Knockdown of BUB1B Inhibits the Proliferation, Migration, and Invasion of Colorectal Cancer by Regulating the JNK/c-Jun Signaling Pathway.

Background: Colorectal cancer (CRC) ranks as the third most common cancer, accounting for a significant number of cancer-related deaths worldwide every year. Yet, the molecular mechanisms responsible for the progression of this malignancy are not fully understood. Numerous studies indicate that BUB1 mitotic checkpoint serine/threonine kinase B (BUB1B) plays a role in the progression of various malignant tumors. However, the specific biological functions and the detailed mechanisms of how BUB1B influences CRC are still not completely known. This study aimed to explore the expression and role of BUB1B in CRC. Materials and Methods: To achieve this, the expression levels of BUB1B in human CRC tissues and cell lines were examined using real-time polymerase chain reaction and Western blotting. The role and associated mechanisms of BUB1B in CRC cell progression were assessed both in vitro and in vivo using RNA interference. Results: The findings of this study revealed an elevated expression of BUB1B in both CRC tissues and cell lines. The silencing of BUB1B in CRC cell lines notably inhibited cell proliferation, migration, and invasion, leading to cell cycle arrest and apoptosis. In addition, the knockdown of BUB1B inhibited the JNK/c-Jun signaling pathway, increased the expression of proapoptotic proteins, and decreased the expression of antiapoptotic proteins. The effects of BUB1B knockdown on CRC cell progression were reversed by the JNK activator PAF(C-16). Conclusions: In summary, the suppression of BUB1B hindered malignant tumor progression and heightened apoptosis and cell cycle arrest in CRC cells via the JNK/c-Jun pathway. Importantly, the removal of BUB1B expression curtailed tumor growth in human CRC xenografts in nude mice, suggesting its potential as a promising therapeutic target for CRC patients. ClinicalTrials.gov ID: No.2019 K-C086.

Molecular basis of methyl-salicylate-mediated plant airborne defence.

Aphids transmit viruses and are destructive crop pests1. Plants that have been attacked by aphids release volatile compounds to elicit airborne defence (AD) in neighbouring plants2-5. However, the mechanism underlying AD is unclear. Here we reveal that methyl-salicylate (MeSA), salicylic acid-binding protein-2 (SABP2), the transcription factor NAC2 and salicylic acid-carboxylmethyltransferase-1 (SAMT1) form a signalling circuit to mediate AD against aphids and viruses. Airborne MeSA is perceived and converted into salicylic acid by SABP2 in neighbouring plants. Salicylic acid then causes a signal transduction cascade to activate the NAC2-SAMT1 module for MeSA biosynthesis to induce plant anti-aphid immunity and reduce virus transmission. To counteract this, some aphid-transmitted viruses encode helicase-containing proteins to suppress AD by interacting with NAC2 to subcellularly relocalize and destabilize NAC2. As a consequence, plants become less repellent to aphids, and more suitable for aphid survival, infestation and viral transmission. Our findings uncover the mechanistic basis of AD and an aphid-virus co-evolutionary mutualism, demonstrating AD as a potential bioinspired strategy to control aphids and viruses.

Targeting of MAD2L1 by miR-515-5p involves the regulation of cell cycle arrest and apoptosis of colorectal cancer cells.

Although many previous studies have found that the mitotic arrest deficient 2-like 1 (MAD2L1) protein contributes to the proliferation of colorectal cancer (CRC) cells, but the upstream mechanism of MAD2L1 is still largely elusive. This study aimed to explore the microRNAs (miRNAs) upstream of MAD2L1 to improve our understanding of the mechanism of the MAD2L1 gene in CRC. The upstream target miRNAs (miR-515-5p) of MAD2L1 were predicted by the online databases miRWalk, miRDIP, and TargetScan. Quantitative real-time PCR (qRT-PCR) was used to detect the expression level of miR-515-5p in human CRC tissues. The targeting relationship between miR-515-5p and MAD2L1 was tested by dual luciferase reporter gene assays. The effects of miR-515-5p on the biological behaviors of CRC cells by regulating MAD2L1 expression were verified by qRT-PCR, western blot, Cell Counting Kit-8, and flow cytometry. The results showed that miR-515-5p was a highly reliable upstream miRNA of the MAD2L1 gene. As an upstream target miRNA of MAD2L1, miR-515-5p was lowly expression in CRC tissues. The overexpression of miR-515-5p could inhibit the proliferation of CRC cells and induce cell cycle arrest at the G1 phase leading to cell apoptosis. However, MAD2L1 gene overexpression could reverse the effects of miR-515-5p overexpression on the biological behaviors of CRC cells above. This study illustrated that miR-515-5p can inhibit proliferation and induce G1 phase arrest leading to apoptosis in CRC cells. The mechanism underlying this phenomenon may be related to the negative targeted regulation of MAD2L1.

Live imaging and quantitation of insect feeding-induced Ca2+ signal using GCaMP3-based system in Nicotiana benthamiana.

Wounding evokes transient increases in cytosolic calcium (Ca2+) concentration. Visualizing real-time Ca2+ flux provides new insights into Ca2+-signaling pathways. Here, we outline a protocol to detect insect feeding-induced Ca2+ flux elevation in Nicotiana benthamiana leaves based on the GCaMP3 reporter system by Leica fluorescence stereo microscopes (LFSM). LFSM combines super-fast manual screening with high-end imaging capabilities. Through this protocol, we can clearly observe the calcium flow after aphid's piercing-sucking. Additionally, we describe a protocol to quantify Ca2+ level using LFSM. For complete details on the use and execution of this protocol, please refer to Wang et al. (2021).

Cannabidiol inhibits human glioma by induction of lethal mitophagy through activating TRPV4.

Glioma is the most common primary malignant brain tumor with poor survival and limited therapeutic options. The non-psychoactive phytocannabinoid cannabidiol (CBD) has been shown to be effective against glioma; however, the molecular target and mechanism of action of CBD in glioma are poorly understood. Here we investigated the molecular mechanisms underlying the antitumor effect of CBD in preclinical models of human glioma. Our results showed that CBD induced autophagic rather than apoptotic cell death in glioma cells. We also showed that CBD induced mitochondrial dysfunction and lethal mitophagy arrest, leading to autophagic cell death. Mechanistically, calcium flux induced by CBD through TRPV4 (transient receptor potential cation channel subfamily V member 4) activation played a key role in mitophagy initiation. We further confirmed TRPV4 levels correlated with both tumor grade and poor survival in glioma patients. Transcriptome analysis and other results demonstrated that ER stress and the ATF4-DDIT3-TRIB3-AKT-MTOR axis downstream of TRPV4 were involved in CBD-induced mitophagy in glioma cells. Lastly, CBD and temozolomide combination therapy in patient-derived neurosphere cultures and mouse orthotopic models showed significant synergistic effect in both controlling tumor size and improving survival. Altogether, these findings showed for the first time that the antitumor effect of CBD in glioma is caused by lethal mitophagy and identified TRPV4 as a molecular target and potential biomarker of CBD in glioma. Given the low toxicity and high tolerability of CBD, we therefore propose CBD should be tested clinically for glioma, both alone and in combination with temozolomide.Abbreviations: 4-PBA: 4-phenylbutyrate; AKT: AKT serine/threonine kinase; ATF4: activating transcription factor 4; Baf-A1: bafilomycin A1; CANX: calnexin; CASP3: caspase 3; CAT: catalase; CBD: cannabidiol; CQ: chloroquine; DDIT3: DNA damage inducible transcript 3; ER: endoplasmic reticulum; GBM: glioblastoma multiforme; GFP: green fluorescent protein; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; PARP1: poly(ADP-ribose) polymerase; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; SLC8A1: solute carrier family 8 member A1; SQSTM1: sequestosome 1; TCGA: The cancer genome atlas; TEM: transmission electron microscopy; TMZ: temozolomide; TRIB3: tribbles pseudokinase 3; TRPC: transient receptor potential cation channel subfamily C; TRPV4: transient receptor potential cation channel subfamily V member 4.

Clinical, laboratory, endoscopical and histological characteristics predict severe ulcerative colitis.

BACKGROUND/AIMS: Ulcerative colitis is a remitting and relapsing chronic inflammatory disorder of the colon with a mortality rate of 7% from acute severe UC. METHODOLOGY: A retrospective study of the correlation of clinical indicators, laboratory indicators, endoscopical features, and histological features with clinical severity of UC was conducted in 125 UC cases. RESULTS: Diarrhea, mucous and pus in the stool, fever, anemia, weight loss and tachycardia symptoms and the erythrocyte sedimentation rate, concentrations of C-reactive protein, amount of white blood cells and platelets, and positive occult blood test positively correlated with the severity of disease. Whereas, serum total protein, albumin, and hemoglobin levels negatively correlated with the severity of UC. The endoscopic observations of mucosa bleeding, granular mucosa, pseudopolyps, pouch lighter or disappeared, and luminal stenosis as well as the grade and extent of disease were significantly associated with the severity of UC. Histological grade and granular mucosa were significantly associated with the severity of UC. Failure of conservative treatment, severely low gastrointestinal bleeding, and the discovery of a suspicious cancer in the biopsy are the main indicators for colectomy. CONCLUSIONS: Significant reduction in serum total protein, albumin, and hemoglobin concentration is a relatively specific indicator of severe UC.