Molecular Biology最新文献

Abstract

Cervical cancer is a malignant disease that seriously affects women’s health, and the cause of cervical cancer is complex. Microtubule-associated protein 9 (MAP9) localizes to the mitotic spindle and it plays an important role in spindle assembly and centrosome integrity maintenance. The expression and function of MAP9 is unclear in cervical cancer. This study aims to explore the relationship between DNA methylation of MAP9 and cervical cancer, and the function of MAP9 in cervical cancer. qRT-PCR was used to detect the transcriptional expression of MAP9 in cervical cancer tissues and cell lines. The methylation status of MAP9 promoter was determined by bisulfite sequencing PCR (BSP). The DNA methyltransferase inhibitor, 5-Aza-CdR, was used to inhibit the activity of methyltransferase. Then, the cell proliferation was performed by CCK-8 assay after the transfection of MAP9 plasmids. Furthermore, the cell cycle and apoptosis were performed by flow cytometry. It was found that MAP9 was down-regulated both in cervical cancer tissues and cell lines. The promoter and the first exon region of MAP9 was hypermethylated and its mRNA expression could be restored after the treatment of 5-Aza-CdR. Ectopic expression of MAP9 could inhibit cell proliferation and cell cycle, without effecting the cell apoptosis. The low expression of MAP9 in cervical cancer dues to its hypermethylation and acts a potential tumor suppressor gene. MAP9 might be a novel biomarker in cervical cancer.

Abstract

The role of prolyl 3-hydroxylase family member 4 (P3H4, or SC65) in breast cancer and the molecular mechanisms in which this protein was involved were investigated. For this purpose, microchips with cancerous and paracancerous tissues collected from 56 patients with breast cancer were constructed. The P3H4 protein expression was evaluated by immunohistochemistry. Cell lines with decreased and increased P3H4 expression were selected and divided into five groups: P3H4 overexpression and corresponding negative control, P3H4 knockout groups #1 and #2 and corresponding negative control. CCK8 assay, colony formation test, immunoblotting, scratch test, transwell test and flow cytometry were used to determine the relevant cell functions. P3H4 expression was higher in breast cancer cells than in paracancerous tissue. Compared with corresponding negative control, proliferative activity of the cells was inhibited at 24, 48 and 72 h, migration activity and invasion ability of the cells were reduced, autophagy and apoptosis in the cells were increased in P3H4 knockout groups #1 and #2. P3H4 knockout promoted apoptosis of breast cancer cells and inhibited their proliferation, migration, and invasion by activating the BCL-2/BAX/Caspase-3 and AMPK/mTOR pathways. P3H4 knockout promoted apparently autophagy by activating the AMPK/mTOR/ULK1 pathway. However, P3H4 overexpression could promote the proliferation, migration and invasion of breast cancer cells, and inhibited apoptosis and autophagy of mammary gland adenocarcinoma cells MDA-MB-231.

Abstract

Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and poses a severe threat to human health. Ginkgo biloba leaf polysaccharide (GBLP) is a bioactive component, and its sulphated derivative (sulfated GBLP, SGBLP) may exhibit high antitumor activity in certain types of cancers. However, the precise mechanisms of the SGBLP antitumor activity, particularly in HCC, remain unclear. Here, we assessed the effect of SGBLP on HepG2 hepatocellular carcinoma cells. SGBLP was shown to inhibit cellular proliferation and promote apoptosis through the regulation of pro- as well as anti-apoptosis markers, and to induce autophagy by supressing the PI3K/AKT/mTOR pathway. In addition, the autophagy inhibitor 3-melyladenine (3-MA) enhanced the antiproliferative and proapoptotic effects of SGBLP in HepG2 cells. Thus, SGBLP exhibits antitumor activity, and its combination with an autophagy inhibitor may represent a promising anticancer strategy in the treatment of HCC.

Abstract

We have previously shown that 5-arylaminouracil derivatives can inhibit HIV-1, herpesviruses, mycobacteria, and other pathogens through various mechanisms. The purpose of this study was to evaluate the potential of 5-arylaminouracils and their derivatives against leukemia, neuroblastoma, and glial brain tumors. 5-Aminouracils with various substituents and their 5'-norcabocyclic and ribo derivatives were screened for cytotoxicity against two neuroblastoma cell lines (SH-SY5Y and IMR-32), K-562 lymphoblastic cells, HL-60 promyeoloblastic cells, and low-passage variants of well-differentiated glioblastoma multiforme (GBM5522 and GBM6138). Cytotoxicity assessment by the standard MTT test showed that most of the compounds lack significant toxicity towards the above cells. However, 5-(4-isopropylphenylamine)uracil and 5‑(4-tert-butylphenylamine)uracil exhibited a dose-dependent toxic effect towards the GBM6138 cell line with half-maximal inhibitory concentrations (IC₅₀) of 9 and 2.3 μM, respectively. Antitumor activity was for the first time demonstrated for compounds of this type and can serve as a starting point for further research.

Abstract—Many viruses, including SARS-CoV-2, the coronavirus responsible for the COVID-19 pandemic, enter host cells through a process of cell-viral membrane fusion that is activated by proteolytic enzymes. Typically, these enzymes are host cell proteases. Identifying the proteases that activate the virus is not a simple task but is important for the development of new antiviral drugs. In this study, we developed a bioinformatics method for identifying proteases that can cleave viral envelope glycoproteins. The proposed approach involves the use of predictive models for the substrate specificity of human proteases and the application of a structural analysis method for predicting the vulnerability of protein regions to proteolysis based on their 3D structures. Specificity models were constructed for 169 human proteases using information on their known substrates. A previously developed method for structural analysis of potential proteolysis sites was applied in parallel with specificity models. Validation of the proposed approach was performed on the SARS-CoV-2 spike protein, whose proteolysis sites have been well studied.

Abstract—Multiple exogenous or endogenous factors alter gene expression patterns by different mechanisms that are poorly understood. We used RNA-Seq analysis in order to study changes in gene expression in melanoma cells that are capable of vasculogenic mimicry that is inhibited upon the action of an inhibitor of vasculogenic mimicry. Here, we show that the drug induces a strong upregulation of 50 genes that control the cell cycle and microtubule cytoskeleton coupled with a strong downregulation of 50 genes that control different cellular metabolic processes. We found that both groups of genes are simultaneously regulated by multiple sets of transcription factors. We conclude that one way for coordinated regulation of large groups of genes is regulation simultaneously by multiple transcription factors.

Abstract

An RNA interference-based method was proposed to achieve an inducible knockdown of genes essential for cell viability. In the method, a genetic cassette in which a copper ion-dependent inducible metallothionein promoter controls expression of a siRNA precursor is inserted into a genomic pre-integrated transgene by CRIPSR/Cas9 technology. The endogenous siRNA source allows the gene knockdown in cell cultures that are refractory to conventional transfection with exogenous siRNA. The efficiency of the method was demonstrated in Drosophila ovarian somatic cell culture (OSC) for two genes that are essential for oogenesis: Cul3, encoding a component of the multiprotein ubiquitin-ligase complex with versatile functions in proteostasis, and cut, encoding a transcription factor regulating differentiation of ovarian follicular cells.

Abstract—Oral cancer is an aggressive and rapidly progressive disease. The oral cavity is home to over 700 species of microorganisms that regulate metabolism, immune function, and health. There are three types of mechanisms by which bacteria may participate in carcinogenesis. First, bacteria cause chronic inflammation, which stimulates the production of cytokines, including interleukins, interferons, and tumor necrosis factor. Second, bacteria can interact directly with host cells by secreting toxins or by binding to membrane receptors. Finally, the production of metabolites by bacteria may also contribute to carcinogenesis. The importance of the bacteria level and composition in the transition of oral precancerous lesions to cancer has been demonstrated. The relationships of changes in microbiome composition with smoking, inflammation in healthy individuals, as well as with the development of oral cancer in patients, have been studied.

Abstract

The tick-borne encephalitis virus (TBEV) strain C11-13 (GenBank acc. no. OQ565596) of the Siberian genotype was previously isolated from the brain of a deceased person. TBEV C11-13 variants obtained at passages 3 and 8 in SPEV cells were inoculated into the brains of white mice for subsequent passages. Full genome sequences of all virus variants were analyzed by high-throughput sequencing. A total of 41 single nucleotide substitutions were found to occur mainly in the genes for the nonstructural proteins NS3 and NS5 (GenBank MF043953, OP902894, and OP902895), and 12 amino acid substitutions were identified in the deduced protein sequences. Reverse nucleotide and amino acid substitutions were detected after three passages through mouse brains. The substitutions restored the primary structures that were characteristic of the isolate C11-13 from a human patient and changed during the eight subsequent passages in SPEV cells. In addition, the 3′-untranslated region (3′-UTR) of the viral genome increased by 306 nt. The Y3 and Y2 3'-UTR elements were found to contain imperfect L and R repeats, which were probably associated with inhibition of cellular XRN1 RNase and thus involved in the formation of subgenomic flaviviral RNAs (sfRNAs). All TBEV variants showed high-level reproduction in both cell cultures and mouse brains. The genomic changes that occurred during successive passages of TBEV are most likely due to its significant genetic variability, which ensures its efficient reproduction in various hosts and its broad distribution in various climatic zones.

Abstract—RNA polymerase III synthesizes a wide range of noncoding RNAs shorter than 400 nucleotides in length. These RNAs are involved in protein synthesis (tRNA, 5S rRNA, and 7SL RNA), maturation, and splicing of different types of RNA (RPR, MRP RNA, and U6 snRNA), regulation of transcription (7SK RNA), replication (Y RNA), and intracellular transport (vault RNA). BC200 and BC1 RNA genes are transcribed by RNA polymerase III in neurons only where these RNAs regulate protein synthesis. Mutations in the regulatory elements of the genes transcribed by RNA polymerase III as well as in transcription factors of this RNA polymerase are associated with the development of a number of diseases, primarily oncological and neurological. In this regard, the mechanisms of regulation of the expression of the genes containing various RNA polymerase III promoters were actively studied. This review describes the structural and functional classification of polymerase III promoters, as well as the factors involved in the regulation of promoters of different types. A number of examples demonstrate the role of the described factors in the pathogenesis of human diseases.