Identification of marker genes associated with N6-methyladenosine and autophagy in ulcerative colitis

BACKGROUND Both N6-methyladenosine(m6A)methylation and autophagy are considered relevant to the pathogenesis of ulcerative colitis(UC).However,a systematic exploration of the role of the com-bination of m6A methylation and autophagy in UC remains to be performed.AIM To elucidate the autophagy-related genes of m6A with a diagnostic value for UC.METHODS The correlation between m6A-related genes and autophagy-related genes(ARGs)was analyzed.Finally,gene set enrichment analysis(GSEA)was performed on the characteristic genes.Additionally,the expression levels of four characteristic genes were verified in dextran sulfate sodium(DSS)-induced colitis in mice.RESULTS GSEA indicated that BAG3,P4HB and TP53INP2 were involved in the inflammatory response and TNF-αsignalling via nuclear factor kappa-B.Furthermore,polymerase chain reaction results showed significantly higher mRNA levels of BAG3 and P4HB and lower mRNA levels of FMR1 and TP53INP2 in the DSS group compared to the control group.CONCLUSION This study identified four m6A-ARGs that predict the occurrence of UC,thus providing a scientific reference for further studies on the pathogenesis of UC.

Effects of Chronotherapy of Benazepril on the Diurnal Profile of RAAS and Clock Genes in the Kidney of 5/6 Nephrectomy Rats

Summary： This study investigated the effects of benazepril administered in the morning or evening on the diurnal variation of renin-angiotensin-aldosterone system （RAAS） and clock genes in the kidney. The male Wistar rat models of 5/6 subtotal nephrectomy （STNx） were established. Animals were ran- domly divided into 4 groups： sham STNx group （control）, STNx group, morning benazepril group （MB） and evening benazepril group （EB）. Benazepril was intragastfically administered at a dose of 10 mg/kg/day at 07：00 and 19：00 in the MB group and EB group respectively for 12 weeks. All the animals were synchronized to the light：dark cycle of 12：12 for 12 weeks. Systolic blood pressure （SBP）, 24-h urinary protein excretion and renal function were measured at 11 weeks. Blood samples and kidneys were collected every 4 h throughout a day to detect the expression pattern of renin activity （RA）, angio- tensin Ⅱ （Ang Ⅱ ） and aldosterone （Aid） by radioimmunoassay （RIA） and the mRNA expression profile of clock genes （bmall, dbp and per2） by real-time PCR at 12 weeks. Our results showed that no signifi- cant differences were noted in the SBP, 24-h urine protein excretion and renal function between the MB and EB groups. There were no significant differences in average Aid and RA content of a day between the MB group and EB group. The expression peak of bmall mRNA was phase-delayed by 4 to 8 h, and the diurnal variation of per2 and dbp mRNA diminished in the MB and EB groups compared with the control and STNx groups. It was concluded when the similar SBP reduction, RAAS inhibition and clock gene profile were achieved with optimal dose of benazepril, morning versus evening dosing of benazepril has the same renoprotection effects.

来源于Alcaligenes A-6的D-氨基酰化酶基因的合成与融合表达

将来源于Alcaligenes A-6的D-氨基酰化酶基因用大肠杆菌中的丰沛密码子替换,利用化学和基于聚合酶链反应(PCR)技术的酶促方法进行基因全合成,利用pET-32a构建重组表达载体pET-dan,转化进E.coil BL21(DE3)中进行融合表达.经SDS-PAGE电泳、Western-blot检测和活性测定发现,D-ANase可在大肠杆菌中高效表达,目的蛋白可达到菌体总蛋白的69.2%,密码子优化后基因构建的工程菌发酵活性为96 U/mL,重组蛋白经超声细胞破碎及Ni2+柱亲和层析纯化,比活可达1692.3 U/mg,纯度可达95%以上.

Ultraviolet-attenuated cercariae ofSchistosoma japonicum fail to effectively induce a Th1 response in spite of up-regulating expression of cytotoxicity-related genes in C57BL/6 mice

Objective： To better understand the reason that Schistosoma japonicurn （S. japonicum） ultraviolet （UV）- radiated cercariae could not induce high level of protection in C57BL/6 mice. Methods： Microarray technology was performed to investigate the gene transcription profile in skin draining lymph nodes （sdLNs） at 1 w after exposure to attenuated cercariae （AC） or normal cercariae （NC） of S. japonicum in C57BL/6 mice. The expressions of some representative genes were further confirmed by real-time PCR. Subsequently, the expressions of Th1/Th2 cytokine genes, cytotoxicity-related genes, as well as co-stimulator genes in spleens from AC-vaccinated and NC- infected mice were analyzed by real-time PCR at w 3 and 6 post-exposure. Results： The gene expressions of Th1 cytokines, including interferon-y （IFN-γ）, interleukin （IL）-12 and tumor necrosis factor-α （TNF-α） in the sdLNs were significantly lower in AC-vaccinated mice than in NC-infected mice. Furthermore, the gene expressions of Th1- and Th2- cytokines, including IFN-γ, IL-12, TNF-α, IL-4 and IL-10, in the spleens from AC-vaccinated mice showed little changes at w 3 and 6 post-vaccination. In addition, cytotoxicity-related molecules including granzyme A, granzyme B, granzyme K, perforin 1 and Fas L were up-regulated from the early stage of vaccination, and peaked at the 3rd w after vaccination with UV-AC. Conclusion： UV-AC of S. japonicum could not ef- fectively induce a Thl response in C57BL/6 mice, which may be an explanation for the low protection against parasite challenge, and the role played by up-regulated expression of cytotoxicity-related genes in mice needs to be further investigated.

A correlation study between ITGA6 gene,chromosome 8q24,MSMB genes and prostate cancer

Objective To explore the correlation between IT-GA6 gene ( rs12621278, G ) , MSMB gene ( rs10993994,T) ,chromosome 8q24 9 ( rs10086908, T) and prostate cancer ( PCa) in Beijing residents,and to explore the correlation between genotype and pheno-

Ephrin-B reverse signaling induces expression of wound healing associated genes in IEC-6 intestinal epithelial cells

AIM： Eph receptors and ephrin ligands play a pivotal role in development and tissue maintenance. Since previous data have indicated an involvement of ephrin-B2 in epithelial healing, we investigated the gene expression and downstream signaling pathways induced by ephrin-B mediated cell-cell signaling in intestinal epithelial cells. METHODS： Upon stimulation of ephrin-B pathways in IEC-6 cells with recombinant rat EphB1-Fc, gene expression was analyzed by Affymetrix rat genome 230 high density arrays at different time points. Differentially expressed genes were confirmed by real-time RT-PCR. In addition, MAP kinase pathways and focal adhesion kinase （FAK） activation downstream of ephrin-B were investigated by immunoblotting and fluorescence microscopy. RESULTS： Stimulation of the ephrin-B reverse signaling pathway in IEC-6 cells induces predominant expression of genes known to be involved into wound healing/cell migration, antiapoptotic pathways, host defense and inflammation. Cox-2, c-Fos, Egr-1, Egr-2, and MCP-1 were found among the most significantly regulated genes. Furthermore, we show that the expression of repair- related genes is also accompanied by activation of the ERKI/2 MAP kinase pathway and FAK, two key regulators of epithelial restitution. CONCLUSION： Stimulation of the ephrin-B reverse signaling pathway induces a phenotype characterized by upregulation of repair-related genes, which may partially be mediated by ERK1/2 pathways.

Effect of Exogenous p16ink4a and hRb1 Genes on Cell Cycle Regulation of Osteosarcoma Cell

To study the effect on regulation of cell cycle of osteosarcoma cell line MG63 tranceduced with exogenous p16ink4a and hRbl genes, pIRES-p16ink4a-hRb1, pIRES-p16ink4a and pIREShRbl plasmids were constructed by gene recombination technology. The recombinant plasmid was transferred into osteosarcoma cell line MG63 by metafectene, and the resistant clones were selected by G418 selective medium, mRNA and protein expression of osteosarcoma cell line were assayed by RT-PCR and Western-Blot respectively. Cell cycle and apoptosis were analyzed by subG1 flow cytometric. Cell proliferation was tested by MTT. In the genome of these transfected target cells, the expression of p16ink4a and hRb1 mRNA and protein were detected respectively in vitro. It was demonstrated with subG1 flow cytometric analysis and MTT method that p16ink4a and hRbl genes cooperation more significantly inhibited cell growth and induced a more marked G1 arrest and apoptosis than p16ink4a/hRb1 alone （P〈0.01）. Coexpression of exogenous p16ink4a with hRbl broke the regulatory feedback loop of p16ink4a-cyclinD1/CDK-hRbl and played a more significant role in inhibiting cell growth as well as inducing cell apoptosis than p16ink4a or hRbl did alone in vitro.

EFFECTS OF IFN-a COMBINED WITH IL-6 ON GROWTH AND EXPRESSION OF THE GENES RELATED TO CELL-GROWTH AND APOPTOSIS OF BONE MARROW CELLS FROM PATIENTS WITH CML

Objective:To investigate the effects of interferon-a (IFN-a) and IFN-a combined with interleukin-6 (IL-6) on growth and expression of bcr-abl, bcl-2 and c--myc genes in the mononuclear cells (MNCs) from bone marrow (BM) of patients with chronic myelogenous leukemia (CML). Methods: MNCs were collected from BM of the patients with CML in chronic phase by centrifugation in lymphocyte separation medium and cultured in liquid with IFN-a (200U/ml) or IFN-a (200U/ml) plus IL-6 (100 ng/ml). The growing cells were counted every day. The expression levels of b-actin, bcr-abl, bcl-2 and c-myc genes in the MNCs incubated for 24 h were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and relatively quantitative analysis of the amplified fragments by optical density scanning for the bands on gel. Results: The cell growth was markedly suppressed by IFN-a but the degree of cell-growth inhibition was slightly decreased by IL-6 on the basis of IFN-a effect. The expression of bcr-abl chimeric gene was intensely inhibited by IFN-a or IFN-a plus IL-6. The expression of bcl-2 gene was suppressed by either IFN-a or IFN-a plus IL-6, whereas that of c-myc gene was also inhibited by IFN-a but strongly elevated by IL-6 on the basis of IFN-a action. Conclusions: Both IFN-a and IFN-a plus IL-6 can inhibit the expression of anti-apoptosis gene such as bcr-abl and bcl-2 and regulate the expression of the cs.hn.cn gene related to cell proliferation and differentiation such as c-myc. Either IFN-a or IFN-a combined with IL-6 will serve as a trustful strategy of clinical treatment for CML.

Construction and Identification of a Yeast Two-Hybrid Bait Vector and Its Effect on the Growth of Yeast Cells and the Self-Activating Function of Reporter Genes for Screening of HPV18 E6-Interacting Protein

By using a yeast two-hybrid system,a yeast two-hybrid bait vector was constructed and identified for screening of the HPV18 E6-interacting proteins,and its effects on the growth of yeast cells and the activation of reporter genes were investigated.Total mRNA extracted from Hela cells was reversely transcribed into cDNA.Fragment of HPV18 E6 cDNA was amplified using RT-PCR and directly ligated to the pGBKT7 vector.The recombinant plasmid was confirmed by restriction endonuclease analysis and DNA sequencing.Th...

Construction of genetically engineered macrophages expressing Smad6 and Smad7 genes with adeno-associated virus

Objective: To construct the genetically engineered macrophages expressing Smad6 and Smad7 genes with adeno-associated virus (AAV). Methods: The plasmids containing pcDNA3-Smad6/Flag and pcDNA3-Smad7/Flag were digested with BamHⅠ and XhoⅠ, respectively. Then the Smad6/Flag and Smad7/Flag gene segments obtained were cloned into plasmid pAAV-MCS respectively to construct the recombinant pAAV-Smad6/Flag and pAAV-Smad7/Flag plasmids. The resulting recombinant plasmids (pAAV-Smad6/Flag or pAAV-Smad7/Flag) or pAAV-LacZ plasmid were co-transfected into the HEK 293cells with pHelper and pAAV-RC by calcium-phosphate precipitation method. Recombinant AAV-2 viral particles were prepared from infected HEK293 cells and then were used to infect mouse macrophages. The expressions of Smad6 and Smad7 in macrophages were detected by immunocytochemical staining and expression of b-galactosidase was evaluated by X-gal staining. Results: The recombinant AAV vector containing Smad6 or Smad7 genes was successfully constructed. More than 95% macrophage cells expressed X-gal and Smad6 and Smad7 genes at 72 h after infection. Conclusion: These results indicate that the genetically engineered macrophages can express Smad6 and Smad7 proteins effectively, laying the foundation for the studies of TGF-β-induced diseases in vivo and highlighting the feasibility of macrophage-based gene therapy.

Transcriptomic comparison to identify rapidly evolving genes in Braya humilis

The Brassicaceae species Braya humilis shows broad adaptation to different climatic zones and latitudes. However, the molecular adaptation mechanism of B. humilis is poorly understood. In China, B. humilis is mainly distributed on the QinghaiTibetan Plateau(QTP) and in the adjacent arid region. Previous transcriptome analysis of B. humilis has revealed that 39 salt and osmotic stress response genes are subjected to purifying selection during its speciation. To further explore the adaptation mechanism of B. humilis to an arid environment, OrthoMCL program was employed in this study and 6,268 pairs of orthologous gene pairs with high confidence were obtained between B. humilis and Arabidopsis thaliana. A comparative evolutionary analysis based on nonsynonymous to synonymous substitution ratio(Ka/Ks) was then conducted. There were 64 pairs exhibiting a Ka/Ks ratio more than 0.5 and among which, three instrumental candidate genes, T20487,T22576, and T23757, were identified with strong selection signatures(Ka/Ks >1). The corresponding A. thaliana orthologs are double-stranded RNA-binding domain protein, MADS-box family protein, and NADH-dehydrogenase subunit6, which is encoded by mitochondria genome. This report not only demonstrates the adaptation contribution of fast evolving nuclear genes, but also highlights the potential adaptive value of mitochondria gene to the speciation and adaptation of B. humilis toward the extreme environment in an arid region.

Screening key target genes for coronavirus disease 2019 based on bioinformatics and gene network

Objective:To use bioinformatics and gene networks to screen key target genes of coronavirus disease 2019,which provides references for clinical research and development of drugs for coronavirus disease 2019.Methods:Target genes related to coronavirus disease 2019 were screened in the GeneCards and National Center for Biotechnology Information databases,and the obtained gene data were imported into the Database for Annotation,Visualization and Integrated Discovery(Version 6.8)database to collect the related information about pathways and genes.The genes enriched in the first 20 pathways and the genes whose occurrence frequency≥5 were imported into the String database respectively to construct protein-protein interaction network diagram and compare the two network diagrams.Results:TNF,IL-6,IL-2,IL-8,CXCL8,IL1B,CCL2,IFNG,STAT1,MAPK1,MAPK3,MAPK8,TP53 and RELA are ranked top in the two network diagrams,and the frequency of occurrence in the first 20 pathways was≥5.Conclusion:The incidence of coronavirus disease 2019 is associated with multiple signaling pathways,including influenza A,pathways in cancer,toll-like receptor signaling pathway,hypoxia-inducible factor-1 signaling pathway,et al.TNF,IL-6,IL-2,IL-8,CXCL8,IL1B,CCL2,IFNG,STAT1,MAPK1,MAPK3,MAPK8,TP53 and RELA are closely related to coronavirus disease 2019,which needs to be further studied.By analyzing the pathways of the genes related to coronavirus disease 2019 and the interactive network diagrams between the genes,it is helpful to understand the pathogenesis of the disease and provide a reference for clinical research and development of effective drugs for coronavirus disease 2019.

PIFs interact with SWI2/SNF2-related 1 complex subunit 6 to regulate H2A.Z deposition and photomorphogenesis in Arabidopsis

Light is an essential environmental signal perceived by a broad range of photoreceptors in plants. Among them, the red/far-red light receptor phytochromes function to promote photomorphogenesis, which is critical to the survival of seedlings after seeds germination. The basic-helix-loop-helix transcription factors phytochrome-interacting factors (PIFs) are the pivotal direct downstream components of phytochromes. H2A.Z is a highly conserved histone variant regulating gene transcription, and its incorporation into nucleosomes is catalyzed by SWI2/SNF2-related 1 complex, in which SWI2/SNF2-related 1 complex subunit 6 (SWC6) and actin-related protein 6 (ARP6) serve as core subunits. Here, we show that PIFs physically interact with SWC6 in vitro and in vivo, leading to the disassociation of HY5 from SWC6. SWC6 and ARP6 regulate hypocotyl elongation partly through PIFs in red light. PIFs and SWC6 coregulate the expression of auxin-responsive genes such as IAA6, IAA19, IAA20, and IAA29 and repress H2A.Z deposition at IAA6 and IAA19 in red light. Based on previous studies and our findings, we propose that PIFs inhibit photomorphogenesis, at least in part, through repression of H2A.Z deposition at auxin-responsive genes mediated by the interactions of PIFs with SWC6 and promotion of their expression in red light.

AAV2-PDE6B restores retinal structure and function in the retinal degeneration 10 mouse model of retinitis pigmentosa by promoting phototransduction and inhibiting apoptosis

Retinitis pigmentosa is a group of inherited diseases that lead to retinal degeneration and photoreceptor cell death.However,there is no effective treatment for retinitis pigmentosa caused by PDE6B mutation.Adeno-associated virus(AAV)-mediated gene therapy is a promising strategy for treating retinitis pigmentosa.The aim of this study was to explore the molecular mechanisms by which AAV2-PDE6B rescues retinal function.To do this,we injected retinal degeneration 10(rd10)mice subretinally with AAV2-PDE6B and assessed the therapeutic effects on retinal function and structure using dark-and light-adapted electroretinogram,optical coherence tomography,and immunofluorescence.Data-independent acquisition-mass spectrometry-based proteomic analysis was conducted to investigate protein expression levels and pathway enrichment,and the results from this analysis were verified by real-time polymerase chain reaction and western blotting.AAV2-PDE6B injection significantly upregulated PDE6βexpression,preserved electroretinogram responses,and preserved outer nuclear layer thickness in rd10 mice.Differentially expressed proteins between wild-type and rd10 mice were closely related to visual perception,and treating rd10 mice with AAV2-PDE6B restored differentially expressed protein expression to levels similar to those seen in wild-type mice.Kyoto Encyclopedia of Genes and Genome analysis showed that the differentially expressed proteins whose expression was most significantly altered by AAV2-PDE6B injection were enriched in phototransduction pathways.Furthermore,the phototransductionrelated proteins Pde6α,Rom1,Rho,Aldh1a1,and Rbp1 exhibited opposite expression patterns in rd10 mice with or without AAV2-PDE6B treatment.Finally,Bax/Bcl-2,p-ERK/ERK,and p-c-Fos/c-Fos expression levels decreased in rd10 mice following AAV2-PDE6B treatment.Our data suggest that AAV2-PDE6B-mediated gene therapy promotes phototransduction and inhibits apoptosis by inhibiting the ERK signaling pathway and upregulating Bcl-2/Bax expression in retinitis pigmentosa.

Gene amplification-driven RNA methyltransferase KIAA1429 promotes tumorigenesis by regulating BTG2 via m6A-YTHDF2-dependent in lung adenocarcinoma

Background:Epigenetic alterations have been shown to contribute immensely to human carcinogenesis.Dynamic and reversible N6-methyladenosine(m6A)RNA modification regulates gene expression and cell fate.However,the reasons for activation of KIAA1429(also known as VIRMA,an RNA methyltransferase)and its underlying mechanism in lung adenocarcinoma(LUAD)remain largely unexplored.In this study,we aimed to clarify the oncogenic role of KIAA1429 in the tumorigenesis of LUAD.Methods:Whole-genome sequencing and transcriptome sequencing of LUAD data were used to analyze the gene amplification of RNA methyltransferase.The in vitro and in vivo functions of KIAA1429 were investigated.Transcriptome sequencing,methylated RNA immunoprecipitation sequencing(MeRIP-seq),m6A dot blot assays and RNA immunoprecipitation(RIP)were performed to confirm the modified gene mediated by KIAA1429.RNA stability assays were used to detect the half-life of the target gene.Results:Copy number amplification drove higher expression of KIAA1429 in LUAD,whichwas correlatedwith poor overall survival.Manipulating the expression of KIAA1429 could regulate the proliferation and metastasis of LUAD.Mechanistically,the target genes of KIAA1429-mediated m6A modification were confirmed by transcriptome sequencing and MeRIP-seq assays.We also revealed that KIAA1429 could regulate BTG2 expression in an m6A-dependent manner.Knockdown of KIAA1429 significantly decreased the m6A levels of BTG2 mRNA,leading to enhanced YTH m6A RNA binding protein 2(YTHDF2,the m6A“reader”)-dependent BTG2 mRNA stability and promoted the expression of BTG2;thus,participating in the tumorigenesis of LUAD.Conclusions:Our data revealed the activation mechanism and important role of KIAA1429 in LUAD tumorigenesis,which may provide a novel view on the targeted molecular therapy of LUAD.

A rare missense PAX6 mutation causes atypical aniridia in a three-generation Chinese family

●AIM:To investigate the molecular diagnosis of a threegeneration Chinese family affected with aniridia,and further to identify clinically a PAX6 missense mutation in members with atypical aniridia.●METHODS:Eleven family members with and without atypical aniridia were recruited.All family members underwent comprehensive ophthalmic examinations.A combination of whole exome sequencing(WES)and direct Sanger sequencing were performed to uncover the causative mutation.●RESULTS:Among the 11 family members,8 were clinically diagnosed with congenital aniridia(atypical aniridia phenotype).A rare heterozygous mutation c.622C>T(p.Arg208Trp)in exon 8 of PAX6 was identified in all affected family members but not in the unaffected members or in healthy control subjects.●CONCLUSION:A rare missense mutation in the PAX6 gene is found in members of a three-generation Chinese family with congenital atypical aniridia.This result contributes to an increase in the phenotypic spectrum caused by PAX6 missense heterozygous variants and provides useful information for the clinical diagnosis of atypical aniridia,which may also contribute to genetic counselling and family planning.

Cloning and expression analysis of the chloroplast fructose-1,6-bisphosphatase gene from Pyropia haitanensis

Fructose-1,6-bisphosphatase（FBPase） is one of the key enzymes in Calvin circle and starch biosynthesis. In this study, the full-length of cpFBPase gene from Pyropia haitanensis was cloned by using rapid amplification of cDNA ends（RACE） technology. The nucleotide sequence of PhcpFBPase consists of 1 400 bp, including a 5′ untranslated region（UTR） of 92 bp, a 3′？UTR of 69 bp, and an open reading frame（ORF） of 1 236 bp, which can be translated into a 412-amino-acid putative peptides with a molecular weight of 44.3 kDa and a theoretical pI of 5.23. Multiple sequence alignment indicated that the protein belonged to the chloroplast FBPase enzyme. Phylogenetic analysis showed that the protein assembled with the cpFBPase of a thermal tolerant unicellular red micro-algae Galdieria sulphuraria. Expression patterns analyzed by qRT-PCR revealed that the expression of PhcpFBPase gene in the thallus phage was 7-fold higher than in the conchocelis phage, which suggested the different mechanisms of inorganic carbon utilization among the different life phages of P. haitanensis. And the different response modes of PhcpFBPase mRNA levels to high temperature and desiccation stress indicated that PhcpFBPase played an important role in responsing to abiotic stress.

Pathogenic mutations of TGFBI and CHST6 genes in Chinese patients with Avellino,lattice,and macular corneal dystrophies

Objective:To investigate gene mutations associated with three different types of corneal dystrophies(CDs),and to establish a phenotype-genotype correlation.Methods:Two patients with Avellino corneal dystrophy(ACD),four patients with lattice corneal dystrophy type I(LCD I) from one family,and three patients with macular corneal dystrophy type I(MCD I) were subjected to both clinical and genetic examinations.Slit lamp examination was performed for all the subjects to assess their corneal phenotypes.Genomic DNA was extracted from peripheral blood leukocytes.The coding regions of the human transforming growth factor β-induced(TGFBI) gene and carbohydrate sulfotransferase 6(CHST6) gene were amplified by polymerase chain reaction(PCR) and subjected to direct sequencing.DNA samples from 50 healthy volunteers were used as controls.Results:Clinical examination showed three different phenotypes of CDs.Genetic examination identified that two ACD subjects were associated with homozygous R124H mutation of TGFBI,and four LCD I subjects were all associated with R124C heterozygous mutation.One MCD I subject was associated with a novel S51X homozygous mutation in CHST6,while the other two MCD I subjects harbored a previously reported W232X homozygous mutation.Conclusions:Our study highlights the prevalence of codon 124 mutations in the TGFBI gene among the Chinese ACD and LCD I patients.Moreover,we found a novel mutation among MCD I patients.

Comparative Analysis of Human Genes Frequently and Occasionally Regulated by m^6A Modification

The m^6A modification has been implicated as an important epitranscriptomic marker, which plays extensive roles in the regulation of transcript stability, splicing, translation, and localization. Nevertheless, only some genes are repeatedly modified across various conditions and the principle of m^6A regulation remains elusive. In this study, we performed a systems-level analysis of human genes frequently regulated by m^6A modification （m^6Afreq genes） and those occasionally regulated by m^6A modification （m^6Aocca genes）. Compared to the m^6Aocca genes, the m^6Afreq genes exhibit gene importance-related features, such as lower dN/dS ratio, higher protein-protein interaction network degree, and reduced tissue expression specificity. Signaling network analysis indicates that the m^6Afreq genes are associated with downstream components of signaling cascades, high-linked signaling adaptors, and specific network motifs like incoherent feed forward loops. Moreover, functional enrichment analysis indicates significant overlaps between the m^6Afreq genes and genes involved in various layers of gene expression, such as being the microRNA targets and the regulators of RNA processing. Therefore, our findings suggest the potential interplay between m^6A epitranscriptomic regulation and other gene expression regulatory machineries.

The Application of ISSR Markers to Identify the Fertility Restorer Gene Rf6 in T. timopheevii Cytoplasmic MaleSterile Wheat(Triticum aestivum)

Inter-simple sequence repeat (ISSR) analysis was carried out on a F2 population of 147 plants derived from a cross between a wheat male fertility restorer line 2114 and a male sterile line ND44A. Out of 43 primers examined, 18 primers produced distinguishable, polymorphic bands between the two parents. Linkage analysis in the mapping population showed that two markers UBC-808 and UBC-848 were closely linked with the restorer gene Rf6 of the Triticum timopheevii CMS system. The distance between the two markers and the restorer gene was 7.9 cM and 4.9 cM, respectively. Also two parents were screened with 181 pairs of SSR primers, of which, 34.3% showed polymorphisms. But no locus was found linked with the restorer gene. Compared with the SSR technique, the ISSR approach used in the experiment provided more information and proved to be a valuable method to identify alien fragments.