Deoxypodophyllotoxin Mediates Autophagy Death through Inhibition of GRP78 in Human Osteosarcoma.

BACKGROUND: Glucose-regulated protein 78 (GRP78), as a chaperone protein, can protect the endoplasmic reticulum of cells and is expressed to influence chemoresistance and prognosis in cancer. Deoxypodophyllotoxin (DPT) is a compound with antitumor effects on cancers. DPT inhibits the proliferation of osteosarcoma by inducing apoptosis, necrosis, or cell cycle arrest. OBJECT: This study was performed to demonstrate the molecular mechanism by which DPT attenuates osteosarcoma progression through GRP78. METHODS: Natural compound libraries and western blot (WB) were used to screen the inhibitors of osteosarcoma GRP78. The expression of mitochondria-related genes in cancer cells of the treatment group was detected by quantitative real-time PCR (qPCR) and WB. 3-(4,5)- Dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) and 5-ethynyl-2'- deoxyuridine (EDU) were used to discover the activity and proliferation of osteosarcoma cells treated with DPT. We constructed an in vivo mouse model of DPT drug therapy and carried out immunohistochemical detection of xenografts. The treated osteosarcoma cells were analyzed using bioinformatics and electron microscopy. The data were analyzed finally. RESULTS: DPT inhibited osteosarcoma cell survival and the growth of tumor xenografts. It promoted up-regulation of BCL2-associated X protein (Bax) and B-cell CLL/lymphoma 2 (Bcl-2), which serves to mediate and attenuate, respectively, the killing activities of DPT through mitochondria dysfunction. The effect of DPT against cancer cells could be attenuated by the overexpression of GRP78, characterized by the inactivation of the caspase cascade. The loss of GRP78 in osteosarcoma cells negatively mediated the basal level of autophagyassociated genes. DPT stimulated autophagy via the phosphoinositide 3-kinase (PI3K)-v-akt murine thymoma viral oncogene homolog (AKT), a mechanistic target of rapamycin (mTOR) axis. The autophagy caused by DPT played an active role in the osteosarcoma of humans and blocked the apoptotic cascade. CONCLUSION: Combination treatment with the GRP78 inhibitor DPT and pharmacological autophagy inhibitors will be a meaningful method of obviating osteosarcoma cells.

Angiopoietin-4 expression and potential mechanisms in carcinogenesis: Current achievements and perspectives.

As one of the factors regulating tumour angiogenesis, angiopoietin-4 (ANGPT4), which plays an important role in promoting tumour proliferation, survival, expansion and angiogenesis, is highly expressed in some tumours, such as lung adenocarcinoma, glioblastoma and ovarian cancer. This may be related to the fact that ANGPT4 affects the blood vessels and lymphatic system of the tumour. Specifically, ANGPT4 could play an effective role in promoting cancer by affecting the tyrosine kinase receptor TIE2, ERK1/2 and PI3K/AKT signalling pathways. Therefore, ANGPT4 may be an important biomarker for the occurrence and development of cancer and poor prognosis. In addition, the inhibition of ANGPT4 may be a useful cancer treatment. This paper reviews the latest preclinical research on ANGPT4, emphasizes its role in tumourigenesis and broadens our understanding of the carcinogenic function of ANGPT4 and the development of ANGPT4 inhibitors. This is the latest version of the revised version of the previous article.

PLK1-mediated phosphorylation cascade activates Mis18 complex to ensure centromere inheritance.

Accurate chromosome segregation requires the attachment of microtubules to centromeres, epigenetically defined by the enrichment of CENP-A nucleosomes. During DNA replication, CENP-A nucleosomes undergo dilution. To preserve centromere identity, correct amounts of CENP-A must be restored in a cell cycle-controlled manner orchestrated by the Mis18 complex (Mis18α-Mis18ß-Mis18BP1). We demonstrate here that PLK1 interacts with the Mis18 complex by recognizing self-primed phosphorylations of Mis18α (Ser54) and Mis18BP1 (Thr78 and Ser93) through its Polo-box domain. Disrupting these phosphorylations perturbed both centromere recruitment of the CENP-A chaperone HJURP and new CENP-A loading. Biochemical and functional analyses showed that phosphorylation of Mis18α and PLK1 binding were required to activate Mis18α-Mis18ß and promote Mis18 complex-HJURP interaction. Thus, our study reveals key molecular events underpinning the licensing role of PLK1 in ensuring accurate centromere inheritance.

Two-factor authentication underpins the precision of the piRNA pathway.

The PIWI-interacting RNA (piRNA) pathway guides the DNA methylation of young, active transposons during germline development in male mice1. piRNAs tether the PIWI protein MIWI2 (PIWIL4) to the nascent transposon transcript, resulting in DNA methylation through SPOCD1 (refs. 2-5). Transposon methylation requires great precision: every copy needs to be methylated but off-target methylation must be avoided. However, the underlying mechanisms that ensure this precision remain unknown. Here, we show that SPOCD1 interacts directly with SPIN1 (SPINDLIN1), a chromatin reader that primarily binds to H3K4me3-K9me3 (ref. 6). The prevailing assumption is that all the molecular events required for piRNA-directed DNA methylation occur after the engagement of MIWI2. We find that SPIN1 expression precedes that of both SPOCD1 and MIWI2. Furthermore, we demonstrate that young LINE1 copies, but not old ones, are marked by H3K4me3, H3K9me3 and SPIN1 before the initiation of piRNA-directed DNA methylation. We generated a Spocd1 separation-of-function allele in the mouse that encodes a SPOCD1 variant that no longer interacts with SPIN1. We found that the interaction between SPOCD1 and SPIN1 is essential for spermatogenesis and piRNA-directed DNA methylation of young LINE1 elements. We propose that piRNA-directed LINE1 DNA methylation requires a developmentally timed two-factor authentication process. The first authentication is the recruitment of SPIN1-SPOCD1 to the young LINE1 promoter, and the second is MIWI2 engagement with the nascent transcript. In summary, independent authentication events underpin the precision of piRNA-directed LINE1 DNA methylation.

The study of vaginal wall thickness in adults based on histopathological measurements.

To accurately measure the vaginal mucosa thickness across different age groups using histopathologic techniques and investigate the factors that may influence the thickness changes. This study aims to provide clinicians with objective evidence of variations in vaginal mucosal thickness, facilitating personalized medical decisions for patients. A retrospective analysis was conducted on clinical data from 348 patients who underwent local vaginal wall resection at the West China Second University Hospital, Sichuan University, from January 2021 and May 2022. The thickness of vaginal mucosa, epithelium and lamina propria was measured precisely under the microscope. And the 10th, 25th, 50th, 75th, and 90th percentile values of vaginal mucosa thickness across different age groups were counted and charted a dot-line plot. The percentile values for vaginal mucosa thickness exhibited a decreasing trend with increasing age; vaginal mucosa thickness showed significant correlations with times of delivery (P = 0.031) and age (P < 0.001), both of which were negatively associated. And vaginal mucosa thickness demonstrated no significant correlation with body mass index (BMI) (P = 0.325), times of abortions (P = 0.511), times of gestation (P = 0.101), menstrual cycle (P = 0.533), or types of delivery (P = 0.056); epithelial thickness showed significant associations with age (P < 0.001) and types of delivery (P = 0.017), both of which were negative correlations. Moreover, BMI (P = 0.429), times of abortions (P = 0.764), delivery (P = 0.079), gestation (P = 0.475), and menstrual cycle (P = 0.950) were nonassociated with epithelial thickness; lamina propria thickness displayed a significant correlation only with age (P = 0.002), and there were no obvious correlations observed between lamina propria thickness and BMI (P = 0.374), times of abortion (P = 0.417), delivery (P = 0.053), gestation (P = 0.101), types of delivery (P = 0.132) and menstrual cycle (P = 0.495). Moreover, when the age segmentation was thresholded at 35 and 50 years, both epithelial thickness and vaginal mucosa thickness were significantly correlated with age (P < 0.05). Lamina propria thickness was associated with age when the age threshold was set at 35 years (P = 0.007), whereas it showed no strong link with age when the age threshold was 50 years (P = 0.072). This study has innovatively established percentile reference values for vaginal mucosa thickness based on histopathology, furnishing clinicians with objective evidence of variations in vaginal mucosal thickness to facilitate personalized medical decisions for patients. The findings demonstrated a strong link between vaginal mucosa thickness and age, with epithelium likely playing a predominant role, while the association with lamina propria appeared to be less significant. Further research involving a larger sample size is warranted to elucidate the potential relationship with the lamina propria.

Oleanane triterpenoids with C-14 carboxyl group from Astilbe grandis inhibited LPS-induced macrophages activation by suppressing the NF-κB signaling pathway.

Background: Excessive inflammation poses significant risks to human physical and mental health. Astilbe grandis, a traditional Miao medicine, is renowned for its anti-inflammatory properties. However, the specific anti-inflammatory effects and mechanisms of many compounds within this plant remain unclear. This study aims to investigate the anti-inflammatory effects and mechanisms of two characteristic oleanane triterpenoids, 3α-acetoxyolean-12-en-27-oic acid (1) and 3ß-acetoxyolean-12-en-27-oic acid (2), isolated from Astilbe grandis, using lipopolysaccharide (LPS)-induced Macrophages. Methods: The anti-inflammatory effects and mechanisms of compounds 1 and 2 were investigated by establishing an LPS-induced inflammation model in RAW 264.7 cells and THP-1 cells. Nitric oxide (NO) levels were assessed using the Griess method. The concentrations of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and interleukin-1beta (IL-1ß) were measured via enzyme-linked immunosorbent assay (ELISA). The expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) was determined using western blotting and quantitative real-time PCR (qRT-PCR). Additionally, the phosphorylation level of p65 in nuclear factor-kappa B (NF-κB) was assessed through western blotting. The nuclear translocation of NF-κB p65 was assessed through immunofluorescence staining. Finally, the binding affinity of the compounds to NF-κB p65 target was validated through molecular docking. Results: Compounds 1 and 2 significantly inhibited the expression of NO, TNF-α, IL-6, IL-1ß, COX-2, and iNOS in LPS-induced Macrophages. Mechanistically, they attenuated the activation of the NF-κB signaling pathway by downregulating the phosphorylation level and nuclear translocation of p65. Conclusion: This study elucidates the anti-inflammatory activities and potential mechanism of the characteristic oleanane triterpenoids with C-14 carboxyl group, compounds 1 and 2, in LPS-induced Macrophages by inhibiting the NF-κB signaling pathway for the first time. These findings suggest that these two compounds hold promise as potential candidates for anti-inflammatory interventions in the future.

RAB37-mediated autophagy guards ovarian homeostasis and function.

Loss of ovarian homeostasis is associated with ovary dysfunction and female diseases; however, the underlying mechanisms responsible for the establishment of homeostasis and its function in the ovary have not been fully elucidated. Here, we showed that conditional knockout of Rab37 in oocytes impaired macroautophagy/autophagy proficiency in the ovary and interfered with follicular homeostasis and ovary development in mice. Flunarizine treatment upregulated autophagy, thus rescuing the impairment of follicular homeostasis and ovarian dysfunction in rab37 knockout mice by reprogramming of homeostasis. Notably, both the E2F1 and EGR2 transcription factors synergistically activated Rab37 transcription and promoted autophagy. Thus, RAB37-mediated autophagy ensures ovary function by maintaining ovarian homeostasis.Abbreviations: AMH: anti-Mullerian hormone; ATG: autophagy related; BECN1: beclin 1; cKO: conditional knockout; Cre: cyclization recombination enzyme; dpp: days postpartum; E2: estradiol; E2F1: E2F transcription factor 1; EBF1: EBF transcription factor 1; EGR2: early growth response 2; FSH: follicle stimulating hormone; LH: luteinizing hormone; mpp: months postpartum; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; RAB37: RAB37, member RAS oncogene family; SQSTM1: sequestosome 1; TFEB: transcription factor EB; Zp3: zona pellucida glycoprotein 3.

Synthesis, biological activities and mechanistic studies of C20-ketone pachysandra alkaloids as anti-hepatocellular carcinoma agents.

The pachysandra alkaloids found in Sarcococca ruscifolia demonstrate notable anti-hepatocellular carcinoma activity. Despite their efficacy, the structural diversity of these compounds remains limited, and their precise antitumor mechanism is still unclear. In pursuit of identifying novel lead compounds with high efficacy and low toxicity for combating hepatocellular carcinoma, twenty-three compounds of C20-ketone pachysandra alkaloid derivatives were designed and synthesized by using 3-dimethylamine pachysandra alkaloids as scaffolds. Subsequent in vitro anticancer activity experiments showed that synthetic pachysandra alkaloids had a stronger effect on HepG2 cells than did their natural counterparts, with low toxicity and high selectivity. The most potent derivative, 6k, had an IC50 value of 0.75 µM, demonstrating 25.7-fold greater anticancer activity than sarcovagine D against HepG2 cells. Through network pharmacology and molecular docking analysis, it was revealed that synthetic pachysandra alkaloids may exert their effects by inhibiting the JAK2/STAT3 pathway, thereby preventing the proliferation of liver cancer cells. Further research through scratch tests, immunofluorescence experiments, and Western blot analysis revealed that compound 6k effectively inhibited the migration of HepG2 cells and induced mitochondria-mediated intrinsic apoptosis of HepG2 cells by regulating the JAK2/STAT3 signaling pathway. The aforementioned results indicate that compound 6k could be developed as a potential candidate for the treatment of hepatocellular carcinoma.

Generalizable Reconstruction for Accelerating MR Imaging via Federated Learning with Neural Architecture Search.

Heterogeneous data captured by different scanning devices and imaging protocols can affect the generalization performance of the deep learning magnetic resonance (MR) reconstruction model. While a centralized training model is effective in mitigating this problem, it raises concerns about privacy protection. Federated learning is a distributed training paradigm that can utilize multi-institutional data for collaborative training without sharing data. However, existing federated learning MR image reconstruction methods rely on models designed manually by experts, which are complex and computationally expensive, suffering from performance degradation when facing heterogeneous data distributions. In addition, these methods give inadequate consideration to fairness issues, namely ensuring that the model's training does not introduce bias towards any specific dataset's distribution. To this end, this paper proposes a generalizable federated neural architecture search framework for accelerating MR imaging (GAutoMRI). Specifically, automatic neural architecture search is investigated for effective and efficient neural network representation learning of MR images from different centers. Furthermore, we design a fairness adjustment approach that can enable the model to learn features fairly from inconsistent distributions of different devices and centers, and thus facilitate the model to generalize well to the unseen center. Extensive experiments show that our proposed GAutoMRI has better performances and generalization ability compared with seven state-of-the-art federated learning methods. Moreover, the GAutoMRI model is significantly more lightweight, making it an efficient choice for MR image reconstruction tasks. The code will be made available at https://github.com/ternencewu123/GAutoMRI.

Model-based federated learning for accurate MR image reconstruction from undersampled k-space data.

Deep learning-based methods have achieved encouraging performances in the field of Magnetic Resonance (MR) image reconstruction. Nevertheless, building powerful and robust deep learning models requires collecting large and diverse datasets from multiple centers. This raises concerns about ethics and data privacy. Recently, federated learning has emerged as a promising solution, enabling the utilization of multi-center data without the need for data transfer between institutions. Despite its potential, existing federated learning methods face challenges due to the high heterogeneity of data from different centers. Aggregation methods based on simple averaging, which are commonly used to combine the client's information, have shown limited reconstruction and generalization capabilities. In this paper, we propose a Model-based Federated learning framework (ModFed) to address these challenges. ModFed has three major contributions: (1) Different from existing data-driven federated learning methods, ModFed designs attention-assisted model-based neural networks that can alleviate the need for large amounts of data on each client; (2) To address the data heterogeneity issue, ModFed proposes an adaptive dynamic aggregation scheme, which can improve the generalization capability and robustness of the trained neural network models; (3) ModFed incorporates a spatial Laplacian attention mechanism and a personalized client-side loss regularization to capture the detailed information for accurate image reconstruction. The effectiveness of the proposed ModFed is evaluated on three in-vivo datasets. Experimental results show that when compared to six existing state-of-the-art federated learning approaches, ModFed achieves better MR image reconstruction performance with increased generalization capability. Codes will be made available at https://github.com/ternencewu123/ModFed.

Structural basis for Mis18 complex assembly and its implications for centromere maintenance.

The centromere, defined by the enrichment of CENP-A (a Histone H3 variant) containing nucleosomes, is a specialised chromosomal locus that acts as a microtubule attachment site. To preserve centromere identity, CENP-A levels must be maintained through active CENP-A loading during the cell cycle. A central player mediating this process is the Mis18 complex (Mis18α, Mis18ß and Mis18BP1), which recruits the CENP-A-specific chaperone HJURP to centromeres for CENP-A deposition. Here, using a multi-pronged approach, we characterise the structure of the Mis18 complex and show that multiple hetero- and homo-oligomeric interfaces facilitate the hetero-octameric Mis18 complex assembly composed of 4 Mis18α, 2 Mis18ß and 2 Mis18BP1. Evaluation of structure-guided/separation-of-function mutants reveals structural determinants essential for cell cycle controlled Mis18 complex assembly and centromere maintenance. Our results provide new mechanistic insights on centromere maintenance, highlighting that while Mis18α can associate with centromeres and deposit CENP-A independently of Mis18ß, the latter is indispensable for the optimal level of CENP-A loading required for preserving the centromere identity.

Aberrant angiogenic signaling pathways: Accomplices in ovarian cancer progression and treatment.

Ovarian cancer is one of the most common malignant tumors in women, and treatment options are limited. Despite efforts to adjust cancer treatment models and develop new methods, including tumor microenvironment (TME) therapy, more theoretical support is needed. Increasing attention is being given to antiangiogenic measures for TME treatment. Another important concept in ovarian cancer TME is angiogenesis, where tumor cells obtain nutrients and oxygen from surrounding tissues through blood vessels to support further expansion and metastasis. Many neovascularization signaling pathways become imbalanced and hyperactive during this process. Inhibiting these abnormal pathways can yield ideal therapeutic effects in patients, even by reversing drug resistance. However, these deep TME signaling pathways often exhibit crosstalk and correlation. Understanding these interactions may be an important strategy for further treating ovarian cancer. This review summarizes the latest progress and therapeutic strategies for these angiogenic signaling pathways in ovarian cancer.

Abietane diterpenoids from Isodon amethystoides and their biological activities.

Seven undescribed abietane diterpenoids [abietamethinols A-G (1-7)] were isolated from the twigs and leaves of Isodon amethystoides. Their structures were elucidated on the basis of spectroscopic methods including 2D NMR, and they were further confirmed by X-ray crystallographic data. Lophanic acid was considered as the precursor of 1-7 in the biosynthesis pathway hypothesis. These compounds were evaluated for their cytotoxic, anti-bacterial and anti-AIV (avian influenza virus) activities. Compound 5 showed 42.9% inhibitory activity against the cancer cell line SMMC-7721 at the concentration of 40 µM, 3 and 4 could inhibit the bacterial growth of Streptococcus sobrinus by 55.3% and 63.2% at the concentrations of 148.6 and 141.9 µM, respectively, and 4 was demonstrated with antiviral activity against AIV with the inhibitory effect of 68.4% at 25 µM.

The C-terminal disordered loop domain of Apc8 unlocks APC/C mitotic activation.

The anaphase-promoting complex/cyclosome (APC/C) is a critical and tightly regulated E3 ligase that orchestrates the cellular life cycle by controlling the degradation of cell cycle regulators. An intriguing feature of this complex is an autoinhibition mechanism: an intrinsically disordered loop domain, Apc1-300L, blocks Cdc20 coactivator binding, yet phosphorylation of Apc1-300L counteracts this autoinhibition. Many such disordered loops within APC/C remain unexplored. Our systematic analysis of loop-deficient APC/C mutants uncovered a pivotal role for Apc8's C-terminal loop (Apc8-L) in mitotic activation. Apc8-L directly recruits the CDK adaptor protein, Xe-p9/Cks2, positioning the Xe-p9-CDK-CycB complex near Apc1-300L. This stimulates the phosphorylation and removal of Apc1-300L, prompting the formation of active APC/CCdc20. Strikingly, without both Apc8-L and Apc3-L, the APC/C is rendered inactive during mitosis, highlighting Apc8-L's synergistic role with other loops and kinases. This study broadens our understanding of the intricate dynamics in APC/C regulation and provides insights on the regulation of macromolecular complexes.

ESM1 enhances fatty acid synthesis and vascular mimicry in ovarian cancer by utilizing the PKM2-dependent warburg effect within the hypoxic tumor microenvironment.

BACKGROUND: The hypoxic tumor microenvironment is a key factor that promotes metabolic reprogramming and vascular mimicry (VM) in ovarian cancer (OC) patients. ESM1, a secreted protein, plays an important role in promoting proliferation and angiogenesis in OC. However, the role of ESM1 in metabolic reprogramming and VM in the hypoxic microenvironment in OC patients has not been determined. METHODS: Liquid chromatography coupled with tandem MS was used to analyze CAOV3 and OV90 cells. Interactions between ESM1, PKM2, UBA2, and SUMO1 were detected by GST pull-down, Co-IP, and molecular docking. The effects of the ESM1-PKM2 axis on cell glucose metabolism were analyzed based on an ECAR experiment. The biological effects of the signaling axis on OC cells were detected by tubule formation, transwell assay, RT‒PCR, Western blot, immunofluorescence, and in vivo xenograft tumor experiments. RESULTS: Our findings demonstrated that hypoxia induces the upregulation of ESM1 expression through the transcription of HIF-1α. ESM1 serves as a crucial mediator of the interaction between PKM2 and UBA2, facilitating the SUMOylation of PKM2 and the subsequent formation of PKM2 dimers. This process promotes the Warburg effect and facilitates the nuclear translocation of PKM2, ultimately leading to the phosphorylation of STAT3. These molecular events contribute to the promotion of ovarian cancer glycolysis and vasculogenic mimicry. Furthermore, our study revealed that Shikonin effectively inhibits the molecular interaction between ESM1 and PKM2, consequently preventing the formation of PKM2 dimers and thereby inhibiting ovarian cancer glycolysis, fatty acid synthesis and vasculogenic mimicry. CONCLUSION: Our findings demonstrated that hypoxia increases ESM1 expression through the transcriptional regulation of HIF-1α to induce dimerization via PKM2 SUMOylation, which promotes the OC Warburg effect and VM.

CDK1-PP2A-B55 interplay ensures cell cycle oscillation via Apc1-loop300.

Cell cycle control relies on a delicate balance of phosphorylation with CDK1 and phosphatases like PP1 and PP2A-B55. Yet, identifying the primary substrate responsible for cell cycle oscillations remains a challenge. We uncover the pivotal role of phospho-regulation in the anaphase-promoting complex/cyclosome (APC/C), particularly through the Apc1-loop300 domain (Apc1-300L), orchestrated by CDK1 and PP2A-B55. Premature activation of PP2A-B55 during mitosis, induced by Greatwall kinase depletion, leads to Apc1-300L dephosphorylation, stalling APC/C activity and delaying Cyclin B degradation. This effect can be counteracted using the B55-specific inhibitor pEnsa or by removing Apc1-300L. We also show Cdc20's dynamic APC/C interaction across cell cycle stages, but dephosphorylation of Apc1-300L specifically inhibits further Cdc20 recruitment. Our study underscores APC/C's central role in cell cycle oscillation, identifying it as a primary substrate regulated by the CDK-PP2A partnership.

Epicoccum sorghinum Causing Leaf Spot on Polygonatum cyrtonema in China.

Polygonatum cyrtonema Hua (family Asparagaceae) is a traditional Chinese medicinal plant that is widely cultivated in various parts of China, including Hunan Province. In summer 2022, a leaf spot disease was observed in 10% of the P. cyrtonema plants (Huang jing) in 18 hectares of this crop in the Hongjiang District (27°18'4″N, 110°11'1″E) of Hunan Province. The initial symptoms of the disease were brown spots on young leaves, and adjacent tissues gradually changed from green to yellow. The entire leaf then became yellow, withered, and eventually exhibited a thn and black appearance. In total, 12 diseased plants from four sampling sites (three plants per site) were collected for laboratory analysis to address the concerns of P. cyrtonema growers. Symptomatic leaf samples were selected, and the leaf fragments containing infected parts of the plants were disinfected with 75% ethanol for 1 min, then immersed in 2.5% hypochlorite for 45 s. After disinfection, symptomatic leaf samples were rinsed three times with sterile water, placed on potato saccharose agar containing 50 µg/ml kanamycin and incubated at 25°C for 2 days. Subsequently, 12 fungal isolates were isolated from various leaf samples through hyphal tip transferring. Ten of the 12 fungal isolates had similar morphological features, and one of them (isolate hjh) was used as the representative isolate for the study. With a growth rate of 6.3 mm per day, its white colonies transformed into red concentric rings in five days; they gradually became black after 10 days of growth. The chlamydospores were round (4.0-9.9 × 3.1-9.3 µm, n = 30), whereas the conidia were ovate (8.0-12.1 × 3.2-6.5 µm, n = 30). The morphological features of the isolate hjh were similar to the features of Epicoccum spp. (Aveskamp et al. 2010). The internal transcribed spacer (ITS) region (including the partial ITS1 sequence and the 5.8S and ITS2 complete sequences), ß-tubulin (tub) gene, and large subunit (LSU) rRNA gene, were amplified from the isolate hjh using the primer pairs ITS5/ITS4, Bt2a/Bt2b, and LROR/LR5, respectively (Taguiam et al. 2021). BLASTn analysis showed that the ITS (OR253745), tub (OR253764), and LSU (OR253746) sequences generated from the isolate hjh were 98-99% similar to the sequences of E. sorghinum strains CBS 179.80 and CBS 627.68. Subsequently, the ITS, tub, and LSU sequences were combined using Sequence Matrix software; phylogenetic analysis via Bayesian and maximum likelihood methods (Vaidya et al. 2011; Li et al. 2021) classified the isolate hjh into the E. sorghinum clade. To fulfill Koch's postulates, pathogenicity tests were conducted on healthy (lesion-free and disease-free) 2-year-old P. cyrtonema plants. Three healthy plants were inoculated by spraying whole plant until run-off with a spore suspension of the isolate hjh (1 × 106 conidia/ml); Three other healthy plants were sprayed with sterile water as controls. The inoculated plants were incubated in a growth chamber at 25 ± 2°C with 85% humidity for 28 days(Chen et al. 2021). Leaves from the inoculated plants gradually became brown within 15 days. Finally, the plants died 28 days after inoculation. The control plants showed no symptoms throughout the experimental period. Isolates (isolate hjh1, hjh2 and hjh3) that were reisolated from the inoculated plants exhibited morphologically similar characteristics and molecularly identical to the original isolate hjh. To our knowledge, this is the first report of E. sorghinum causing leaf spot disease on P. cyrtonema. The results of this study may facilitate the production of P. cyrtonema in China.

RPGR is a guanine nucleotide exchange factor for the small GTPase RAB37 required for retinal function via autophagy regulation.

Although the small GTPase RAB37 acts as an organizer of autophagosome biogenesis, the upstream regulatory mechanism of autophagy via guanosine diphosphate (GDP)-guanosine triphosphate (GTP) exchange in maintaining retinal function has not been determined. We found that retinitis pigmentosa GTPase regulator (RPGR) is a guanine nucleotide exchange factor that activates RAB37 by accelerating GDP-to-GTP exchange. RPGR directly interacts with RAB37 via the RPGR-RCC1-like domain to promote autophagy through stimulating exchange. Rpgr knockout (KO) in mice leads to photoreceptor degeneration owing to autophagy impairment in the retina. Notably, the retinopathy phenotypes of Rpgr KO retinas are rescued by the adeno-associated virus-mediated transfer of pre-trans-splicing molecules, which produce normal Rpgr mRNAs via trans-splicing in the Rpgr KO retinas. This rescue upregulates autophagy through the re-expression of RPGR in KO retinas to accelerate GDP-to-GTP exchange; thus, retinal homeostasis reverts to normal. Taken together, these findings provide an important missing link for coordinating RAB37 GDP-GTP exchange via the RPGR and retinal homeostasis by autophagy regulation.

First Report on Apothecium Deformity of Morchella importuna Caused by Alternaria alternata in China.

Morel mushrooms (Morchella spp.) are highly regarded globally for their distinctive texture and savory flavor. In 2022, the cultivation area for morel mushrooms in China reached nearly 20,000 hectares, with predominant cultivars including M. sextelata, M. importuna and M. exima (Bian et al., 2024). In March 2022, however, deformities of friting bodies were observed in M. importna at morel mushroom farms in Huaihua city (28.43°N, 110.47°), China, with an incidence rate ranging from 5% to 10%. The disease symptoms begin with the invasion of the hymenium of morel mushroom by white cotton-like mycelia, ultimately resulting in halted fruiting body growth and the manifestation of anomalous fruiting body morphology. Infected samples were collected from the morel growers. Following sterilization with 75% ethanol of the surrounding tissue of infected samples, the white hyphae from the morel lesions were picked out using a dissecting needle, and incubated onto potato saccharose agar medium supplemented with 60 mg/L streptomycin at 25°C. Studies showed that seven out of nine fungal isolates exhibiting identical morphological features rapidly grew on the same culture medium described above, reaching a length of 75 mm in 4 to 5 days at 25°C. The white and thick hyphal colonies of these isolates gradually filled with brown spore powder. Generally, the conidia of the hyphal colonies were polyblastic with protrusions at the tips, measuring 75 to 165 × 36 to 50 µm (n = 30) in width and length, displaying colors varying from light reddish brown to grayish brown, and possessing one or five septa. To confirm the identity of the pathogen, the region of the internal transcribed spacer region (ITS), 28S nuclear ribosomal large subunit (LSU), and RNA polymerase II second largest subunit (rpb2) genes of the representative isolate H2 were amplified by PCR (Taguiam, et al. 2021). The generated ITS (OR338304), rpb2 (OR452112) and LSU (OR338334) from the isolate H2 had 98-100% similarity to the Alternaria alternata strains ATCC 6663 and CBS 880.95 in BLASTn analysis. ITS, rpb2 and LSU sequences were assembled using Sequence Matrix, and their homogeneity was assessed with PAUP (Vaidya et al., 2011). Bayesian (MrBayes-3.2.7a) and maximum-likelihood (RAxML1.3.1) methods, utilizing the best fit GTR+G+I model obtained from MrModeltest 2.3, were employed for phylogenetic analysis (Aveskamp et al. 2010). Based on morphological characteristics and phylogenetic analysis, the isolate H2 was identified as A. alternata. In the second year post-disease, disease-free morels, with a height of 3 cm, were cultivated in field greenhouses and used for test. A 15 ml suspension (1 × 106 conidia/ml) was applied to 15 young fruiting bodies and their corresponding substrate soil. The results showed that the reappearance of white cotton-like mycelia and deformed M. importuna fruiting bodies within 7 days post-inoculation with the spore suspension, as opposed to the controls. The isolates (H2-1, H2-2 and H2-3) were reisolated from the infected tissues and identified as A. alternata based on its morphological features and phylogenetic analyses. In this study, a similar investigation was previously conducted on cultivated quinoa (Chenopodium quinoa) in Eastern Denmark (Colque-Little et al., 2023). This study marks the first documentation of A. alternata causing deformities in M. importuna fruiting bodies. These deformities occur under conditions of high-temperature (>22°C) and high humidity (>88%). Our findings provide crucial insights for managing A. alternata in M. importuna cultivation in China.