ABSTRACT
Objective To investigate the expression and the potential roles of long non-coding RNA(lncRNA)cancer susceptibility candidate 2(CASC2)and imprinted gene H19 in extrahepatic cholangiocarcinoma(ECC). Methods Four samples from patients with ECC were collected for high-throughput sequencing which was conducted to reveal the transcriptomic profiles of lncRNA CASC2 and H19.Bioinformatics tools were employed to predict the potential roles of the two genes.Another 22 ECC tissue samples and the cholangiocarcinoma cell lines(RBE,QBC939,HuH-28,and HuCCT1)with different degrees of differentiation were selected for validation.The para-carcinoma tissue and normal human intrahepatic biliary epithelial cell(HIBEC)were used as the control groups.The expression levels of lncRNA CASC2 and H19 in carcinoma tissue,para-carcinoma tissue,and cell lines were determined by real-time quantitative polymerase chain reaction(qRT-PCR).The correlation analysis was carried out for the clinical indicators of patients with the expression levels of the target genes. Results The two target genes showed significantly different expression between carcinoma tissue and para-carcinoma tissue(all P<0.05).Specifically,CASC2 had higher expression level in the carcinoma tissue than in the para-carcinoma tissue(t=1.262,P=0.025),whereas the expression of H19 showed an opposite trend(t=1.285,P=0.005).The expression levels of CASC2 in QBC939(t=8.114,P=0.015)and HuH-28(t=9.202,P=0.012)cells were significantly higher than that in the control group.The expression levels of H19 were significantly lower in RBE(t=-10.244,P<0.001),QBC939(t=-10.476,P<0.001),HuH-28(t=-19.798,P<0.001),and HuCCT1(t=-16.193,P=0.004)cells than in the control group.Bioinformatics analysis showed that CASC2 was mainly involved in the metabolic process and H19 in the development of multicellular organisms.Both CASC2 and H19 were related to catalytic activity.The expression level of lncRNA CASC2 was correlated with pathological differentiation(χ 2=6.222,P=0.022)and lymph node metastasis(χ2=5.455,P=0.020),and that of lncRNA H19 with pathological differentiation(χ2=1.174,P=0.029)and tumor size(χ2=-0.507,P=0.037). Conclusions In the case of ECC,lncRNA CASC2 and H19 have transcription disorders.lncRNA CASC2 is generally up-regulated in the carcinoma tissue,while H19 is down-regulated.Both genes have the potential to become new molecular markers for ECC.
Subject(s)
Humans , Bile Duct Neoplasms/genetics , Bile Ducts, Intrahepatic/metabolism , Cholangiocarcinoma/genetics , Gene Expression Regulation, Neoplastic , RNA, Long Noncoding/genetics , Tumor Suppressor Proteins/geneticsABSTRACT
Botryosphaeriaceae are an important fungal family that cause woody plant diseases worldwide. Recent studies have established a correlation between environmental factors and disease expression; however, less is known about factors that trigger these diseases. The current study reports on the 43.3 Mb de novo genome of Lasiodiplodia theobromae and five other genomes of Botryosphaeriaceae pathogens. Botryosphaeriaceous genomes showed an expansion of gene families associated with cell wall degradation, nutrient uptake, secondary metabolism and membrane transport, which contribute to adaptations for wood degradation. Transcriptome analysis revealed that genes involved in carbohydrate catabolism, pectin, starch and sucrose metabolism, and pentose and glucuronate interconversion pathways were induced during infection. Furthermore, genes in carbohydrate-binding modules, lysine motif domain and the glycosyl hydrolase gene families were induced by high temperature. Among these genes, overexpression of two selected putative lignocellulase genes led to increased virulence in the transformants. These results demonstrate the importance of high temperatures in opportunistic infections. This study also presents a set of Botryosphaeriaceae-specific effectors responsible for the identification of virulence-related pathogen-associated molecular patterns and demonstrates their active participation in suppressing hypersensitive responses. Together, these findings significantly expand our understanding of the determinants of pathogenicity or virulence in Botryosphaeriaceae and provide new insights for developing management strategies against them.
ABSTRACT
Conidia of the obligate biotrophic fungal pathogen Blumeria graminis f. sp. tritici (Bgt) play a vital role in its survival and rapid dispersal. However, little is known about the genetic basis for its asexual reproduction. To uncover the primary metabolic and regulatory events during conidiation, we sequenced the transcriptome of Bgt epiphytic structures at 3 (vegetative hyphae growth), 4 (foot cells initiation), and 5 (conidiophore erection) days post-inoculation (dpi). RNA-seq analyses identified 556 and 404 (combined 685) differentially expressed genes (DEGs) at 4 and 5 dpi compared with their expression levels at 3 dpi, respectively. We found that several genes involved in the conversion from a variety of sugars to glucose, glycolysis, the tricarboxylic acid cycle (TAC), the electron transport chain (ETC), and unsaturated fatty acid oxidation were activated during conidiation, suggesting that more energy supply is required during this process. Moreover, we found that glucose was converted into glycogen, which was accumulated in developing conidiophores, indicating that it could be the primary energy storage molecule in Bgt conidia. Clustering for the expression profiles of 91 regulatory genes showed that calcium (Ca2+), H2O2, and phosphoinositide (PIP) signaling were involved in Bgt conidiation. Furthermore, a strong accumulation of H2O2 in developing conidiophores was detected. Application of EGTA, a Ca2+ chelator, and trifluoperazine dihydrochloride (TFP), a calmodulin (CaM) antagonist, markedly suppressed the generation of H2O2, affected foot cell and conidiophore development and reduced conidia production significantly. These results suggest that Ca2+ and H2O2 signaling play important roles in conidiogenesis and a crosslink between them is present. In addition to some conidiation-related orthologs known in other fungi, such as the velvet complex components, we identified several other novel B. graminis-specific genes that have not been previously found to be implicated in fungal conidiation, reflecting a unique molecular mechanism underlying asexual development of cereal powdery mildews.
