnostic accuracy of CRC (AUC = 0.92, Se = 0.90, and Sp = 0.87), especially for CRC patients with large tumors (AUC = 0.95) or distal metastasis (AUC = 0.95). CONCLUSION: mSEPT9, NLR, PLR and LMR showed the potential to be reliable biomarkers for the diagnosis of CRC. And the combined application of these biomarkers further improved the diagnostic accuracy of CRC significantly.

Evaluating PAX1 methylation for cervical cancer screening triage in non-16/18 hrHPV-positive women.

BACKGROUND: In China, the national cervical cancer screening protocol involves initial testing for high-risk human papillomavirus (hrHPV), followed by cytology for hrHPV-positive cases. This study evaluates the effectiveness of PAX1 methylation (PAX1m) analysis in identifying precancerous or cancerous lesions in cervical samples from Chinese women positive for non-16/18 hrHPV strains. METHODS: Between February 2022 and March 2023, 281 cervical samples from non-16/18 hrHPV-positive women underwent cytological examination and PAX1m analysis. The study assessed the statistical relationship between PAX1m levels and the presence of cervical lesions, comparing the diagnostic performance of PAX1m to conventional cytology. RESULTS: A significant association was found between PAX1 methylation levels and the risk of CIN2 + and CIN3 + lesions, with 47 instances of CIN2 + detected. Odds ratios (ORs) for moderate and high PAX1m levels were 8.86 (95% CI: 2.24-42.17) and 166.32 (95% CI: 47.09-784.97), respectively. The area under the ROC curve for PAX1m in identifying CIN2 + lesions was 0.948 (95% CI: 0.895-0.99). PAX1m demonstrated similar sensitivity and negative predictive value (NPV) to cytology but reduced the colposcopy referral rate from 47.7% with cytology alone to 25.6% with PAX1m, showing superior specificity and positive predictive value across age groups. CONCLUSIONS: PAX1 methylation is a strong indicator of CIN2 + and CIN3 + risk, offering diagnostic performance comparable to cytology with the added benefit of reduced unnecessary colposcopy referrals. These findings support the use of PAX1m analysis as a reliable tool for triaging non-16/18 hrHPV-positive women in outpatient settings.

Benzo(a)pyrene promotes the malignant progression of malignant-transformed BEAS-2B cells by regulating YTH N6-methyladenosine RNA binding protein 1 to inhibit ferroptosis.

Benzo[a]pyrene (BaP) is associated with the development of lung cancer, but the underlying mechanism has not been completely clarified. Here, we used 10 µM BaP to induce malignant transformation of human bronchial epithelial BEAS-2B cells, named BEAS-2B-T. Results indicated that BaP (6.25, 12.5 and 25 µM) treatment significantly promoted the migration and invasion of BEAS-2B-T cells. Meanwhile, BaP exposure inhibited ferroptosis in BEAS-2B-T, ferroptosis-related indexes Fe2+, malondialdehyde (MDA), lipid peroxidation (LPO) and reactive oxygen species (ROS) decreased significantly. The protein level of ferroptosis-related molecule transferrin receptor (TFRC) decreased significantly, while solute carrier family 7 membrane 11 (SLC7A11), ferritin heavy chain 1 (FTH1) and glutathione peroxidase 4 (GPX4) increased significantly. The intervention of ferroptosis dramatically effected the migration and invasion of BEAS-2B-T induced by BaP. Furthermore, the expression of YTH N6-methyladenosine RNA binding protein 1 (YTHDF1) was markedly increased after BaP exposure. YTHDF1 knockdown inhibited BEAS-2B-T migration and invasion by promoting ferroptosis. In the meantime, the contents of Fe2+, MDA, LPO and ROS increased significantly, TFRC was markedly increased, and SLC7A11, FTH1, and GPX4 were markedly decreased. Moreover, overexpression of YTHDF1 promoted BEAS-2B-T migration and invasion by inhibiting ferroptosis. Importantly, knockdown of YTHDF1 promoted ferroptosis and reduced BEAS-2B-T migration and invasion during BaP exposure, and overexpression of YTHDF1 increased migration and invasion of BEAS-2B-T by inhibiting ferroptosis during BaP exposure. RNA immunoprecipitation assays indicated that the binding of YTHDF1 to SLC7A11 and FTH1 markedly increased after YTHDF1 overexpression. Therefore, we concluded that BaP promotes the malignant progression of BEAS-2B-T cells through YTHDF1 upregulating SLC7A11 and FTH1 to inhibit ferroptosis. This study reveals new epigenetic and ferroptosis markers for preventing and treating lung cancer induced by environmental carcinogens.

Cayley Rotation Averaging: Multiple Camera Averaging Under the Cayley Framework.

Rotation averaging, which aims to calculate the absolute rotations of a set of cameras from a redundant set of their relative rotations, is an important and challenging topic arising in the study of structure from motion. A central problem in rotation averaging is how to alleviate the influence of noise and outliers. Addressing this problem, we investigate rotation averaging under the Cayley framework in this paper, inspired by the extra-constraint-free nature of the Cayley rotation representation. Firstly, for the relative rotation of an arbitrary pair of cameras regardless of whether it is corrupted by noise/outliers or not, a general Cayley rotation constraint equation is derived for reflecting the relationship between this relative rotation and the absolute rotations of the two cameras, according to the Cayley rotation representation. Then based on such a set of Cayley rotation constraint equations, a Cayley-based approach for Rotation Averaging is proposed, called CRA, where an adaptive regularizer is designed for further alleviating the influence of outliers. Finally, a unified iterative algorithm for minimizing some commonly-used loss functions is proposed under this approach. Experimental results on 16 real-world datasets and multiple synthetic datasets demonstrate that the proposed CRA approach achieves a better accuracy in comparison to several typical rotation averaging approaches in most cases.

Carexqingyuanensis (Cyperaceae), a new species from Guangdong, China.

Carexqingyuanensis, a new species of Cyperaceae from Guangdong Province, China, is described and illustrated. The new species is morphologically similar to Carexpeliosanthifolia F. T. Wang & Tang ex P. C. Li, but it can be distinguished by the racemose inflorescence branches appearing single (rarely binate or ternate) (vs. binate or ternate), one (rarely two or three) (vs. 1-3) spiked, male part of linear-cylindrical spikes much longer than the female part (vs. just male part short-cylindrical and slightly longer than female part), style base thickened (vs. not thickened) and perigynium horizontally patent with a short (vs. long and excurved) beak. Phylogenetic analysis, based on the two nuclear DNA regions (ETS 1f and ITS) and three chloroplast DNA regions (matK, ndhF and rps16), suggests that the new species belongs to sect. Siderostictaes.s. of subg. Siderosticta and shows a closer phylogenetic relationship to Carexscaposa C. B. Clarke.

QDPR deficiency drives immune suppression in pancreatic cancer.

The relevance of biopterin metabolism in resistance to immune checkpoint blockade (ICB) therapy remains unknown. We demonstrate that the deficiency of quinoid dihydropteridine reductase (QDPR), a critical enzyme regulating biopterin metabolism, causes metabolite dihydrobiopterin (BH2) accumulation and decreases the ratio of tetrahydrobiopterin (BH4) to BH2 in pancreatic ductal adenocarcinomas (PDACs). The reduced BH4/BH2 ratio leads to an increase in reactive oxygen species (ROS) generation and a decrease in the distribution of H3K27me3 at CXCL1 promoter. Consequently, myeloid-derived suppressor cells are recruited to tumor microenvironment via CXCR2 causing resistance to ICB therapy. We discovered that BH4 supplementation is capable to restore the BH4/BH2 ratio, enhance anti-tumor immunity, and overcome ICB resistance in QDPR-deficient PDACs. Tumors with lower QDPR expression show decreased responsiveness to ICB therapy. These findings offer a novel strategy for selecting patient and combining therapies to improve the effectiveness of ICB therapy in PDAC.

Giving birth after fertility-sparing treatment for primary leiomyosarcoma of the fallopian tube.

Primary leiomyosarcoma of the fallopian tube (PLFT) is an extremely rare gynecological malignancy that has only been described in case reports. Fertility-sparing treatment for PLFT has not been reported previously. A 24-year-old nulligravida woman was diagnosed with stage IC1 PLFT in the right fallopian tube after experiencing right lower quadrant pain for 2 weeks. She underwent laparoscopic right salpingectomy to preserve fertility followed by adjuvant chemotherapy with gemcitabine/docetaxel. She subsequently became pregnant spontaneously, delivering a term baby 27 months after treatment. This appears to be the only report of the use of fertility-preserving treatment for PLFT. The success of the treatment provides valuable information on the preservation of fertility in young women with PLFT.

SIRT3 alleviates high glucose-induced chondrocyte injury through the promotion of autophagy and suppression of apoptosis in osteoarthritis progression.

A growing amount of epidemiological evidence proposes diabetes mellitus (DM) to be an independent risk factor for osteoarthritis (OA). Sirtuin 3 (SIRT3), which is mainly located in mitochondria, belongs to the family of nicotinamide adenine dinucleotide (NAD+)-dependent protein deacetylases and is involved in the physiological and pathological processes of cell regulation. The aim of this study was to investigate the effects of SIRT3 on diabetic OA and underlying mechanisms in the prevention of type 2 DM (T2DM)-induced articular cartilage damage. High-fat and high-sugar diets combined with streptozotocin (STZ) injection were used for establishing an experimental T2DM rat model. The destabilization of medial meniscus (DMM) surgery was applied to induce the rat OA model. Primary rat chondrocytes were cultivated with a concentration of gradient glucose. Treatment with intra-articular injection of SIRT3 overexpression lentivirus was achieved in vivo, and intervention with SIRT3 knockdown was performed using siRNA transfection in vitro. High glucose content was found to activate inflammatory response, facilitate apoptosis, downregulate autophagy, and exacerbate mitochondrial dysfunction in a dose-dependent manner in rat chondrocytes, which can be deteriorated by SIRT3 knockdown. In addition, articular cartilage damage was found to be more severe in T2DM-OA rats than in DMM-induced OA rats, which can be mitigated by the intra-articular injection of SIRT3 overexpression lentivirus. Targeting SIRT3 is a potential therapeutic strategy for the alleviation of diabetic OA.

Eosinophilic Cystitis Mimicking Malignancy on FDG PET/CT.

ABSTRACT: A 64-year-old man had progressive dysuria and nocturia for 1 month. Initial MRI and CT revealed localized thickening of the bladder wall with significant enhancement. Meanwhile, the lesion showed intense FDG accumulation on the delayed PET/CT. Taken together, a malignancy was suspected. However, the pathologic findings confirmed the diagnosis of eosinophilic cystitis.

Characterization of Firmiana danxiaensis plastomes and comparative analysis of Firmiana: insight into its phylogeny and evolution.

BACKGROUND: Firmiana danxiaensis is a critically endangered and ecologically important tree currently only found in four locations in Danxia or Karst habitats in northern Guangdong Province, China. The specialized habitat preference makes it an ideal model species for study of adaptive evolution. Meanwhile, the phylogenetic relationships of F. danxiaensis in four locations under two landforms are unclear. Therefore, we sequenced its complete chloroplast (cp.) genomes and conducted comprehensive interspecific and intrageneric plastome studies. RESULTS: The F. danxiaensis plastomes in four locations showed a typical quadripartite and circular structure that ranged from 160,832 to 161,206 bp in size, with 112 unique genes encoded. Comparative genomics showed that the plastomes of F. danxiaensis were relatively conserved with high similarity of genome organization, gene number, GC content and SSRs. While the genomes revealed higher biased codon preferences in Karst habitat than those in Danxia habitats. Eighteen and 11 divergent hotpots were identified at interspecific and intrageneric levels for species identification and further phylogenetic studies. Seven genes (clpP, accD, ccsA, ndhH, rpl20, rpoC2, and rps4) were under positive selection and may be related to adaptation. Phylogenetic analysis revealed that F. danxiaensis is sister to F. major and F. simplex. However, the interspecific relationships are not consistent with the habitat types. CONCLUSIONS: The characteristics and interspecific relationship of F. danxiaensis plastomes provide new insights into further integration of geographical factors, environmental factors, and genetic variations on the genomic study of F. danxiaensis. Together, our study will contribute to the study of species identification, population genetics, and conservation biology of F. danxiaensis.

Simple and Effective Squash-PCR for Rapid Genotyping of Industrial Microalgae.

Microalgae are recognized for their versatility in providing renewable energy, biopharmaceuticals, and nutraceuticals, attributed to their sustainable, renewable, and cost-effective nature. Genetic engineering has proven highly effective in enhancing microalgae production. PCR-based genotyping is the primary method for screening genetically transformed microalgae cells. Recently, we developed a novel PCR method, namely Squash-PCR, and employed it for the molecular analysis of industrially important fungi and yeasts. In this study, we successfully implemented the Squash-PCR technique in 12 industrially significant algae species. This approach offers a quick and reliable means of obtaining DNA templates directly from squashed algal cells, eliminating the need for time-consuming and labor-intensive cultivation and genomic DNA extraction steps. Our results demonstrate the effectiveness of Squash-PCR in detecting and characterizing target genes of interest in 12 different algae species. Overall, this study establishes the Squash-PCR method as a valuable tool for molecular studies in algae, enabling researchers to rapidly screen and manipulate genetic traits in diverse algal species.

Superficial radiotherapy (SRT-100) for refractory plantar warts: An alternative noninvasive treatment strategy.

BACKGROUND: Verrucas that occur on the soles of the feet are called plantar warts, most of which can recur repeatedly and are difficult to eradicate. Hypertrophic and refractory plantar warts are often accompanied by pain and discomfort, which cause many inconveniences in patients' daily lives. AIM: This study aimed to analyze the therapeutic effect of superficial radiotherapy (SRT-100) on refractory plantar warts and further create favorable conditions for the subsequent treatment of this disease with a high recurrence rate. METHODS: A retrospective analysis was conducted for refractory plantar warts treated with superficial radiotherapy in our outpatient department from January to June 2023. RESULTS: A total of 30 patients were included in our study (median age, 33 years). The female-to-male ratio was 1:3.29. Two to six months after radiotherapy, all of the warts subsided in 23 (76.67%) patients, most of the warts subsided in 4 (13.33%) patients, 3 (10%) patients did not respond to treatment, and 7 (23.33%) patients had recurrent or new warts after their warts subsided. CONCLUSIONS: Most patients with refractory plantar warts improved after superficial radiotherapy. Our study presented that men are more susceptible to plantar warts than women, and young and middle-aged people are the main population affected by the disease. Superficial radiotherapy is an effective treatment for refractory plantar warts, which can quickly remove the warts in a short period. It is safe and noninvasive, with minimal adverse reactions. Some patients relapse after the lesion is clear, and superficial radiotherapy can create favorable conditions for the subsequent treatment of viral warts in clinical practice.

Photocatalytic CO2 reduction of 2D/0D CoAl-LDH@Cu2O catalyst with p-n heterojunction.

Layered double hydroxides (LDHs) are widely used in catalytic field, especially in photocatalysis, benefiting from the ultrathin 2D structure and luxuriant surface functional groups. However, the wide band gap and low utilization rate of solar spectrum affect their photocatalytic performance. Herein, we integrated n-type CoAl-LDH with p-type Cu2O nanoparticles to construct a p-n heterojunction with a strong built-in electric field, which can prevent photoinduced electron-hole pairs from recombination as well as facilitate charge transfer. With the X-ray photoelectron spectroscope and in situ Fourier transform infrared spectroscopy, we confirmed the charge transfer under light illumination complying with the type II-scheme mechanism and analyzed the intermediates during photocatalytic CO2 reduction reaction (CO2RR). The highest yields reached 320.9 µmol h-1 g-1 for CoAl-LDH@Cu2O-60 (LC-60) under 1 h light irradiation, which was about 1.6 times than the pristine CoAl-LDH. The sample also exhibited excellent stability which maintained 84.1% of initial performance after 4 circulations.

Latest progress in low-intensity pulsed ultrasound for studying exosomes derived from stem/progenitor cells.

Stem cells have self-renewal, replication, and multidirectional differentiation potential, while progenitor cells are undifferentiated, pluripotent or specialized stem cells. Stem/progenitor cells secrete various factors, such as cytokines, exosomes, non-coding RNAs, and proteins, and have a wide range of applications in regenerative medicine. However, therapies based on stem cells and their secreted exosomes present limitations, such as insufficient source materials, mature differentiation, and low transplantation success rates, and methods addressing these problems are urgently required. Ultrasound is gaining increasing attention as an emerging technology. Low-intensity pulsed ultrasound (LIPUS) has mechanical, thermal, and cavitation effects and produces vibrational stimuli that can lead to a series of biochemical changes in organs, tissues, and cells, such as the release of extracellular bodies, cytokines, and other signals. These changes can alter the cellular microenvironment and affect biological behaviors, such as cell differentiation and proliferation. Here, we discuss the effects of LIPUS on the biological functions of stem/progenitor cells, exosomes, and non-coding RNAs, alterations involved in related pathways, various emerging applications, and future perspectives. We review the roles and mechanisms of LIPUS in stem/progenitor cells and exosomes with the aim of providing a deeper understanding of LIPUS and promoting research and development in this field.

18F-FDG PET/CT image of pericardial inflammatory myofibroblastic tumor.

Pericardial inflammatory myofibroblastic tumor (IMT) is very rare. Herein, we report fluorine-18-fluorodeoxyglucose (18F-FDG) positron emission tomography/computed tomography (PET/CT) findings of pericardial IMT in a 57-year-old woman. On conventional image, it presented as a pericardial mass with heterogeneous delay enhancement. On 18F-FDG PET/CT image, this lesion had mild 18F-FDG uptake with a maximum standardized uptake value (SUVmax) of 1.84. The postoperative pathology supported a diagnosis of IMT. Our case hints us that IMT should be regarded as a differential diagnosis when we meet a solitary pericardial mass with 18F-FDG uptake.

Super-enhancer RNA m6A promotes local chromatin accessibility and oncogene transcription in pancreatic ductal adenocarcinoma.

The biological functions of noncoding RNA N6-methyladenosine (m6A) modification remain poorly understood. In the present study, we depict the landscape of super-enhancer RNA (seRNA) m6A modification in pancreatic ductal adenocarcinoma (PDAC) and reveal a regulatory axis of m6A seRNA, H3K4me3 modification, chromatin accessibility and oncogene transcription. We demonstrate the cofilin family protein CFL1, overexpressed in PDAC, as a METTL3 cofactor that helps seRNA m6A methylation formation. The increased seRNA m6As are recognized by the reader YTHDC2, which recruits H3K4 methyltransferase MLL1 to promote H3K4me3 modification cotranscriptionally. Super-enhancers with a high level of H3K4me3 augment chromatin accessibility and facilitate oncogene transcription. Collectively, these results shed light on a CFL1-METTL3-seRNA m6A-YTHDC2/MLL1 axis that plays a role in the epigenetic regulation of local chromatin state and gene expression, which strengthens our knowledge about the functions of super-enhancers and their transcripts.