In 2002, Chumakov and colleagues reported that, in 213 patients with schizophrenia and 241 normal subjects in Canada, there was a highly susceptible locus in chromosome 13 which is correlated with schizophrenia. There was a gene named
U87 cells were cultured in
The plasmids pcDNA4/TO/myc, His A (pcDNA4/TO) and pcDNA4/TO/myc, His A-
The U87 cells were seeded in a 100 mm plate and transfected with pcDNA4/TO and pcDNA4/TO/
The Human Whole Genome One Array version 6 (Phalanx BiotechGroup, Taiwan) contains 32,679 DNA oligonucleotide probes, and each probe is a 60-mer designed in the sense direction. Among the total number of probes, 31,741 correspond to the annotated genes in the RefSeq v51 and Ensembl version 65 database. In addition, 938 control probes are also included. The detailed descriptions of the gene array list are available from
Fluorescent aRNA targets were prepared from 1
Each single sample was at least performed twice in terms of technical or biological replicates under a reproducibility of more than 0.975. The signal intensity was loaded into Rosetta Resolver System (Rosetta Biosoftware, USA) to do data preprocessing and applied to 75 percentile centering normalization. The errors of the sample were estimated by using an error-weighted approach at the same time. Both fold change and
The genes upregulated by more than 2-fold and with the statistic
The differential expression affected by pLG72 overexpression in U87 cells.
Gene symbol | Fold change |
|
Gene symbol | Fold change |
|
Gene symbol | Fold change |
|
---|---|---|---|---|---|---|---|---|
DAOA |
99.9 | <2.80 |
TBC1D8B | 6.4 | 3.250 |
C5orf60 | 2.8 | 1.508 |
HNRNPCL1 | 22.2 | 3.287 |
CADPS2 | 5.7 | 5.697 |
ADAL | 2.8 | 3.527 |
FOXR2 | 14.4 | <2.80 |
LINC00028 | 4.7 | 5.597 |
SCUBE2 | 2.8 | 4.477 |
PANX2 | 11.7 | 7.113 |
IGSF9 | 3.8 | 0.000550514 | CYP26A1 | 2.6 | 5.026 |
GALNT3 | 10.9 | <2.80 |
TNFSF4 | 3.6 | <2.80 |
PAFAH1B2 | 2.5 | 5.294 |
RTN1 | 10.1 | <2.80 |
MXRA5 | 3.6 | 1.051 |
CHRNA6 | 2.5 | 4.314 |
LOC100287063 | 10.0 | 6.157 |
HSPA1B | 3.4 | 6.485 |
KLK3 | 2.4 | 2.067 |
TM4SF18 | 7.3 | 0.001431557 | LOC100130705 | 3.1 | <2.80 |
EFNB1 | 2.1 | 1.061 |
CNTD2 | 5.6 | 6.912 |
ACVR2B | 3.0 | <2.80 |
|||
RN7SK | 6.1 | 8.507 |
MAML2 | 3.0 | 1.129 |
|||
|
||||||||
NCF1B | −24.3 | <2.80 |
UPP1 | −2.8 | 1.229 |
HIPK3 | −2.6 | 6.306 |
WBP1 | −16.4 | 8.353 |
ALDH7A1 | −2.7 | 5.883 |
NR2C1 | −2.6 | 0.000500818 |
TOMM20L | −5.7 | <2.80 |
CLCN2 | −2.7 | 4.905 |
SCN1B | −2.6 | 0.00099985 |
ZNF618 | −5.7 | 5.221 |
ADAM9 | −2.7 | 1.640 |
PSMA1 | −2.6 | 2.490 |
LOC100133251 | −3.8 | 1.984 |
CYP1A2 | −2.7 | 2.620 |
TMEM216 | −2.6 | 1.675 |
MAF | −3.5 | 3.682 |
MFSD3 | −2.7 | 1.343 |
PLA2G5 | −2.6 | 8.703 |
ZPLD1 | −3.2 | 1.677 |
NDST4 | −2.7 | 9.920 |
KANSL3 | −2.5 | 1.840 |
OTOG | −3.0 | 5.315 |
HAAO | −2.7 | 2.481 |
TRIM40 | −2.5 | 1.702 |
SMOC2 | −3.0 | 1.154 |
ARL8A | −2.6 | 3.177 |
SAG | −2.5 | 5.333 |
CCDC58 | −3.0 | 5.483 |
C20orf202 | −2.6 | 6.332 |
FAM5C | −2.5 | 3.283 |
NTRK2 | −3.0 | 1.381 |
EDNRA | −2.6 | 2.281 |
PCK1 | −2.4 | 7.419 |
MIEN1 | −2.9 | 5.327 |
MTX3 | −2.6 | 9.082 |
NPHS2 | −2.3 | 4.093 |
MAP4K4 | −2.8 | 1.432 |
MIA2 | −2.6 | 4.305 |
The NOA (
U87 cells were transfected with pLG72 overexpression plasmids as described previously. After 48 hr, the medium was removed and the cells were treated with or without 10 mM of Tempol, a ROS scavenger, at 37°C for 1 hr. After removal of the medium, cells were incubated with fresh serum-free medium containing 20
U87 cells treated with or without 100
In order to investigate the biological functions/processes induced by pLG72 overexpression, the network generation tool, GeneMANIA, was employed. GeneMANIA was developed for biofunction predictions of favorite genes or gene sets based on Gene Ontology annotation patterns. GeneMANIA finds the genes related with a given input gene from public datasets including protein-protein interaction, genetic regulation, pathways, reactions, gene coexpression, protein colocalization, protein domain similarity, and phenotypic screening profiles. Additional genes to the input gene will be presented to make a complete and comprehensive network. The 27 genes upregulated and 38 genes downregulated by pLG72 overexpression were separately input into the GeneMANIA system (Figures
GeneMANIA analysis. The upregulated genes (a), downregulated genes (b), and total differentially expressed genes (c) were input into GeneMANIA server using the default setting.
The biological networks are usually used for prediction of biological pathways, annotation of biological functions, and/or identification of targets of diseases. Since GeneMANIA only gave a low-confidence clue of oxidative reaction, NOA, which applied the categories of Gene Ontology (GO) to network analysis [
The upregulated genes analyzed with Network Ontology Analysis.
Gene ontology |
|
Term name |
---|---|---|
Biological process | 4.3 |
|
4.3 |
|
|
8.4 |
|
|
0.0010 |
|
|
|
||
Cellular component | 0.0080 | Cytoplasmic cyclin-dependent protein kinase holoenzyme complex |
0.0080 | Cell outer membrane | |
0.0162 | Subsynaptic reticulum | |
0.0201 | Spindle midzone | |
|
||
Molecular function | 4.5 |
|
4.5 |
|
|
9.0 |
|
|
9.3 |
Steroid hormone receptor activity |
To demonstrate the induction of oxidative stress proposed above, U87 cells were transfected with pLG72 overexpression plasmids and the intracellular ROS was stained using DCFH-DA, the general ROS indicator. The fluorescent intensity was measured using flow cytometry. As compared with the control group of empty vector (EV), ROS level significantly increased 69.2% in U87 cells transfected with
The measurement of ROS production. The ROS production was detected using a fluorescent indicator, DCFH-DA, and monitored by flow cytometry. The ROS level was significantly induced by pLG72 and scavenged by Tempol. Ev, empty vector;
In 2013, Cheng and colleagues reported that 361 genes were differentially expressed in the brains of
In 2011, Otte and colleagues discovered that in
Cappelletti’s and Sacchi’s groups also transfected pLG72 into U87 cells [
The authors declare that there is no conflict of interests regarding the publication of this paper.
This work was supported by a grant from the Ministry of Science and Technology of Taiwan (NSC102-2628-B-039-008-MY3, NSC102-2622-B-039-001-CC3, and MOST 104-2632-B-039-002 to H.-T. Chang). The funders played no role in conducting this research or in paper preparation.