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Chaoneng Ji

Publications and source records attributed to Chaoneng Ji.

At least 19 recordsLinked to original sources
PubMed2007

Identification of genes differentially regulated by transcription factor, AP-2delta.

In an effort to better understand the transcriptional program regulated by transcription factor, AP-2delta, we used cDNA microarray to evaluate the relative expression of human genes in AD293 cells by exogenous expression of AP-2delta. Microarray showed 29 genes that were up-regulated and 39 genes with a down-regulated expression pattern. Among the identified genes were those encoding transcription factors, signal transduction molecules, kinases, as well as genes regulating cell growth, differentiation, and cell adhesion, a set of genes suggested to be regulated by AP-2. The results of microarray was validated for some regulated genes by real-time PCR analysis. These observations shed novel insight into the mechanism of AP-2delta action and provide a range of target genes for further investigation.

Binding Sites↗
PubMed2006

Different gene expression profiles of AD293 and HEK293 cell lines that show contrasting susceptibility to apoptosis induced by overexpression of Bim L.

Bim is a pro-apoptotic member of the Bcl-2 protein family. Overexpression of Bim proved to be highly cytotoxic for diverse cells. The AD293 cell line is derived directly from the HEK293 cell line but has been transfected with a gene that can improve cell adherence. We found that there was almost no apoptosis seen in Bim L-transfected AD293 cells, but more than half of Bim L-transfected HEK293 cells underwent apoptosis. Suppression subtractive hybridization was used to detect the different gene expression profile between these two cell lines. In 192 sequenced positive clones, there were 30 clones repeating twice or more. Ten genes were selected for identification by semi-quantitative RT-PCR. The transcripts of two adhesion-related genes (actin and parvin) and two apoptosis-related genes (cyclin 2 and protein phosphatase 1G) were up-regulated in AD293 cells. These results suggest that the high expression of cell adhesion-related proteins might be responsible for the different apoptosis status after the transfection of Bim L. Our data provide candidate genes responsible for the different apoptosis sensitivity of these two cell lines. Further investigation on the differential expression profile between AD293 and HEK293 might improve our understanding of cell apoptosis mechanism.

Actins↗
PubMed2006

Dual specificity phosphotase 18, interacting with SAPK, dephosphorylates SAPK and inhibits SAPK/JNK signal pathway in vivo.

The SAPK/JNKs play important roles in numerous cellular processes, and for this reason they have become putative drug targets. Most dual-specificity protein phosphatases (DSPs) play important roles in the regulation of mitogenic signal transduction and cell cycle control in response to extracellular stimuli. Dual-specificity phosphatase 18 (DUSP18), a newly recognized SAPK/JNK phosphatase, is widely expressed. This expression is modulated in response to extracellular stimuli. By phosphorylation assay, pull down and coimmunoprecipitation experiments, it is shown here that DUSP18 interacts with SAPK/JNK and dephosphorylates it both in vitro and in vivo. DUSP18 does not dephosphorylate p38 or p44ERK1. Furthermore, DUSP18 inhibits SAPK/JNK pathway in vivo. Based on these findings, DUSP18 appears to serve an important role by regulation of SAPK/JNK pathway.

Blotting, Western↗
PubMed2006

Cloning, characterization and subcellular localization of a gene encoding a human Ubiquitin-conjugating enzyme (E2) homologous to the Arabidopsis thaliana UBC-16 gene product.

Ubiquitin charging and activation of class III E2 enzymes has been directly linked to their nuclear import. It has not been published whether other classes E2s also abide by this mechanism. During the large-scale sequencing analysis of a human fetal brain cDNA library, we isolated a cDNA clone that is 2252 base pair in length, encoding a putative 162 amino acid protein, which shares high homology to Arabidopsis thaliana ubiquitin-conjugating enzyme 16 (Accession number NP_565110, 51% identity and 71% similarity) at protein level. Bioinformatics analysis revealed that the gene is composed of 7 exons, located on human chromosome 8q13-8q21.1, and that the predicted protein of the gene is a class I E2, for only composed of a conserved approximately 150-amino acid catalytic core, ubiquitin-conjugating enzyme E2 domain (UBC domain). In the C-terminal of the UBC domain sequence, there are two nuclear localization signals (NLSs). RT-PCR showed that this gene is ubiquitously expressed in 16 kinds of normal human tissues, but expression level is very low, unless in human heart, brain, liver, and pancreas. The subcellular localizations of the new human Ubiquitin conjugating enzyme E2 and its mutation were also examined, which showed that the nuclear localization of hUBC16 depended on two conditions: It has NLS, and at the same time, has enzyme active site, too, at least in HEK293 cells.

Amino Acid Sequence↗
PubMed2006

Crystal structures of human glycerol 3-phosphate dehydrogenase 1 (GPD1).

Homo sapiens L-alpha-glycerol-3-phosphate dehydrogenase 1 (GPD1) catalyzes the reversible biological conversion of dihydroxyacetone (DHAP) to glycerol-3-phosphate. The GPD1 protein was expressed in Escherichia coli, and purified as a fusion protein with glutathione S-transferase. Here we report the apoenzyme structure of GPD1 determined by multiwavelength anomalous diffraction phasing, and other complex structures with small molecules (NAD+ and DHAP) by the molecular replacement method. This enzyme structure is organized into two distinct domains, the N-terminal eight-stranded beta-sheet sandwich domain and the C-terminal helical substrate-binding domain. An electrophilic catalytic mechanism by the epsilon-NH3+ group of Lys204 is proposed on the basis of the structural analyses. In addition, the inhibitory effects of zinc and sulfate on GPDHs are assayed and discussed.

Catalysis↗
PubMed2006

Co-evolutionary analysis of insulin/insulin like growth factor 1 signal pathway in vertebrate species.

Insulin/insulin like growth factor 1 signaling pathway plays an essential role on the regulation of variant, interrelated and fundamental processes, such as metabolism, growth, reproduction and aging. This pathway is highly conserved during evolution. We analyzed eight vertebrate species' insulin/insulin like growth factor 1 pathway and examined the co-evolutionary relationship between different protein components by quantifying similarity of phylogenetic trees. The collected species include Takifugu rubripes, Danio rerio, Xenopus laevis, Gallus gallus, Mus musculus, Rattous norvegicus, Pan troglodytes and Homo sapiens. Results show that interacting proteins in this pathway share highly co-evolutionary relationship while contrarily the proteins without interaction have low co-evolutionary relationship. We also predict some receptor and ligand partners enjoy highly binding potential in corresponding species' pathway.

Animals↗
PubMed2006

Cloning and characterization of a novel KRAB-domain-containing zinc finger gene (ZNF284L).

The zinc finger gene (ZNF) family plays an important role in the regulation of transcription. This study reports the cloning and characterization of a novel human zinc finger protein cDNA (ZNF284L) from fetal brain cDNA library. The ZNF284L cDNA is 2223 bp in length encoding a 593-aa polypeptide. The protein contains a KRAB A+b box and eleven C2H2 type zinc finger motifs. ZNF284L gene is mapped to 19q13.2-19q13.3 with 5 exons, and the expression pattern of ZNF284L gene was also examined by reverse transcription polymerase chain reaction (RT-PCR). The transcripts were detected in the human lung, liver, pancreas, thymus, heart, placenta, spleen, prostate, ovary, small intestine and colon, but in human brain, skeletal muscle, kidney, testis and peripheral blood leukocyte, no expression was detected.

Amino Acid Sequence↗
PubMed2006

Purification and structural characterization of human ERp29.

ERp29 is a major resident of the endoplasmic reticulum (ER) and is postulated to play an important molecular chaperone role in most animal cells. Human ERp29 was isolated to homogeneity in high yield by using a bacterial expression system. Its secondary structure was studied by circular dichroism (CD), Fourier transformed infrared spectroscopy (FTIR) and Raman spectroscopy and it was found that human ERp29 comprises significant alpha-helical structure. The details of its temperature-induced conformational changes was studied by CD and FTIR for the first time, revealing that the protein is stable below 50 degrees C and has two distinct structural transitions between 50 degrees C and 70 degrees C. This may shed light on ERp29's inability to protect substrate proteins against thermal aggregation.

Amino Acid Sequence↗
PubMed2006

Human RAB24, interestingly and predominantly distributed in the nuclei of COS-7 cells, is colocalized with cyclophilin A and GABARAP.

As an atypical member of the Rab family, Rab24 has several attributes distinguishing this protein from the other members. Based on the yeast two-hybrid system, interaction between human RAB24 and two proteins, cyclophilin A (CyP-A) and gamma-aminobutyric acid type A receptor-associated protein (GABARAP), was detected and identified in COS-7 cells. Interestingly, RAB24 is predominantly localized in the nuclei of COS-7 cells, which is different from previous reports using other cell lines. RAB24 (D123I) can trigger the accumulation of intracellular inclusions, with small quantities of intranuclear inclusions in some cells. The GTPase activity of RAB24 and its two mutants was detected.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Characterization of a novel human protein phosphatase 2C family member, PP2Ckappa.

A novel member of human protein phosphatase 2C gene named PP2Ckappa was isolated from a human fetal brain cDNA library. The 2.0 kb cDNA encodes a 372 amino acid polypeptide with an intact protein phosphatase 2C (PP2C) catalytic domain. Reverse transcription-PCR (RT-PCR) revealed that the PP2Ckappa was widely expressed in normal human tissues. Transient transfection suggested that PP2Ckappa was localized in the nucleus in AD293 cells. Recombinant Trx-His-PP2Ckappa showed phosphatase activity toward p-nitrophenyl phosphate (pNPP), as well as oligopeptides containing phospho-threonine residues. Furthermore, the overexpression of PP2Ckappa distinctly activated the heat shock transcription factor pathway in eukaryotic cells.

Amino Acid Sequence↗
PubMed2005

Crystal structure of human guanosine monophosphate reductase 2 (GMPR2) in complex with GMP.

Guanosine monophosphate reductase (GMPR) catalyzes the irreversible and NADPH-dependent reductive deamination of GMP to IMP, and plays a critical role in re-utilization of free intracellular bases and purine nucleosides. Here, we report the first crystal structure of human GMP reductase 2 (hGMPR2) in complex with GMP at 3.0 A resolution. The protein forms a tetramer composed of subunits adopting the ubiquitous (alpha/beta)8 barrel fold. Interestingly, the substrate GMP is bound to hGMPR2 through interactions with Met269, Ser270, Arg286, Ser288, and Gly290; this makes the conformation of the adjacent flexible binding region (residues 268-289) fixed, much like a door on a hinge. Structure comparison and sequence alignment analyses show that the conformation of the active site loop (residues 179-187) is similar to those of hGMPR1 and inosine monophosphate dehydrogenases (IMPDHs). We propose that Cys186 is the potential active site, and that the conformation of the loop (residues 129-133) suggests a preference for the coenzyme NADPH over NADH. This structure provides important information towards understanding the functions of members of the GMPR family.

Amino Acid Sequence↗
PubMed2005

Identification and characterization of a novel Cut family cDNA that encodes human copper transporter protein CutC.

Copper is an essential heavy metal trace element that plays important roles in cell physiology. The Cut family was associated with the copper homeostasis and involved in several important metabolisms, such as uptake, storage, delivery, and efflux of copper. In this study, a novel Cut family cDNA was isolated from the human fetal brain library, which encodes a 273 amino acid protein with a molecular mass of about 29.3 kDa and a calculated pI of 8.17. It was named hCutC (human copper transporter protein CutC). The ORF of hCutC gene was cloned into pQE30 vector and expressed in Escherichia coli M15. The secreted hCutC protein was purified to a homogenicity of 95% by using the Ni-NTA affinity chromatography. RT-PCR analysis showed that the hCutC gene expressed extensively in human tissues. Subcellular location analysis of hCutC-EGFP fusion protein revealed that hCutC was distributed to cytoplasm of COS-7 cells, and both cytoplasm and nucleus of AD293 cells. The results suggest that hCutC may be one shuttle protein and play important roles in intracellular copper trafficking.

Amino Acid Sequence↗
PubMed2005

Isolation and expression pattern of RGS21 gene, a novel RGS member.

Regulators of G-protein signaling (RGS) proteins are known for the RGS domain that is composed of a conserved stretch of 120 amino acids, which binds directly to activated G-protein alpha subunits and acts as a GTPase-activating protein (GAP), leading to their deactivation and termination of downstream signals. In this study, a novel human RGS cDNA (RGS21), 1795 bp long and encoding a 152-amino acid polypeptide, was isolated by large-scale sequencing analysis of a human fetal brain cDNA library. Unlike other RGS family members, RGS21 gene has no additional domain/motif and may represent the smallest known member of RGS family. It may belong to the B/R4 subfamily, which suggests that it may serve exclusively as a negative regulator of alphai/o family members and/or alphaq/11. PCR analysis showed that RGS21 mRNA was expressed ubiquitously in the 16 tissues examined, implying general physiological roles.

Amino Acid Sequence↗
PubMed2005

Cloning and characterization of a novel RNA polymerase II C-terminal domain phosphatase.

Reversible phosphorylation of RNA polymerase (RNAP) II's largest subunit C-terminal domain (CTD) is a key event during mRNA metabolism. The CTD phosphatase, FCP1, catalyzes the dephosphorylation of RNAP II and is thought to play a major role in polymerase recycling. In this study, we isolated a novel phosphatase gene by large-scale sequencing analysis of a human fetal brain cDNA library. Its cDNA is 2215 bp in length, encoding a 318-amino acid polypeptide that contains a ubiquitin-like domain and a CTD phosphatase domain. Therefore, it was termed ubiquitin-like domain containing CTD phosphatase 1 (UBLCP1). Reverse transcription PCR (RT-PCR) revealed that UBLCP1 was expressed with relatively lower levels in most adult normal tissues and higher levels in fast growing or tumor tissues. Transient transfection experiment suggested that UBLCP1 was localized in the nucleus of COS-7 cells. Significantly, UBLCP1 could dephosphorylate GST-CTD in vitro. Accordingly, UBLCP1 may play a role in the regulation of phosphorylation state of RNA polymerase II C-terminal domain.

Amino Acid Sequence↗
PubMed2005

Molecular cloning and characterization of a novel adenylate kinase 3 gene from Clonorchis sinensis.

Adenylate kinase (AK) is a ubiquitous enzyme that contributes to the homeostasis of adenine nucleotides in living cells. AK catalyzes reversible high energy phosphoryl transfer reactions between ATP (or GTP) and AMP to generate ADP (or GDP). From a Clonorchis sinensis adult worm cDNA library, we isolated a cDNA clone encoding a novel AK3 isozyme. The 956 bp cDNA encodes a putative protein of 228 amino acids with a predicted molecular mass of 26.2 kDa. The recombinant CsAK3 protein produced in Escherichia coli can be refolded into a functional protein with AK3 activity. The optimum pH and temperature for the enzyme are 8.5 and 40 degrees C, respectively. The calculated activation energy is 56.04 kJ mol-1. The Km of the CsAK3 for AMP and GTP are 118 microM and 359 microM, respectively. CsAK3 is inhibited by Ap5A (>70% inhibition by 2.0 mM AP5A). Ap5A may be a potential lead compound acting on C. sinensis in which AK3 as a drug target.

Adenylate Kinase↗
PubMed2005

Molecular cloning and characterization of a novel human C4orf13 gene, tentatively a member of the sodium bile acid cotransporter family.

By large-scale sequencing analysis of a human fetal brain cDNA library, we isolated a novel human cDNA (C4orf13). This cDNA is 2706 bp in length, encoding a 340-amino-acid polypeptide that contains a typical SBF (sodium bile acid cotransporter family) domain and ten possible transmembrane segments. The putative protein C4orf13 shows high similarity with its orthologs in Mus musculus and Xenopus laevis. Human C4orf13 is mapped to chromosome 4q31.2 and contains 12 exons. RT-PCR analysis shows that human C4orf13 is widely expressed in human tissues, and the expression levels in liver and lung are relatively high, expression levels in placenta, kidney, spleen, and thymus are moderate, low levels of expression are detected in heart, prostate, and testis.

Amino Acid Sequence↗
PubMed2005

A novel zinc finger gene ZNF468 with two co-expressional splice variants, ZNF468.1 and ZNF468.2.

A novel human zinc finger protein encoding gene ZNF468 was obtained from a fetal brain cDNA library. By BLAST-N analysis we found two different splice variants. We termed the two splice variants ZNF468.1 and ZNF468.2. By BLAST search against the human genome database, ZNF468 was mapped to 19q13.4. The ZNF468.1 cDNA has four exons, and the ZNF468.2 cDNA has one more, between the third and fourth exon. This extra exon creates a difference between the deduced protein N-termini of the two splice variants. The ZNF468.1 cDNA is 3906 bp in length, encoding a 522a a protein, and ZNF468.2 is 4024 bp, encoding a 469-aa-protein. Both proteins contain 11 C2H2-type zinc finger motifs at their C-termini. The N-terminus of the deduced protein of ZNF468.1 has a well-conserved Krüppel-associated box (KRAB) domain that consists of KRAB boxes A and B, whereas the protein of ZNF468.2 does not have the KRAB domain. Tissue distribution of the ZNF468 gene indicates that the two splice variants are widely expressed in normal human tissues, except in heart and brain, and they are also co-expressional.

Alternative Splicing↗