Genes In Different Types Research Articles

Gene expression profile analysis of gallic acid-induced cell death process

Gallic acid is a natural phenolic compound that displays anti-cancer properties in clinically relevant cell culture and rodent models. To date, the molecular mechanism governing the gallic acid-induced cancer cell death process is largely unclear, thus hindering development of novel therapeutics. Therefore, we performed time-course RNA-sequencing to reveal the gene expression profiles at the early (2nd hour), middle (4th and 6th hour), and late (9th hour) stages of the gallic acid-induced cell death process in HeLa cells. By Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, we found significant changes in transcription of the genes in different types of cell death pathways. This involved the ferroptotic cell death pathway at the early stage, apoptotic pathway at the middle stage, and necroptotic pathway at the late stage. Metabolic pathways were identified at all the stages, indicating that this is an active cell death process. Interestingly, the initiation and execution of gallic acid-induced cell death were mediated by multiple biological processes, including iron and amino acid metabolism, and the biosynthesis of glutathione, as targeting on these pathways suppressed cell death. In summary, our work provides a dataset with differentially expressed genes across different stages of cell death process during the gallic acid induction, which is important for further study on the control of this cell death mechanism.

The predictive values of loss-of-function variants in histone methyltransferases for response to immune checkpoint inhibitors in solid tumors.

2586 Background: Dysregulation of histone methyltransferases (HMTs) has been reported to play critical roles in cancer development. Previous studies showed that many HMTs were recruited to DNA damage sites where they posttranslationally modified chromatin to regulate chromatin-based DNA damage repair (DDR) activities. We hypothesized that loss-of-function (LOF) variants of HMTs may associate with genome instability and tumor mutational burden (TMB). Thus, we explored the associations of LOF variants in some HMTs with TMB and benefit from immune checkpoint inhibitors (ICIs) in solid tumors. Methods: An ICIs treatment cohort from the Memorial Sloan Kettering Cancer Center (MSKCC) was analyzed. The following solid tumor types were enrolled: NSCLC (n = 350), colorectal cancer (n = 110), bladder cancer (n = 215), breast cancer (n = 44), esophagogastric cancer (n = 126), head and neck cancer (n = 139), glioma (n = 117), melanoma (n = 320), and renal cell carcinoma (n = 151). We evaluated 15 HMTs (KMT2A, KMT2B, KMT2C, KMT2D, SETD2, SETD8, EZH1, EZH2, PRDM1, PRDM14, SMYD3, NSD1, WHSC1, WHSC1L1, and DOT1L). Results: The data revealed that LOF variants of KMT2D, SETD2, and KMT2C were more frequent in pan-cancer dataset. Furthermore, we found that LOF variants of 7 HMTs, including KMT2A, KMT2B, KMT2C, KMT2D, NSD1, SETD2, and EZH2, were associated with higher TMB (P < 0.0001). Then we analyzed the associations between LOF variants and overall survival (OS) after ICIs therapy. The results indicated that LOF variants of KMT2A (P = 0.0295), KMT2B (P = 0.0329), KMT2C (P = 0.0122), and SETD2 (P = 0.0004) were significantly associated with prolonged median OS for all the enrolled patients. In this cohort, LOF variants of KMT2A, KMT2B, KMT2C, and SETD2 were most common in bladder cancer, colorectal cancer, colorectal cancer, and renal cell carcinoma, respectively. Then we assessed the predictive values of these four genes for each type of cancer. It was noteworthy that KMT2C LOF variants were significantly correlated with longer median OS in colorectal cancer (P = 0.0171), but not in other cancer types. Surprisingly, we did not observe the predictive roles of LOF variants in KMT2A, KMT2B, and SETD2 genes for response to ICIs therapy in any types of cancer. Conclusions: In pan-cancer dataset, we found that LOF variants of 4 HMTs, such as KMT2A, KMT2B, KMT2C, and SETD2, were correlated with better outcomes of ICIs treatment. However, for different types of cancer, only KMT2C LOF variants were associated with longer median OS in colorectal cancer, suggesting that it may be used as a predictive biomarker for ICIs efficacy in colorectal cancer. Because the sample sizes of patients with KMT2A, KMT2B, or SETD2 LOF variants were small, we did not find the predictive values of LOF variants in these three genes for different types of cancer. Next, we will enroll more patients to address this question.

In Vivo Follow-Up of Gene Inhibition in Solid Tumors Using Peptide-Based Nanoparticles for siRNA Delivery.

Small interfering RNA (siRNA) exhibits a high degree of specificity for targeting selected genes. They are efficient on cells in vitro, but in vivo siRNA therapy remains a challenge for solid tumor treatment as siRNAs display difficulty reaching their intracellular target. The present study was designed to show the in vivo efficiency of a new peptide (WRAP5), able to form peptide-based nanoparticles (PBN) that can deliver siRNA to cancer cells in solid tumors. WRAP5:siRNA nanoparticles targeting firefly luciferase (Fluc) were formulated and assayed on Fluc-expressing U87 glioblastoma cells. The mode of action of WRAP5:siRNA by RNA interference was first confirmed in vitro and then investigated in vivo using a combination of bioluminescent reporter genes. Finally, histological analyses were performed to elucidate the cell specificity of this PBN in the context of brain tumors. In vitro and in vivo results showed efficient knock-down of Fluc expression with no toxicity. WRAP5:siFluc remained in the tumor for at least 10 days in vivo. Messenger RNA (mRNA) analyses indicated a specific decrease in Fluc mRNA without affecting tumor growth. Histological studies identified PBN accumulation in the cytoplasm of tumor cells but also in glial and neuronal cells. Through in vivo molecular imaging, our findings established the proof of concept for specific gene silencing in solid tumors. The evidence generated could be translated into therapy for any specific gene in different types of tumors without cell type specificity but with high molecular specificity.

Upregulation of miR-21 and miR-106b in plasma and tissues as a possible prognostic marker in aggressive breast cancer

Background: The miRNAs are referred to small non-coding RNAs (consisting of 18 to 25 nucleotides). Functional studies have shown their functions to be oncogenes or tumor suppressor genes in different types of cancers. The miR-106b and miR-21 have been identified to participate in the biological behaviors of cells. This study aimed to evaluate the tissue and plasma levels of miR-21 and miR-106b in patients with breast cancer who were diagnosed with ductal carcinoma. Methods: In total, 40 cases of breast cancer patients 180 samples were examined in this project. Samples included ductal carcinoma breast tumors (n=40), normal breast tissues of the margin of the tumor (n=40) and 20 samples from unaffected mammary tissue of females undergoing reduction mammoplasty (control group), plasma samples of patients with breast cancer (n=40), and plasma of non-affected individuals (n=40). The expression levels of miR-106b and miR-21 were determined using SYBR Green real-time RT-PCR assay in breast tissues and plasma of cancerous patients in comparison to the controls. Results: MiR-106b and miR-21 revealed much higher expression in tissues and plasma of patients with breast cancer in comparison to that in the group of control (P<0.001). High levels of mir-106b and miR-21 expression in plasma and tumor tissues were highly correlated with tumors in higher stages and lymph node involvement (P<0.0001). Conclusions: Based on the obtained results, upregulation of miR-106b and miR-21 in the plasma of patients with breast cancer can act as a possible non-invasive biomarker for breast cancer prognosis. Further follow-up studies are required to confirm this.

IgLON5 Regulates the Adhesion and Differentiation of Myoblasts.

IgLON5 is a cell adhesion protein belonging to the immunoglobulin superfamily and has important cellular functions. The objective of this study was to determine the role played by IgLON5 during myogenesis. We found IgLON5 expression progressively increased in C2C12 myoblasts during transition from the adhesion to differentiation stage. IgLON5 knockdown (IgLON5kd) cells exhibited reduced cell adhesion, myotube formation, and maturation and reduced expressions of different types of genes, including those coding for extracellular matrix (ECM) components (COL1a1, FMOD, DPT, THBS1), cell membrane proteins (ITM2a, CDH15), and cytoskeletal protein (WASP). Furthermore, decreased IgLON5 expression in FMODkd, DPTkd, COL1a1kd, and ITM2akd cells suggested that IgLON5 and these genes mutually control gene expression during myogenesis. IgLON5 immunoneutralization resulted in significant reduction in the protein level of myogenic markers (MYOD, MYOG, MYL2). IgLON5 expression was higher in the CTX-treated gastrocnemius mice muscles (day 7), which confirmed increase expression of IgLON5 during muscle. Collectively, these results suggest IgLON5 plays an important role in myogenesis, muscle regeneration, and that proteins in ECM and myoblast membranes form an interactive network that establishes an essential microenvironment that ensures muscle stem cell survival.

DNA methylation abnormalities of imprinted genes in congenital heart disease: a pilot study

BackgroundCongenital heart disease (CHD) is resulted from the interaction of genetic aberration and environmental factors. Imprinted genes, which are regulated by epigenetic modifications, are essential for the normal embryonic development. However, the role of imprinted genes in the etiology of CHD remains unclear.MethodsAfter the samples were treated with bisulfate salt, imprinted genes methylation were measured by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. T test and One-way ANOVA were performed to evaluate the differences among groups. Odds ratios (ORs) were performed to evaluate the incidence risk of CHD in relation to methylation levels.ResultsWe investigated the alterations of imprinted gene germline differential methylation regions (gDMRs) methylation in patients with CHD. Eighteen imprinted genes that are known to affect early embryonic development were selected and the methylation modification genes were detected by massarray in 27 CHD children and 28 healthy children. Altered gDMR methylation level of 8 imprinted genes was found, including 2 imprinted genes with hypermethylation of GRB10 and MEST and 6 genes with hypomethylation of PEG10, NAP1L5, INPP5F, PLAGL1, NESP and MEG3. Stratified analysis showed that the methylation degree of imprinted genes was different in different types of CHD. Risk analysis showed that 6 imprinted genes, except MEST and NAP1L5, within a specific methylation level range were the risk factors for CHDConclusionAltered methylation of imprinted genes is associated with CHD and varies in different types of CHD. Further experiments are warranted to identify the methylation characteristics of imprinted genes in different types of CHD and clarify the etiologies of imprinted genes in CHD.

An information-theoretical analysis of gene nucleotide sequence structuredness for a selection of aging and cancer-related genes.

We provide an algorithm for the construction and analysis of autocorrelation (information) functions of gene nucleotide sequences. As a measure of correlation between discrete random variables, we use normalized mutual information. The information functions are indicative of the degree of structuredness of gene sequences. We construct the information functions for selected gene sequences. We find a significant difference between information functions of genes of different types. We hypothesize that the features of information functions of gene nucleotide sequences are related to phenotypes of these genes.

Essential Genes to Consider in Epstein-Barr Virus-Associated Gastric Cancer: A Systematic Review.

Gastric cancer (GC) is a prevalent malignancy worldwide; the Epstein-Barr Virus (EBV) also affects many people worldwide. An important association has been seen in these two diseases that could explain causality and a possible viral etiology of GC as has been seen with Helicobacter pylori. This study aims to identify genes expressed in malignant cells that are infected with EBV and see if one could be more oncogenic than the other. We conducted a systematic review based on the preferred reporting items for systematic reviews and meta-analysis (PRISMA) guidelines. We had 29 observational studies after inclusion/exclusion criteria and quality assessment for every single study. A total of 1022 patients were evaluated for different types of genes in 29 papers. It was demonstrated that the most expressed genes or the gene most involved were genes that are seen in Epstein-Barr virus-associated gastric cancer (EBVaGC) as latent genes of the EBV-infected cells, which are found in tumor cells. The genes that were mostly involved were LMP2, BNLF2a, and the absence of LMP1 that lead to the expression of BARF1, among other genes. These studies were made on mostly Asian populations, so it is still unknown if these genes involved have a geographical association more than an EBV and GC association.

Comprehensive analysis of differential expression profiles via transcriptome sequencing in SH-SY5Y cells infected with CV-A16.

Coxsackievirus A16 (CV-A16) is one of the viruses that is most frequently associated with hand-foot-and-mouth disease (HFMD). Previous studies have shown that CV-A16 infections are mostly self-limiting, but in recent years, it has been gradually found that CV-A16 infections can also induce neurological complications and eventually cause death in children with HFMD. Moreover, no curative drugs or preventative vaccines have been developed for CV-A16 infection. Therefore, it is particularly important to investigate the mechanism of CV-A16 infection-induced neuropathy. In the current study, transcriptome sequencing technology was used to identify changes in the transcriptome of SH-SY5Y cells infected with CV-A16, which might hide the mechanism of CV-A16-induced neuropathology. The transcriptome profiling showed that 82,406,974, 108,652,260 and 97,753,565 clean reads were obtained in the Control, CV-A16-12 h and CV-A16-24 h groups, respectively. And it was further detected that a total of 136 and 161 differentially expressed genes in CV-A16-12 h and CV-A16-24 h groups, respectively, when compared with Control group. Then, to explore the mechanism of CV-A16 infection, we focused on the common differentially expressed genes at different time points of CV-A16 infection and found that there were 34 differentially expressed genes based on which clustering analysis and functional category enrichment analysis were performed. The results indicated that changes in oxidation levels were particularly evident in the GO term analysis, while only the "Gonadotropin-releasing hormone receptor pathway" was enriched in the KEGG pathway analysis, which might be closely related to the neurotoxicity caused by CV-A16 infection. Meanwhile, the ID2 closely related to nervous system has been demonstrated to be increased during CV-A16 infection. Additionally, the data on differentially expressed non-protein-coding genes of different types within the transcriptome sequencing results were analyzed, and it was speculated that these dysregulated non-protein-coding genes played a pivotal role in CV-A16 infection. Ultimately, qRT-PCR was utilized to validate the transcriptome sequencing findings, and the results of qRT-PCR were in agreement with the transcriptome sequencing data. In conclusion, transcriptome profiling was carried out to analyze response of SH-SY5Y cells to CV-A16 infection. And our findings provide important information to elucidate the possible molecular mechanisms which were linked to the neuropathogenesis of CV-A16 infection.

Single-cell RNA profiling links ncRNAs to spatiotemporal gene expression during C. elegans embryogenesis

Recent studies show that non-coding RNAs (ncRNAs) can regulate the expression of protein-coding genes and play important roles in mammalian development. Previous studies have revealed that during C. elegans (Caenorhabditis elegans) embryo development, numerous genes in each cell are spatiotemporally regulated, causing the cell to differentiate into distinct cell types and tissues. We ask whether ncRNAs participate in the spatiotemporal regulation of genes in different types of cells and tissues during the embryogenesis of C. elegans. Here, by using marker-free full-length high-depth single-cell RNA sequencing (scRNA-seq) technique, we sequence the whole transcriptomes from 1031 embryonic cells of C. elegans and detect 20,431 protein-coding genes, including 22 cell-type-specific protein-coding markers, and 9843 ncRNAs including 11 cell-type-specific ncRNA markers. We induce a ncRNAs-based clustering strategy as a complementary strategy to the protein-coding gene-based clustering strategy for single-cell classification. We identify 94 ncRNAs that have never been reported to regulate gene expressions, are co-expressed with 1208 protein-coding genes in cell type specific and/or embryo time specific manners. Our findings suggest that these ncRNAs could potentially influence the spatiotemporal expression of the corresponding genes during the embryogenesis of C. elegans.

MiR-196: emerging of a new potential therapeutic target and biomarker in colorectal cancer.

Deregulation of microRNAs, as key elements in colorectal cancer (CRC) pathogenesis, is correlated with various stages of this cancer. miR-196 is involved in the initiation and progression of a verity of malignances, especially CRC. miR-196 in CRC cells could target different types of genes with oncogenic and/or tumor suppressor function such as HOX genes, GATA6, SOCS1, SOCS3, ANXA1, DFFA, PDCD4, ZG16 and ING5. Therefore, these genes could be up or down-regulated in cells and subsequently change the capacity of CRC cells in terms of tumor development, progression and, response to therapy. Comprehension of miR-196-associated aberrations underlying the CRC pathogenesis might introduce promising targets for therapy. Additionally, it seems that miR-196 expression profiling, especially circulatory exosomal miR-196, might be useful for diagnosis and prognosis determination of the CRC patients. In this review, at first, we summarize the roles of miR-196 in different types of cancers. After that, a detailed discussion about this miRNA and also their targets in CRC pathogenesis, progression, and response to treatment are represented. Moreover, we highlight the potential utilization of miR-196 and its targets as therapeutic targets and novel biomarkers in early detection and prediction of prognosis in CRC patients.

A pan-cancer analysis of molecular characteristics and oncogenic role of hexokinase family genes in human tumors

Hexokinase (HK) plays a key role in various biological processes such as glycolysis of tumor cells. However, there is still a lack of systematic understanding of the contribution of HK family genes in different types of cancer. In the present study, we systematically analyzed the molecular changes and clinical correlations of HK family genes in 33 types of cancer extracted from more than 10,000 subjects. As a result, there were extensive genetic changes in HK family genes and the expression levels of HK family were significantly correlated with the activity of cancer marker-related pathways. In addition, HK family genes may be useful in predicting prognosis and therapeutic efficacy. Moreover, HK1,HK2 and HK3 may become potential oncogenes across a variety of cancer types. Furthermore, the oncogenic functions of HK1 in bladder cancer have been confirmed in vitro. Collectively, our results provide valuable resources to guide the mechanism and therapeutic analysis concerning the role of HK family genes in cancer.

Identification of Differentially Expressed Genes in Different Types of Broiler Skeletal Muscle Fibers Using the RNA-seq Technique.

The difference in muscle fiber types is very important to the muscle development and meat quality of broilers. At present, the molecular regulation mechanisms of skeletal muscle fiber-type transformation in broilers are still unclear. In this study, differentially expressed genes between breast and leg muscles in broilers were analyzed using RNA-seq. A total of 767 DEGs were identified. Compared with leg muscle, there were 429 upregulated genes and 338 downregulated genes in breast muscle. Gene Ontology (GO) enrichment indicated that these DEGs were mainly involved in cellular processes, single organism processes, cells, and cellular components, as well as binding and catalytic activity. KEGG analysis shows that a total of 230 DEGs were mapped to 126 KEGG pathways and significantly enriched in the four pathways of glycolysis/gluconeogenesis, starch and sucrose metabolism, insulin signalling pathways, and the biosynthesis of amino acids. Quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) was used to verify the differential expression of 7 selected DEGs, and the results were consistent with RNA-seq data. In addition, the expression profile of MyHC isoforms in chicken skeletal muscle cells showed that with the extension of differentiation time, the expression of fast fiber subunits (types IIA and IIB) gradually increased, while slow muscle fiber subunits (type I) showed a downward trend after 4 days of differentiation. The differential genes screened in this study will provide some new ideas for further understanding the molecular mechanism of skeletal muscle fiber transformation in broilers.

EXPRESSION PATTERNS OF THE HUMAN RIBOSOMAL PROTEIN GENES eL14 AND uS19 IN COLON CANCER IS DEPENDENT ON THE TYPE AND STAGE OF THE CANCER CELL

Although the association of some ribosomal protein genes with colorectal cancer is widely known, the detailed mechanisms and complete list of associated genes is lacking. More importantly, the behaviours of these genes in different types and stages of the cancer are poorly understood. Herein we report the study of two ribosomal protein genes in cell lines derived from different sites and stages of colon cancer. Specifically, we analysed the expression pattern of eL14 and uS19 in HCT116 and SW480 cell lines. These two genes, although associated with a wide variety of cancer types, are poorly or have not been studied in colorectal cancer. Semi-quantitative reverse transcription – polymerase chain reaction (RT-PCR) approach was used, together with Students’ t-test validation. We found a significantly (p < 0.05) differential eL14 and uS19 expression patterns between HCT116 and SW480 cell lines. Our findings suggest that eL14 and uS19 have higher activity in a poorly differentiated cell line derived from advanced (metastatic) stage (Duke’s Stage D) colorectal carcinoma tissues compared to the moderately differentiated cell line derived from a mid-stage (Duke’s Stage B) colorectal adenocarcinoma tumour. This will have important implications for both ribosomal protein genes as type and stage specific biomarkers for colon cancer

The mechanism underlying histone deacetylases regulating cardiac hypertrophy

Cardiac hypertrophy is a compensatory response that occurs as a result of increased hemodynamic requirement in peripheral tissues. In the process of cardiac hypertrophy, the expression of different types of genes in different stages is transcriptionally regulated by multiple-level physiological and pathological signals. Histone acetylation, as the most extensive post-translational modification, is closely controlled by the antagonistic histone acetyltransferases (HAT) and histone deacetylases (HDACs). Recent studies have shown that HDACs, as a family of enzymes that inhibit transcription and contain highly conserved deacetylase domains, regulate gene expression during cardiac hypertrophy through a variety of pathways. In this review, we mainly summarize the research progress on histone deacetylase in cardiac hypertrophy. By elucidating the role and molecular mechanism of different HDACs in cardiac hypertrophy, it provides new ideas for the treatment of different types of cardiac hypertrophy and heart failure, and molecular targets for new drug design.

Which type of cancer patients are more susceptible to the SARS-COX-2: Evidence from a meta-analysis and bioinformatics analysis.

ObjectiveNumerous studies have reported that patients with cancer seem to be more likely to be diagnosed with COVID-19. However, it is still unknown the prevalence of different types of cancer in patients with COVID-19. Therefore, this study will explore which type of cancer patients are more susceptible to the SARS-COX-2.MethodsEligible studies were identified by searching several electronic databases for relevant studies published before April 28, 2020. The language was restricted to English or Chinese. The meta-analysis was performed using Stata 12.0. The GEPIA database evaluated the expression of SARS-COX-2 infection key genes in different types of normal and tumor samples.ResultsA total of 6 studies, including 205 patients, were identified to be eligible for this meta-analysis. Among the cancer patients infected by SARS-COX-2, the proportion of patients with the lung, colorectal, breast, esophagus, bladder, pancreatic and cervical cancer were 24.7 %, 20.5 %, 13.0 %, 7.6 %,7.3 %,6.1 %,and 6.0 %, respectively. These findings were also corroborated by the results of the GEPIA database.ConclusionCompared with other types of cancer, lung cancer and colorectal cancer are more susceptible to SARS-COX-2 infection.

A memetic algorithm with novel semi-constructive evolution operators for permutation flowshop scheduling problem

This paper proposes a memetic algorithm (MA) with novel semi-constructive crossover and mutation operators (MASC) to minimize makespan in permutation flowshop scheduling problem (PFSP). MASC combines the strengths of genetic algorithm (GA), simulated annealing (SA), and Nawaz–Enscore–Ham (NEH) algorithm. The aim is to enhance GA in identifying promising areas in the search space, whose local optima will be subsequently located by SA. This is achieved by means of novel crossover and mutation operators that construct chromosomes by using two different types of genes: static and dynamic genes. MASC is tested on the well-known Taillard’s benchmark instances. The proposed operators are compared with traditional operators. The results show that the proposed operators produce considerable improvements. These improvements reach up to 20.79% in the average relative error of best solution and 11.86% in the average relative error of average solution. MASC is compared with fourteen well-known and state-of-the-art algorithms. These algorithms include MA, whale optimization, ant colony optimization, particle swarm optimization, artificial bee colony, monkey search, and iterated greedy. The results show that MASC outperforms all the compared algorithms except three iterated greedy algorithms. Moreover, the improvement in the average relative error of best solution achieved on the best-so-far MA is 37.92%. Therefore, MASC can be considered as one of the best-so-far methods for PFSP.

Identification of Early Warning Signals at the Critical Transition Point of Colorectal Cancer Based on Dynamic Network Analysis.

Colorectal cancer (CRC) is one of the leading causes of cancer-related death worldwide. Due to the lack of early diagnosis methods and warning signals of CRC and its strong heterogeneity, the determination of accurate treatments for CRC and the identification of specific early warning signals are still urgent problems for researchers. In this study, the expression profiles of cancer tissues and the expression profiles of tumor-adjacent tissues in 28 CRC patients were combined into a human protein–protein interaction (PPI) network to construct a specific network for each patient. A network propagation method was used to obtain a mutant giant cluster (GC) containing more than 90% of the mutation information of one patient. Next, mutation selection rules were applied to the GC to mine the mutation sequence of driver genes in each CRC patient. The mutation sequences from patients with the same type CRC were integrated to obtain the mutation sequences of driver genes of different types of CRC, which provide a reference for the diagnosis of clinical CRC disease progression. Finally, dynamic network analysis was used to mine dynamic network biomarkers (DNBs) in CRC patients. These DNBs were verified by clinical staging data to identify the critical transition point between the pre-disease state and the disease state in tumor progression. Twelve known drug targets were found in the DNBs, and 6 of them have been used as targets for anticancer drugs for clinical treatment. This study provides important information for the prognosis, diagnosis and treatment of CRC, especially for pre-emptive treatments. It is of great significance for reducing the incidence and mortality of CRC.

Combined Conditional Knockdown and Adapted Sphere Formation Assay to Study a Stemness-Associated Gene of Patient-derived Gastric Cancer Stem Cells.

Cancer stem cells (CSCs) are implicated in tumor initiation, development and recurrence after treatment, and have become the center of attention of many studies in the last decades. Therefore, it is important to develop methods to investigate the role of key genes involved in cancer cell stemness. Gastric cancer (GC) is one of the most common and mortal types of cancers. Gastric cancer stem cells (GCSCs) are thought to be the root of gastric cancer relapse, metastasis and drug resistance. Understanding GCSCsbiology is needed to advance the development of targeted therapies and eventually to reduce mortality among patients. In this protocol, we present an experimental design using a conditional knockdown system and an adapted sphere formation assay to study the effect of clusterin on the stemness of patient-derived GCSCs. The protocol can be easily adapted to study both in vitro and in vivo function of stemness-associated genes in different types of CSCs.

SETD2 as a regulator of N6-methyladenosine RNA methylation and modifiers in cancer.

Cancer is an unpleasant, painful disease. It is one of the most devastating diseases worldwide diminishing many lives. Many genetic and epigenetic changes occur before cancer develops. Mutation in SETD2 gene is one such example. RNA splicing, DNA damage repair, DNA methylation and histone methylation are some of the biological processes mediated by SETD2. SETD2 (histone H3 lysine 36 methyltransferase) is a frequently mutated gene in different types of cancer. Loss of SETD2 is associated with worse prognosis and aggressive phenotypes. Histone modification is one of the epigenetic regulation having a significant effect on gene regulation. N6-methyladenosine (m6A) mRNA modification is a well-known posttranscriptional modification playing a pivotal role in many normal and pathological processes affecting RNA metabolism. SETD2 catalyses H3K36 trimethylation and in turn H3K36me3 guides the deposition of m6A on nascent RNA transcripts. Finally, this review summarizes the deep understanding of the role of SETD2 in RNA methylation/modification and how SETD2 mutation contributes to tumour development.