Identification of up-regulated genes in human uterine leiomyoma by sup-pression subtractive hybridization

In searching for differentially expressed genes in human uterine leiomyomas (ULs), suppression sub-tractive hybridization was used to construct an UL up-regulated library, which turned out to represent 88 genes. After two rounds of screening by reverse Northern analysis, twenty genes were proved to be up-regulated, including seventeen known genes and three genes with unknown function. All these genes were firstly associated with UL. Three genes with notable difference were selected for Northern confirmation. Our results proved the authenticity of the twenty genes. One gene named Phospholipase A2 (PLA2) showed up-regulation in 4/6 of the patients and investigation of tissue distribution indicated that it had obvious expression in prostate, testis, liver, heart and skeletal muscle.     

Identification of sheep ovary genes potentially associated with off-season reproduction

Off-season reproduction is a favorable economic trait for sheep industry.Hu sheep,an indigenous Chinese sheep breed,demonstrates a higher productivity of lambs and displays year-around oestrous behavior under proper nutrition and environment.The genetic basis behind these traits,however,is not well understood.In order to identify genes associated with the off-season reproduction,we constructed a suppression subtractive hybridization(SSH) cDNA library using pooled ovary mRNAs of 6 oestrous Hu females as a tester and the pooled ovary mRNAs of 6 non-oestrous Chinese Merino females as a driver.A total of 382 resulting positive clones were obtained after the SSH.We identified 114 differentially up-regulated genes in oestrous Hu sheep by using subsequent screening and DNA sequencing,of which 8 were previously known,93 were reported for the first time in sheep,and 13 were novel with no significant homology to any sequence in the DNA databases.Functions of the genes identified are related to cell division,signal transduction,structure,metabolism,or cell defense.To validate the results of SSH,6 genes(Ntrk2,Ppap2b,Htra1,Nid1,Serpine2 and Foxola) were selected for conformational analysis using quantitative real-time PCR(qRT-PCR),and two of them(Htral and Foxola) were verified by Northern blot.All of the 6 genes were differentially up-regulated in the ovary of oestrous Hu.It is obvious that off-season reproduction is a complex trait involving multiple genes in multiple organs.This study helps to provide a foundation for the final identification of functional genes involved in the sheep ovary.     

Gene expression profile in immunologically injured liver cell of mice

To study the gene expression profiles between immunologically injured liver cell and normal liver cell of mice and to screen on a large scale the differentially expressed genes associated with the formation of liver injury,the experimental mice were randomly divided into the normal group for controlling and the immunologically liver-injured group induced by BCG and LPS.The liver mRNA of the two groups were extracted respectively and reversely-transcribed to cDNA with the incorpora-tion of different fluorescence(Cy3,Cy5) labeled dUTP as the hybridization probes.The mixed probes were hybridized to the cDNA microarray chips.The fluorescent signal results were acquired by scanner ScanArray 4000 and analyzed with software GenePix Pro 3.0.Among the 14112 target genes,293 genes were found to be significantly differentially expressed,in which 188 genes were up-regulated and 105 genes were down-regulated.Based on the analysis of biological functions of those differentially expressed genes,it was indicated that the occurrence and development of mouse liver damage induced by BCG and LPS were highly correlated with the processes of immune reac-tions,cell synthesis,metabolism,apoptosis and transportation in liver cell,which might be quite im-portant for elucidating the regulatory network of gene expression associated with the liver damage,also important for finally discovering the pathogenic mechanisms of immunological liver damage.     

Expression of PIN Genes in Rice （Oryza sativa L.）： Tissue Specificity and Regulation by Hormones

Twelve genes of the PIN family in rice were analyzed for gene and protein structures and an evolutionary relationship with reported AtPINs in Arabidopsis. Four members of PIN1 （designated as OsPINla-d）, one gene paired with AtPIN2 （OsPIN2）, three members of PIN5 （OsPIN5a-c）, one gene paired with AtPIN8 （OsPIN8）, and three monocot-specific PINs （OsPIN9, OsPINIOa, and b） were identified from the phylogenetic analysis. Tissue-specific expression patterns of nine PIN genes among them were investigated using RT-PCR and GUS reporter. The wide variations in the expression domain in different tissues of the PIN genes were observed. In general, PIN genes are up-regulated by exogenous auxin, while different responses of different PIN genes to other hormones were found.     

Identification of salt-stress responsive genes in rice (Oryza sativa L.) by cDNA array

To identify salt stress-responsive genes, we constructed a cDNA library with the salt-tolerant rice cultivar, Lansheng. About 15000 plasmids were extracted and dotted on filters with Biomeck 2000 HDRT system or by hand. Thirty genes were identified to display altered expression levels responding to 150 mmol/L NaCl. Among them eighteen genes were up-regulated and the remainders down-regulated. Twenty-seven genes have their homologous genes in Gen-Bank Databases. The expression of twelve genes was studied by Northern analysis. Based on the functions, these genes can be classified into five categories, including photosynthesis-related gene, transport-related gene, metabolism-related gene, stress- or resistance-related gene and the others with various functions. The results showed that salt stress influenced many aspects of rice growth. Some of these genes may play important roles in plant salt tolerance.     

Regulated genes in mesenchymal stem cells and gastric cancer

AIM: To investigate the genes regulated in mesenchymal stem cells(MSCs) and diffuse-type gastric cancer(GC),gene expression was analyzed. METHODS: Gene expression of MSCs and diffuse-type GC cells were analyzed by microarray. Genes related to stem cells, cancer and the epithelial-mesenchymal transition(EMT) were extracted from human gene lists using Gene Ontology and reference information. Gene panels were generated, and messenger RNA gene expression in MSCs and diffuse-type GC cells was analyzed. Cluster analysis was performed using the NCSS software.RESULTS: The gene expression of regulator of G-protein signaling 1(RGS1) was up-regulated in diffuse-type GC cells compared with MSCs. A panel of stem-cell related genes and genes involved in cancer or the EMT were examined. Stem-cell related genes, such as growth arrest-specific 6, musashi RNA-binding protein 2 and hairy and enhancer of split 1(Drosophila), NOTCH family genes and Notch ligands, such as delta-like 1(Drosophila) and Jagged 2, were regulated.CONCLUSION: Expression of RGS1 is up-regulated, and genes related to stem cells and NOTCH signaling are altered in diffuse-type GC compared with MSCs.     

Identification and characterization of 177 unreported genes associated with liver regeneration

The mammalian liver has a very strong regeneration capacity after partial hepatectomy (PH). To further learn the genes participating in the liver regeneration (LR), 551 cDNAs selected from subtracted cDNA libraries of the regenerating rat liver were screened by microarray, and their expression profiles were studied by cluster and generalization analyses. Among them, 177 genes were identified unreported and up-or down-regulated more than twofold at one or more time points after PH, of which 62 genes were down-regulated to less than 0.5; 99 genes were up-regulated to 2-10 folds, and 16 genes were either up- or down-regulated at different time points during LR. By using BLAST and GENSCAN, these genes were located on responsible chromosomes with 131 genes on the long arms of the chromosomes. The cluster and generalization analyses showed that the gene expression profiles are similar in 2 and 4, 12 and 16, 96 and 144 h respectively after PH, suggesting that the actions of the genes expressed in the same profiles are similar, and those expressed in different profiles have less similarity. However, the types,characteristics and functions of the 177 genes remain to be further studied.     

Preliminary studies on differential defense responses induced during plant communication

We compared the expression patterns of three representative genes in undamaged tomato and tobacco plants in response to exposure to either tomato or tobacco fed on by Helicoverpa armigera (cotton bollworm). When tomato and tobacco, two species of one family, were incubated in the chambers with the tomato plants damaged by the cotton bollworm, the expression of the PR1, BGL2, and PAL genes was up-regulated in leaves of both plants. However, the levels of gene expression were significantly higher in the tomato than that in the tobacco. In addition, the activities of enzymes, peroxidase, polyphenol oxidase, and lipoxygenase were found to be higher in the tomato than those in the tobacco. Similar results were obtained when the damaged plants were replaced by the tobacco.     

Genome‐wide identification of chitin‐binding proteins and characterization of BmCBP1 in the silkworm,Bombyx mori

The insect cuticle plays important roles in numerous physiological functions to protect the body from invasion of pathogens, physical injury and dehydration. In this report, we conducted a comprehensive genome-wide search for genes encoding proteins with peritrophin A-type (ChtBD2) chitin-binding domain (CBD) in the silkworm, Bombyx mori. One of these genes, which encodes the cuticle protein BmCBP1, was additionally cloned, and its expression and location during the process of development and molting in B. mori were investigated. In total, 46 protein-coding genes were identified in the silkworm genome, including those encoding 15 cuticle proteins analogous to peritrophins with one CBD (CPAP1s), nine cuticle proteins analogous to peritrophins with three CBD (CPAP3s), 15 peritrophic membrane proteins (PMPs), four chitinases, and three chitin deacetylases, which contained at least one ChtBD2 domain. Microarray analysis indicated that CPAP-encoding genes were widely expressed in various tissues, whereas PMP genes were highly expressed in the midgut. Quantitative polymerase chain reaction and western blotting showed that the cuticle protein BmCBP1 was highly expressed in the epidermis and head, particularly during molting and metamorphosis. An immunofluorescence study revealed that chitin co-localized with BmCBP1 at the epidermal surface during molting. Additionally, BmCBP1 was notably up-regulated by 20-hydroxyecdysone treatment. These results provide a genome-level view of the chitin-binding protein in silkworm and suggest that BmCBP1 participates in the formation of the new cuticle during molting.     

Gene expression profiling in apoptotic k562 cells treated by homoharringtonine

Gene chip technology was used to determine the gene expression profiles in apoptotic K562 cells induced by homoharringtonine. The expression of forty-four mRNAs was found to be changed significantly were identified after screening with a gene chip capable of detecting 14,218 different human mRNA species simultaneously. Of these genes, 17 were up-regulated and 27 were down-regulated. Most of them were found to be related to apoptosis, oncogenes, or tumor suppression. Several genes with altered gene expression, such as human transforming growth factor-beta inducible early protein gene （TIEG）, vitamin D3 upregulated protein 1 gene （VDUP1）, RNA binding motif protein 4 gene （RBM4） and v-myc myelocytomatosis viral oncogene homolog （C-MFC）, were confirmed by Northern blot analysis. According to the dynamic gene expression pattern in these apoptotic cells, the activated transforming growth factor-β and tumor necrosis factor signaling pathways play an important role in homoharringtonine-induced apoptosis. TIEG was significantly altered after apoptosis induction, it should be critical for apoptosis signal transmission.     

Methylation profiling of twenty four genes and the concordant methylation behaviours of nineteen genes that may contribute to hepatocellular carcinogenesis

To determine the possible role of the epigenetic mechanisms in carcinogenesis of the hepatocellular carcinoma, we methylation-profiled the promoter CpG islands of twenty four genes both in HCC tumors and the neighboring non-cancerous tissues of twenty eight patients using the methylation-specific PCR (MSP) method in conjunction with the DNA sequencing. In comparison with the normal liver tissues from the healthy donors, it was found thatwhile remained unmethylated the ABL, CAV, EPO, GATA3, LKB 1, NEP, NFL, NIS and p27KIPI genes, varying extents of the HCC specific loss of the epigenetisoermethvlation were found associated with the ABO, AR. CSPG2. cvclin al. DBCCR1,GALR2, IRF7, MGMT, MT 1 A, MYOD 1, OCT6, p57KP2, p73, WT 1 genes, and demethylation with the MAGEA 1 gene, respectively. Judged by whether the hypermethylated occurred in HCC more rrequenuy man in tneir neignboring normal tissues, the hypermethylation status of the AR, DBCCR1, IRF7, OCT6, and p73 genes was considered as the event specific to the late stage, while that the rest that lacked such a distinguished contrast, as the event specific to the early stage of HCC carcinogenesis. Among all the clinical pathological parameters tested for theassociation with, the hypermethylation of the cyclin al gene was more prevalent in the non-cirrhosis group (P=0.02 1) while the hvoermethvlated p16^INK4a gene was more common in the cirrhosis group (P=0.017). The concordant dant methylation behaviors of nineteen genes, including the four previously studied and their association With clrrllosis has been evaluated by the best subgroup selection method. The data presented in this report would enable us toshape our understanding of the mechanisms for the HCC specific loss of the epigenetic stability of the genome, aswell as the strategy of developing the novel robust methylation based diagnostic and prognostic tools.     

cDNA array analysis of the differential expression change in virulence-related genes during the development of resistance in Candida albicans

Candida albicans is the most frequently isolated fungus in immunocompromised patients associated with mucosal and deep-tissue infections, To investigate the correlation between virulence and resistance on a gene expression profile in C. albicans, we examined the changes in virulence-related genes during the development of resistance in C, albicans from bone marrow transplant patients using a constructed cDNA array representing 3096 unigenes. In addition to the genes known to be associated with azole resistance,16 virulence-related genes were identified, whose differential expressions were newly found to be associated with the resistant phenotype. Differential expressions for these genes were confirmed by RT-PCR independently. Furthermore, the up-regulation of EFG1, CPH2, TEC1, CDC24, SAP10, ALS9, SNF1, SP072 and BDF1, and the down-regulation of RAD32, IPF3636 and UB14 resulted in stronger virulence and invasiveness in the resistant isolates compared with susceptible ones. These findings provide a link between the expression of virulence genes and development of resistance during C. albicans infection in bone marrow transplant (BMT) patients, where C. albicans induces hyphal formation and expression change in multiple virulence factors.