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In this study, a calibration model based on Near-infrared spectroscopy (NIR) technique and chemometrics method was developed for rapid and non-destructive detecting the polysaccharide contents of lentinula edodes samples collected from different regions. The polysaccharide contents of these samples were firstly determined by standard phenol-sulphruic acid method. Then, NIR spectra of these samples were collected by using Fourier transform infrared spectrometry. Based on these experimental data, a random forest method was further used to distinguish the regions of these samples, with a classification accuracy of 96.6%. After that, a rapid, accurate, and quantitative model was established for predicting the polysaccharide contents of these samples. In the model establishing process, some signal pre-treatment methods were optimized, and the validation results with highest determination coefficient (R2) and low root mean square errors of prediction (RMSEP) were, 0.925 and 0.720, respectively. These results showed that combined NIR technique with chemometrics was an effective and green method for lentinula edodes quality control.

The aim of this study was to investigate the genomic epidemiology of MRSA in China to identify predominantly linages and their associated antimicrobial resistance and virulence profiles. In this study, we conducted whole-genome sequencing (WGS) on 565 MRSA isolates from 7 provinces and municipalities of China between 2014 and 2020. MRSA isolates were subjected to MLST, spa typing staphylococcal cassette chromosome mec (SCC mec) typing, analysis of virulence determinants and antimicrobial susceptibility testing. Among 565 MRSA isolates tested, clonal complex (CC) 59 (31.2%), CC5 (23.4%) and CC8 (13.63%) were the major lineages, and the clonal structure was dominated by ST59-t437-IV (14.9%), ST239-t030-III (6.4%), ST5-t2460-II (6.0%). Of note, CC8, the predominant lineage in 2014-2015, was replaced by CC59 in 2016-2017 and since then alternately dominant with CC5. Interestingly, the complicated dynamic change of CC5 population structure was observed with ST764-t1084-II, ST5-t2460-II, ST764-t1084-II and ST764-t002-II successively dominant. Further analysis revealed that virulence determinant profiles and antibiograms were closely associated with the clonal lineage. The CC59 MRSA less resistance to most tested antimicrobial and relatively lower methicillin MIC, while ciprofloxacin, sulfamethoxazole/trimethoprim and rifampicin resistance was closely linked with CC8. MRSA isolates conservatively carried multiple virulence genes for capsule, exoenzymes, secretion, iron uptake and metabolism, biofilm formation and hemolysis. PVL encoding genes was markedly more common in ST338, CC30, CC398, ST8 and CC22, while tsst-1 was associated with ST5. In conclusion, the community-associated CC59-ST59-t437-IV lineage was found to be most predominant in China, with diverse clonal isolates alternately circulating in various geographical locations. Our study highlights the need for MRSA surveillance in China in order to monitor changes in MRSA epidemiology. Funding Information: This study was funded by grants from the Natural Science Fund of China (82072343). Declaration of Interests: The authors declare that there are no conflicts of interest.

In order to solve the problems of traditional basketball classroom teaching, a virtual simulation system for students' sports training is designed. Firstly, a sports capture system is set up to collect information and compare it with a standard database to judge the irregularities of athletes' movements, and then the athletes' movements are digitized and the skills are improved statistically in combination with the system functions. The human body model diagram is constructed and the characters are designed to obtain the virtual basketball stadium and the technical action virtual simulation results of the basketball players. Then, the motion capture technology is used to track the motion tracks of key information nodes in the virtual simulation results to obtain the motion tracks of leg bones in the training process of the basketball players. The system is conducive to acquiring knowledge and skills, realizing effective organization and management of teaching and training information, and providing a good learning and training environment for teaching and training.

Decreased autophagy is accompanied by the development of a myeloproliferative state or acute myeloid leukemia (AML). AML cells are often sensitive to autophagy‑inducing stimuli, prompting the idea that targeting autophagy can be useful in AML cytotoxic therapy. AML NB4 cells overexpressing microtubule-associated protein 1 light chain 3-green fluorescent protein were screened with 69 inhibitors to analyze autophagy activity. AML cells were treated with the polo-like kinase 1 (PLK1) inhibitors RO3280 and BI2536 before autophagy analysis. Cleaved LC3 (LC3-II) and the phosphorylation of mammalian target of rapamycin (mTOR), adenosine monophosphate-activated protein kinase, and Unc-51-like kinase 1 during autophagy was detected with western blotting. Autophagosomes were detected using transmission electron microscopy. Several inhibitors had promising autophagy inducer effects: BI2536, MLN0905, SK1-I, SBE13 HCL and RO3280. Moreover, these inhibitors all targeted PLK1. Autophagy activity was increased in the NB4 cells treated with RO3280 and BI2536. Inhibition of PLK1 expression in NB4, K562 and HL-60 leukemia cells with RNA interference increased LC3-II and autophagy activity. The phosphorylation of mTOR was reduced significantly in NB4 cells treated with RO3280 and BI2536, and was also reduced significantly when PLK1 expression was downregulated in the NB4, K562 and HL-60 cells. We demonstrate that PLK1 inhibition induces AML cell autophagy and that it results in mTOR dephosphorylation. These results may provide new insights into the molecular mechanism of PLK1 in regulating autophagy.

Objective: In this study, we aimed to clarify the effects of long noncoding ribonucleic acid prostrate androgen-regulated transcript-1 on bladder cancer cell proliferation and apoptosis. Methods: Microarrays were implemented to investigate the long noncoding ribonucleic acid expression profiles in bladder cancer tissue (N = 9) and in noncancer bladder tissue (N = 5). Relative prostrate androgen-regulated transcript-1 expression levels in tissue samples or cell lines were detected by real-time quantitative reverse transcription-polymerase chain reaction. Prostrate androgen-regulated transcript-1 expression was enhanced by the transfection of pcDNA3.1-prostrate androgen-regulated transcript-1 and downregulated by the infection with pcMV-sh prostrate androgen-regulated transcript-1. Additionally, cell proliferation and apoptosis were measured by the cell counting kit-8 assay and flow cytometry, respectively. Cell invasion was determined by a Transwell assay. Results: Prostrate androgen-regulated transcript-1 expression was upregulated in bladder cancer tissues compared to adjacent nontumor tissues. Furthermore, prostrate androgen-regulated transcript-1 levels were successfully upregulated by pcDNA3.1-prostrate androgen-regulated transcript-1 and depleted by pCMV-sh prostrate androgen-regulated transcript-1 in bladder cancer cell lines (5637, T24). Enhanced prostrate androgen-regulated transcript-1 expression promoted cell proliferation and invasion and inhibited cell apoptosis. However, knockdown of prostrate androgen-regulated transcript-1 expression inhibited cell proliferation and invasion and induced cell apoptosis. Conclusion: In summary, these data suggest that the knockdown of prostrate androgen-regulated transcript-1 represents a tumor suppressor player in bladder cancer and contributes to the inhibition of tumor proliferation, the promotion of cell apoptosis, and the suppression of cell invasion. Prostrate androgen-regulated transcript-1 may function as a new prognostic biomarker and as a feasible therapeutic target for patients with bladder cancer.

A review of valorization of oyster mushroom species and waste generated in the mushroom cultivation is presented, with a focus on the cultivation and valorization techniques, conditions, current research status and particularly the hazard mitigation and value-added recovery of the waste mushroom substrate (WMS) - an abundant waste in mushroom cultivation industry. Based on the studies reviewed, the production rate of the present mushroom industry is inadequate to meet market demands. There is a need for the development of new mushroom cultivation methods that can guarantee an increase in mushroom productivity and quality (nutritional and medicinal properties). This review shows that the cylindrical baglog cultivation method is more advantageous compared with the wood tray cultivation method to improve the mushroom yield and cost efficiency. Approximately 5 kg of potentially hazardous WMS (spreading diseases in mushroom farm) is generated for production of 1 kg of mushroom. This encourages various valorization of WMS for use in agricultural and energy conversion applications, mainly as biocompost, plant growing media, and bioenergy. The use of WMS as biofertilizer has shown desirable performance compared to conventional chemical fertilizer, whilst the use of WMS as energy feedstock could produce cleaner bioenergy sources compared to conventional fuels.

There is a need to develop better methods for epitope mapping and/or identification of antibody-recognizing motifs. Here, we describe improved biosynthetic peptide (BSP) method using a newly developed plasmid pXXGST-3 as vector, which has a viral E7 gene in the cloning sites of pXXGST-1. It is crucial to employ pXXGST-3 instead of pXXGST-1, since it makes use of the BSP method simpler and easier to perform, and more cost-effective for epitope mapping. These merits are embodied in two aspects: i) convenient recovery of double enzyme-digested product due to the existence of 315 bp inserted between BamH I and Sal I sites, and thus greatly reducing the production of self-ligation clones, and ii) no longer requiring control protein when screening recombinant (r-) clones expressing 8/18mer peptides by running polyacrylamide gel electrophoresis. The protocol involves the following core steps: (i) design of plus and minus strands of DNA fragments encoding overlapping 8/18mer peptides; (ii) chemical synthesis of the designed DNA fragments; (iii) development of r-clones using pXXGST-3 vector expressing each 8/18mer peptide fused with truncated GST188 protein; (iv) screening r-clones by running the cell pellets from each induced clone on SDS-PAGE gel followed by sequencing of inserted DNA fragments for each verified r-clone; and (v) Western blotting with either monoclonal antibodies or polyclonal antibodies. This improved GST188-BSP method provides a powerful alternative tool for epitope mapping.

In the light of the wide applications of a series of ephemerides developed by the Jet Propulsion Laboratory (JPL) of the National Aeronautics and Space Administration (NASA) of the USA to many aspects like the deep-space navigation and interplanetary exploration, it is necessary to make comparisons of their contents, applicable ranges, realizations and maintenances. In view of the fact that the explorations of the Venus and Mars are expected to carry out in China, their dynamical models and the adopted observational data are analysis by taking the DE405, DE421 and DE423 to be the examples. From the evaluations of the accuracies and performances of the above-mentioned ephemerides of different editions, their effects on the probes orbiting around the Venus or Mars are emphatically discussed, and a suggestion is given for their use that as far as the Venus missions are concerned, the use of the DE423 is recommend, and that the use of the DE421 or DE423 is recommended for the Mars Missions.

Bombyx mori cell line (BmN) comes fromBombyx moriovary cell subculture. In order to study the change of several genes transcription in the process ofBombyx moriovary cells primary culture and subculture, we usedBombyx moriovary organizations and BmN cell lines as research materials, used Real Time fluorescent quantitative RT-PCR to detect cyclin gene family (CyclinA, CyclinB, CyclinB3, CyclinE, CyclinL1), p53 and Telomerase genes transcription level in the ovary and BmN cell lines, and took Actin3 gene as reference to dispose the results. The results showed that in theBombyx moriBmN cell lines the expression of CyclinA, CyclinB, CyclinB3, CyclinE, CyclinL1 and Telomerase genes were higher than those in the ovary. The expression of CyclinB in the BmN was more then 3.8 which was 76 times higher than that in the ovary; The expression of p53 gene in the BmN cell was lower than that in the ovary; The expression of Telomerase gene in the BmN cell was higher than that in the ovary. The results accumulated a reliable data for further study on the the role of cyclin gene family, p53 gene, and Telomerase gene in the process ofBombyx moriovarian carcinoma.

Hybrid rice combination Longliangyou072 and hybrid rice Shenyou9576 were used to study the yield formation of ratoon crop and its relationship with the main crop.The result showed that among different nodes of Hybrid rice combination,the 2nd node from the top to the bottom showed the highest LAI and dry matter weight and the 4th node the lowest.There was significantly positive correlation between the yield of ratoon crop and the LAI of main crop in tillering stage,booting stage,full heading time and milk stage,the corresponding correlation coefficient were 0.864,0.896,0.935 and 0.941.The dry matter weight was the highest in milk stage,and the lowest in tillering stage for main crop and ratoon crop respectively.The dry matter weight of main crop in milk stage was significantly positively related with the yield of ratoon crop(r=0.451).Comparing to the main crop,root activity and photosynthesis rate were higher while chlorophyll content were lower in ratoon crop.Root activity of ratoon crop in booting stage and full heading time was very significantly positively related with the yield of ratoon crop,with correlation coefficient being 0.913 and 0.843 respectively.Chlorophyll content of ratoon crop in tillering stage,booting stage and milk stage was very significantly positively related with its yield,with correlation coefficient being 0.913,0.943 and 0.927 respectively.Photosynthesis rate of ratoon crop in tillering stage,full heading time and milk stage was significantly positively related with its yield,with correlation coefficient being 0.722,0.791 and 0.727 respectively.From full heading time to maturity,the ratooning bud generated from the 3rd node of main crop from different combination showed the largest mean length of bud.From twelve days after the full heading stage to maturity,viable bud rate was decreased along with the nodes from top to bottom.The percentage of 14C photosynthates distributed in panicle was the highest and the lowest in other leaves.The number of effective panicle and seed setting rate affected the yield of ratoon crop mostly,and the 2nd and 3rd nodes from the top to the bottom contributed 70% to the total yield.

To enable rational multi-epitope vaccine and diagnostic antigen design, it is imperative to delineate complete IgG-epitome of the protein. Here, we describe results of IgG-epitome decoding of three proteins from high-risk (HR-) oncogenic human papillomavirus type 58 (HPV58). To reveal their entire epitomes, employing peptide biosynthetic approach, 30 precise linear B-cell epitopes (BCEs) were mapped on E6, E7 and L1 proteins using rabbits antisera to the respective recombinant proteins. Using sequence alignment based on BCE minimal motif, the specificity and conservativeness of each mapped BCE were delineated mainly among known HR-HPVs, including finding 3 broadly antibody cross-reactive BCEs of L1 that each covers almost all HR-HPVs. Western blots revealed that 13 of the 18 BCEs within L1-epitome were recognized by murine antisera to HPV58 virus-like particles, suggesting that these are antibody accessible BCEs. Also, a highly conserved epitope (YGD/XTL) of E6 was found to exist only in known common HR-HPVs, which could be used as the first peptide reference marker for judging HR-HPVs. Altogether, this study provides systemic and exhaustive information on linear BCEs of HR-HPV58 that will facilitate development of novel multi-epitope diagnostic reagents/chips for testing viral antibodies and ‘universal’ preventive HPV peptide vaccine based on L1 conserved BCEs.

Human NFB repressing factor (NKRF) is a negative regulation transcription factor, which is able to repress transcription by binding to the negative regulatory element (NRE) near the NFκB binding site in certain genes’ promoters. Current researches reveals that NKRF represses the activation of IFN-, IL-8, hiNOS and HIV-1 by NFκB. We used optical fiber beadchip to analysis the different gene expression patterns of RNAi mediated NKRF knockdown HEK293 cells and found that several genes showed significant change of expression levels. Real-time PCR was performed to verify the changes of expression of candidate genes. We analyzed the function of candidate genes by searching the gene ontology databases and publications and revealed that these genes functioned in cell cycle, cell proliferation, apoptosis, cell migration, DNA repair, transcription, metabolism, response to stimulus and signal transduction. This study provides new perspectives on NKRF’s potential multiple functions.

Escherichia coli is widely used for expressing recombinant proteins, and several tags have been developed to improve protein solubility. However, expressing and purifying protein from other organisms is not always successful. In this study, we investigated the possibility of using TtrD as an expressing fusion tag in E. coli. Twenty RING finger domain containing human genes were expressed in E. coli grown at 37 °C and 18 °C and tested with four other fusion tags, namely His, SUMO, GST and MBP, for comparison. The results indicated that the soluble expressing ability of the tags was MBP, GST, TtrD, SUMO, and His in descending order. A one-column refolding process was used to purify the expressed proteins in inclusion bodies, and TtrD showed the strongest refolding ability. The results suggested that the TtrD tag enhanced recombinant protein solubility and refolding ability and might be a useful tag for protein expression in E. coli.

The steroid-5-alpha-reductase, alpha polypeptide 2 (SRD5A2) gene plays a crucial role in androgen metabolism pathway in human prostate. It encodes SRD5A2 enzyme, which catalyses testosterone to dihydrotestosterone (DHT). DHT is the main active structure binding with androgen receptor (AR). After the activation of AR, it further regulates a series of target genes in androgen metabolism pathway. However, no clear consensus has been reached on the association between the SRD5A2 V89L, A49T and TA repeat polymorphisms and prostate cancer (PCa) risk. Thus, we performed a meta-analysis of 31 association studies with 14 726 PCa cases and 15 802 controls. We found no association between PCa and 89L compared with 89Vallele [odds ratio (OR) = 1.02, 95% confidence interval (CI) 0.98― 1.06, P heterogeneity = 0.44]. The 49T allele showed a significantly elevated effect on the high stage (Stages III―IV) of PCa risk both under the dominant genetic model (OR = 2.13, 95% CI 1.44―3.15, P heterogeneity = 0.65) and in the contrast T versus A allele (OR = 2.06, 95% CI 1.41―3.02, P heterogeneity = 0.69). There was a significantly decreased association between PCa and long TA repeat as compared versus short TA repeat (OR = 0.86, 95% CI 0.74―1.00, P hetero g eneit y = 0.79). No significant between-study heterogeneity was found in all subjects under four genetic models (dominant model, recessive model, allele comparison and homozygosity comparison) for these three polymorphisms, respectively, so the fixed effects model was used to pool the result. Our result indicated that carriers of 49T might improve the risk of PCa in higher stages (Stages III―IV), carriers of long TA repeat might decrease the risk of PCa and 89L may not be an important risk factor for PCa. However, due to the limited sample sizes, this meta-analysis did not achieve sufficiently conclusive results. Still more well-designed studies should be performed to clarify the role of these three polymorphisms in the development of PCa.

LNX protein is the first described PDZ domain-containing member of the RING finger-type E3 ubiquitin ligase family. Studies have approved that LNX could participate in signal transduction, such as Notch pathway, and play an important role in tumorigenesis. In this study, we found that down-regulation of LNX resulted in G0/G1 cell cycle arrest in G0/G1 phase in HEK293 cells. To explore the molecular mechanism of this phenomenon, we employed expression microarray to comparatively analyze the genome-wide expression between the LNX-knockdown cells and the normal cells. We also used quantitative real-time PCR to further confirm the differential expression patterns of 25 transcripts involved in cell cycle. Combined with known information about genic functions, signal pathways and cell cycle machinery, we analyzed the role of endogenous LNX in cell cycle. The results suggest that down-regulation of LNX could result in cell cycle arrest in G0/G1 phase through inhibition of β-catenin, MAPK, NFκB, c-Myc-dependent pathway and activation of p53, TGF-β-dependent pathway. This study provides new perspectives on LNX's pleiotropic activities, especially its essential role in cell proliferation and cell cycle.

In order to illustrate the roles played by Pro40 in the structure, properties and functions of Cytochrome b5, three mutated genes, P40V, P40Y, P40G were constructed in this work. Only the P40V gene was successfully expressed into holoprotein in E. coli JM83. According to the results of X-ray crystallographic analysis and various kinds of spectroscopy studies, it is evident that substituting valine for Pro40 does not result in significant alterations in the protein′s overall structure; however, local conformational perturbations in the proximity of the heme do occur. The redox potential of the P40V mutant is 40 mV lower than that of the wild type protein. Its stability towards heat, urea, acid and ethanol were significantly decreased. The mutation leads to a decrease in the hydrophobicity of the heme pocket, which is probably the major factor contributing to the above changes. Binding constants and electron transfer rates between cytochrome bs and cytochrome c were determined using UV-visible spectroscopy and stopped-flow techniques for both the wild type and the mutant. The results showed that the substitution of Pro40 by valine does not influence the binding constant of cytochrome b5 to cytochrome c; however, the electron transfer rate between them decreased significantly. This indicates that proline-40 is essential to maintaining cytochrome bss stability and its electron transfer with cytochrome c. These studies also provided a good example that property and functional changes of a protein do not necessarily require large overall structural alterations; in most cases, only perturbations on the local conformations are sufficient to induce significant changes in protein′s properties and functions.

The purpose of this study was to identify and validate novel prognostic biomarkers in human hepatocellular carcinoma (HCC). We analyzed gene expression profiles not only between 33 HCCs and their corresponding noncancerous liver tissues, but also between 25 HCCs and pooled normal liver tissues using cDNA microarrays containing 12800 genes. Functional analysis of differentially expressed genes involved in HCC carcinogenesis and tumor progression revealed that up-regulated and down-regulated genes are mainly associated with cell cycle and immune response, respectively. We detected two regions of cytogenetic changes only in poorly-differentiated HCCs using the expression data. We identified a 9-gene expression signature, which was able to predict differentiation degree and survival of HCC samples. Among the 9 most discriminatory genes, minichromosome maintenance protein 2 (MCM2), a significantly up-regulated gene involved in cell cycle pathway, was selected for further analysis. Overexpression of MCM2 protein related to poor-differentiation in HCC was validated using tissue microarray-based immunohistochemistry containing 96 HCCs. Our studies show that the 9-gene expression signature may serve as promising prognostic biomarkers involved in hepatocarcinogenesis and tumor progression.

Polymorphisms in methionine synthase (MTR) gene may be involved in carcinogenesis by affecting DNA methylation. However, association studies on MTR A2756G polymorphism in cancers have reported conflicting results. Therefore we performed a meta-analysis to better assess the associations. A total of 24 896 cancer patients and 33 862 controls from 52 articles for MTR A2756G were investigated. Overall, individuals carrying MTR 2756GG genotype had a subtly reduced cancer risk under a recessive genetic model (odds ratio (OR), 0.92; P=0.053; 95% confidence interval (95% CI), 0.84-1.00; I(2)=0.0%; P(heterogeneity)=0.61). In the subgroup analyses by ethnicity, 2756GG was associated with a significantly reduced cancer risk in European populations (OR, 0.83; P=0.001; 95% CI, 0.74-0.93; I(2)=0.0%; P(heterogeneity)=0.99). However, in Asian populations, a significantly elevated association between 2756GG genotype and cancer risk was observed (OR, 1.33; P=0.012; 95% CI, 1.06-1.65; I(2)=0.0%; P(heterogeneity)=0.50). In studies stratified by tumor site, there was a significantly reduced risk of acute lymphoblastic leukemia (ALL) (OR, 0.54; P=0.049; 95% CI, 0.29-1.00; I(2)=10.7%; P(heterogeneity)=0.33) and colorectal cancer (OR, 0.63; P=0.004; 95% CI, 0.47-0.87; I(2)=0.0%; P(heterogeneity)=0.73) in European populations. Our study indicates that MTR A2756G polymorphism is a candidate gene polymorphism for cancer susceptibility regardless of environmental factors. Large-scale, well-designed, and population-based studies are required to further investigate gene-gene and gene-environment interactions on MTR A2756G polymorphism and tissue-specific cancer risk in an ethnicity-specific population.

The solution structure of oxidized bovine microsomal cytochrome b(5) mutant (E48, E56/A, D60/A) has been determined through 1524 meaningful nuclear Overhauser effect constraints together with 190 pseudocontact shift constraints. The final family of 35 conformers has rmsd values with respect to the mean structure of 0.045+/-0.009 nm and 0.088+/-0.011 nm for backbone and heavy atoms, respectively. A characteristic of this mutant is that of having no significant changes in the whole folding and secondary structure compared with the X-ray and solution structures of wild-type cytochrome b(5). The binding of different surface mutants of cytochrome b(5) with cytochrome c shows that electrostatic interactions play an important role in maintaining the stability and specificity of the protein complex formed. The differences in association constants demonstrate the electrostatic contributions of cytochrome b(5) surface negatively charged residues, which were suggested to be involved in complex formation in the Northrup and Salemme models, have cumulative effect on the stability of cyt c-cyt b(5) complex, and the contribution of Glu48 is a little higher than that of Glu44. Moreover, our result suggests that the docking geometry proposed by Northrup, which is involved in the participation of Glu48, Glu56, Asp60, and heme propionate of cytochrome b(5), do occur in the association between cytochrome b(5) and cytochrome c.

Most members of the type-2 phosphatidic acid phosphatase (PAP2) superfamily are integral membrane phophatases involved in lipid-related signal transduction and metabolism. Here we describe the cloning of a novel gene from Geobacillus toebii T-85, encoding a PAP2-like protein, Gtb PAP2L2, which contains 212 amino acids and shows a limited homology to other known PAP2s, especially at conserved phosphatase motifs, and a similar six-transmembrane topology structure. This enzyme was expressed, and purified in Escherichia coli. Recombinant Gtb PAP2L2s from the membrane fractions were solublized with 0.3% (v/v) Triton X-100 and purified by Ni(2+) affinity chromatography. The purified enzyme showed broad substrate specificity to phosphatidic acid, diacylglycerol pyrophosphate, and lysophosphatidic, but preferred phosphatidic acid and diacylglycerol pyrophosphate in vitro. Gtb PAP2L2 is a thermal stable enzyme with a half-life of 30 min at 60 degrees C. The enzyme was strongly inhibited by 1% SDS, 10 mM veranda, and Zn(2+), whereas it was independent of the Mg(2+) ion, and insensitive to N-ethylmaleimide. The purified recombinant Gtb PAP2L2 was catalytically active and highly stable, making it ideal as a candidate on which to base further PAP2 structure/function studies.

BACKGROUND Prostate cancer is a common disease among men but the knowledge of the prostate carcinogenesis is still limited. METHODS cDNA microarray-based comparative genomic hybridization (CGH) and expression profiling were performed to screen the genomic and the expression changes in prostate cancer respectively. The two data were integrated to study the influence of genomic aberrations on gene expression and seek for the genes with their expression affected by the genomic aberrations. Real-time PCR was performed to evaluate the array data. RESULTS Array-based CGH detected gains at 2q, 3p/q, 5q, 6q, 8q, 9p, 10p/q, 11q, 12p, 14q, and 19p/q and losses at 1p, 2p, 4q, 6p/q, 7p, 11p/q, 12q, 17p/q, 19p/q, and Xp/q in more than 20% prostate tumors and narrowed these aberrations. For example, the gain of 8q was mapped to five minimal regions. Novel aberrations were also identified, such as loss at Xq21.33-q22.2. Expression profiling discovered the significant biological processes involved in the prostate carcinogenesis, such as exogenous antigen presentation via MHC class II and protein ubiquitination. Integration analysis revealed a weak positive correlation between genomic copy number and gene expression level. Fifty-three genes showed their expression directly affected by the genomic aberrations possibly, including more than one member of Ras superfamily and major histocompatibility complex (MHC). These genes are involved in multiple biological processes. CONCLUSIONS Integration of the CGH and expression data provided more information than separate analysis. Although the direct influence of genomic aberrations on gene expression seems weak, the influence can be extended by indirect regulation through a few directly affected genes. Because the influence can be persistent, the genes directly affected by the genomic aberrations may play key roles in the prostate carcinogenesis and are worth further analysis. Prostate 68: 1496–1509, 2008. © 2008 Wiley-Liss, Inc.

Ubiquitin and ubiquitin-like proteins are known to be covalently conjugated to a variety of cellular substrates via a three-step enzymatic pathway. These modifications lead to the degradation of substrates or change its functional status. The ubiquitin-activating enzyme (E1) plays a key role in the first step of ubiquitination pathway to activate ubiquitin or ubiquitin-like proteins. Ubiquitin-activating enzyme E1-domain containing 1 (UBE1DC1) had been proved to activate an ubiquitin-like protein, ubiquitin-fold modifier 1 (Ufm1), by forming a high-energy thioester bond. In this report, UBE1DC1 is proved to activate another ubiquitin-like protein, SUMO2, besides Ufm1, both in vitro and in vivo by immunological analysis. It indicated that UBE1DC1 could activate two different ubiquitin-like proteins, SUMO2 and Ufm1, which have no significant similarity with each other. Subcellular localization in AD293 cells revealed that UBE1DC1 was especially distributed in the cytoplasm; whereas UBE1DC1 was mainly distributed in the nucleus when was cotransfected with SUMO2. It presumed that UBE1DC1 greatly activated SUMO2 in the nucleus or transferred activated-SUMO2 to nucleus after it conjugated SUMO2 in the cytoplasm.

A gene-encoding alkaline phosphatase (AP) from thermophilic Geobacillus thermodenitrificans T2, termed Gtd AP, was cloned and sequenced. The deduced Gtd AP protein comprises 424 amino acids and shares a low homology with other known AP (35% identity), while it exhibits the conservation of the active site and structure element of Escherichia coli AP. The Gtd AP protein, without a predicted signal peptide of 30 amino acids, was successfully overexpressed in E. coli and purified as a hexa-His-tagged fusion protein. The pH and temperature optima for purified enzyme are 9.0 and 65 degrees C, respectively. The enzyme retained a high activity at 45-60 degrees C, while it could be quickly inactivated by a heat treatment at 80 degrees C for 15 min, exhibiting a half-life of 8 min at 70 degrees C. The K(m) and V (max) for pNPP were determined to be 31.5 muM and 430 muM/min at optimal conditions. A divalent cation is essential, with a combination of Mg2+ and Co2+ or Zn2+ preferred. The enzyme was strongly inhibited by 10 mM ethylenediaminetetraacetic acid (EDTA) and vanadate but highly resistant to urea and dithiothreitol. The properties of Gtd AP make it suitable for application in molecular cloning or amplification.

We conducted a meta-analysis to assess the association between tumor necrosis factor-alpha (TNF-alpha) gene TNFA-308 (G > A) and TNFA-857 (C > T) polymorphisms and gastric cancer (GC) susceptibility. We also performed subgroup analyses based on ethnicity (Caucasian, east Asian, and other populations) and tumor location [noncardia gastric cancer (NCGC)]. There were 3,335 GC patients and 5,286 controls for TNFA-308, and 1,118 GC patients and 1,591 controls for TNFA-857 in our analysis. Overall, allele contrast (A vs. G) of TNFA-308 polymorphism produced significant results in worldwide populations [Pheterogeneity = 0.05, random-effects (RE) odds ratio (OR) 1.19; 95% confidence interval (CI) 1.03–1.37, P = 0.02] and Caucasian populations (Pheterogeneity = 0.15, fixed-effects (FE), OR 1.27; 95% CI 1.11–1.45, P = 0.0005). Similar results were also obtained in recessive models and homozygote contrasts. No significant association was observed in NCGC and east Asian subgroup analysis. T variant of TNFA-857 produced significant results only in allele contrast (Pheterogeneity = 0.38, FE OR 1.17; 95% CI 1.01–1.35, P = 0.04). In conclusion, TNFA-308 locus of TNF-alpha would be a risk factor for GC, especially in Caucasian populations. Besides, TNFA-857 locus may be related to GC risk, which demonstrated changeability of results in different contrasts.

ERCC1 (excision repair cross complementation group 1) is a subunit of the nucleotide excision repair complex, which can perform DNA strand incision correction of DNA damage. Association studies on the ERCC1 polymorphisms (C8092A and T19007C) in cancer had shown conflicting results. We performed a meta-analysis from all eligible case-control studies to assess the purported associations. Overall, the 19007C allele (3 853 patients and 4 349 controls) showed no significant effect on cancer risk compared to 19007T allele (P=0.39, odds ratio (OR)=0.95; 95% confidence interval (CI) 0.85-1.06, P(heterogeneity)=0.001) in all subjects. Meta-analysis under other genetic contrasts did not reveal any significant association of T19007C to cancer in all subjects, Caucasians and Asians. The 19007C allele (2 279 patients and 2 808 controls) showed no significant effect on lung cancer risk compared to 19007T allele (P=0.72, OR=0.94, 95% CI 0.69-1.29, P(heterogeneity)=0.0001) in all subjects. No significant effect of 8092A allele (3 865 patients and 3 750 controls) on cancer risk in all subjects (P=0.85, OR=1.01, 95% CI 0.94-1.08, P(heterogeneity)=0.92) and in Caucasians and Asians compare to 8092C. No evidences of association of C8092A (501 patients and 620 controls) to squamous cell carcinoma were found. The accumulated evidence indicated ERCC1 T19007C and C8092A might not be risk factors for cancer. Significant between-study heterogeneity existed in T19007C, which arose from a study showing significant protecting effect of 19007C allele compare to 19007T allele in smokers. More studies based on larger, stratified case-control population should be required to further evaluate the role of ERCC1 C8092A and T19007C polymorphisms in different cancer, especially in smokers.

Phosphorylation status of RNA polymerase (RNAP) II's largest subunit C-terminal domain (CTD) plays an important role during transcription cycles. The reversible phosphorylation mainly occurs at serine 2 and serine 5 of CTD heptapeptide repeats and regulates RNAP II's activity during transcription initiation, elongation and RNA processing. Here we expressed and characterized HSPC129, a putative human protein bearing a CTD phosphatase domain (CPD). PCR analysis showed that it was ubiquitously expressed. HSPC129DeltaTM, the truncate HSPC129 with first 156 N terminal amino acids deleted, exhibited Mg(2+) dependent phosphatase activity at pH 5.0. Its specific CTD phosphatase activity was verified in vitro. Our research suggests that HSPC129 may regulate the dynamic phosphorylation of RNAP II CTD.

Uracil phosphoribosyltransferase, which catalyzes the conversion of uracil and 5-phosphoribosyl-1-R-diphosphate to uridine monophosphate, is important in the pyrimidine salvage pathway and is an attractive target for rational drug design by incorporation of prodrugs that are lethal to many parasitic organisms specifically. So far, uracil phosphoribosyltransferase has been reported in Arabidopsis thaliana only, not in mammals. In this study, a novel uracil phosphoribosyltransferase family cDNA encoding a 309 amino acid protein with a putative uracil phosphoribosyltransferase domain was isolated from the human fetal brain library. It was named human UPRTase (uracil phosphoribosyltransferase). The ORF of human UPRTase gene was cloned into pQE30 and expressed in Escherichia coli M15. The protein was purified by Ni-NTA affinity chromatography, but UPRTase activity could not be detected by spectrophotometry. RT-PCR analysis showed that human UPRTase was strongly expressed in blood leukocytes, liver, spleen, and thymus, with lower levels of expression in the prostate, heart, brain, lung, and skeletal muscle. Subcellular location of UPRTase-EGFP fusion protein revealed that human UPRTase was distributed in the nucleus and cytoplasm of AD293 cells. Evolutional tree analyses of UPRTases or UPRTase-domain-containing proteins showed that UPRTase was conserved in organisms. UPRTases of archaebacteria or eubacterium had UPRTase activity whereas those higher than Caenorhabditis elegans, which lacked two amino acids in the uracil-binding region, had no UPRTase activity. This means that human UPRTase may have enzymatic activity with another, unknown, factor or have other activity in pyrimidine metabolism.

Ubiquitination, the covalent attachment of the protein ubiquitin (Ub) to other cellular proteins, has been implicated in a number of important physiological processes. ubiquitin conjugating enzyme (E2) plays an important role in the ubiquitin system. Here we report the research about a human putative ubiquitin conjugating enzyme gene, UBE2Z. The length of UBE2Z cDNA is 3054 base pairs and contains an open reading frame encoding 246 amino acids. The UBE2Z gene was mapped to human chromosome 17q21.32 and consisted of 6 exons. RT-PCR showed that UBE2Z was widely expressed in human tissues, especially high in placenta, pancreas, spleen and testis. The UBE2Z-GFP fusion protein was located in both nucleus and cytoplasm of AD293 cells.

Human protein kinases make up a large superfamily of homologous proteins, which are related by virtue of their kinase domains (also known as catalytic domains). Here we report the cloning and characterization of a novel human MAST4 (microtubule associated serine/threonine kinase family member 4) gene, which locates on human chromosome 5q13. The MAST4 cDNA is 7587 base pairs in length and encodes a putative protein of 2435 amino acids which contains a serine/threonine kinase domain and a PDZ domain. MAST4 protein has 64%, 63%, 59% and 39% identical aminoacid residues with MAST1, MAST2, MAST3 and MASTL respectively. RT-PCR analysis revealed relatively high expression level of MAST4 in most normal human tissues, with an exception of in testis, small intestine, colon and peripheral blood leukocyte.

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.

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.

Pre-mRNA splicing allows individual genes to produce multiple protein isoforms with diverse functions. Recognition of functional splice sites in pre-mRNAs is very important in this splicing process and requires some protein auxiliary factors such as U2 small nuclear ribonucleoprotein auxiliary factor small subunit (U2AF35, encoded by U2AF1). By its RNA binding domains, U2AF35 interacts with U2AF65 to bind 3' splice site of pre-mRNA and initiates splicing. Another protein, which is named as U2AF1-like3 (U2AF1L3), shows high similarity with U2AF35 and may have related function in pre-mRNA splicing. Here, we report a splice variant of U2AF1L3, which is 767 bp in length and has an open reading frame (ORF) coding a predicted 181 amino acids protein. Reverse transcription-PCR (RT-PCR) shows that this isoform has different expression pattern with U2AF1L3 and is highly expressed in heart, brain and lung.

Ubiquitin and other ubiquitin-like proteins play important roles in post-translational modification. They are phylogenetically well-conserved in eukaryotes. Activated by other proteins, ubiquitin and ubiquitin-like proteins can covalently modify target proteins. The enzymes responsible for the activation of this modification have been known to include UBA1, SAE2, UBA3, SAE1 and ULA1. Here we report a new ubiquitin activating enzyme like cDNA, named ubiquitin activating enzyme E1-domain containing 1 (UBE1DC1), whose cDNA is 2654 base pairs in length and contains an open reading frame encoding 404 amino acids. The UBE1DC1 gene consists of 12 exons and is located at human chromosome 3q22. The result of RT-PCR showed that UBE1DC1 is expressed in most of human tissues.

Biofilm formation is an important phenotype associated with chronic Pseudomonas aeruginosa infections. In the present study, a total of 48 P. aeruginosa strains isolated from clinical specimens were examined for their biofilm-forming ability using a microtiter plate method. The different biofilm-forming abilities were demonstrated among the strains; however, most strains formed a larger biofilm than strain PAO1, a reference strain. The genetic typing was also carried out by enterobacterial repetitive intergenic consensus-based polymerase chain reaction. Although they were divided into five groups (A to E), most of the strains showing the higher biofilm-forming ability were found to be in groups D and E, suggesting a significant relationship between the biofilm-forming ability and the genetic group.

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.

Gliomas are major tumors of the central nervous system with a wide spectrum of different tumor types. Ligand of Numb protein X (LNX) is PDZ domain containing protein that interacts with cell fate determinant Numb. cDNA microarray analysis was used to determine the expression of 13,939 genes in a set of 18 gliomas. It showed that human LNX was downregulated in 100% of gliomas including low- and high-grade ones, which was confirmed by Northern blot. In situ hybridization analysis revealed that LNX was lowly expressed in cytoplasm of glioma cells. Thus, LNX might act as a diagnostic marker and a potential therapeutic target for glioma. Two-hybrid screen in yeast was used to identify human LNX interacting proteins important for LNX function. It showed that human LNX interacted with Ski interacting protein (SKIP) via PDZ domains. The co-immunoprecipitation results suggested that LNX interacted with SKIP in HEK293 cells. LNX could affect the subcellular localization of Numb, which indicated that LNX might function as a molecular anchor that localized Numb to the subcellular site of its interaction with Notch. The presence of multiple protein binding domains involved in signal transduction and interaction with Numb and SKIP suggested an important role for LNX in tumorogenesis.

The short-chain dehydrogenases/reductases (SDRs) play an important role in the body's metabolism. We have cloned a novel mouse SDR cDNA, which encodes a deduced HSD-like protein with a conserved SDR domain and an SCP2 domain. The 1.8 kb cDNA consists of 11 exons and is mapped to mouse chromosome 4B3. The corresponding gene is widely expressed in normal mouse tissues and its expression level in the liver increases after inducement with cholesterol food. The predicted mouse HSDL2 protein, which has a peroxisomal target signal, is localized in the cytoplasm of NIH 3T3 cells.

Rheumatoid arthritis (RA) is a common autoimmune disease. Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) is a highly suspected candidate gene for RA susceptibility. However, association studies on the polymorphism of CTLA-4 exon-1 +49A/G in RA have shown conflicting results. Therefore, we performed a meta-analysis to better assess the purported association. In order to look for ethnic effect, we performed subgroup meta-analysis in populations of European descent and Asian descent. Meta-regression analysis was also performed to explore the possible heterogeneity between the two subgroups. Ten studies (11 comparisons) with the CTLA-4 exon-1 +49A/G genotyping on 2,315 patients with RA and 2,536 controls were selected for our meta-analysis. Overall, the fixed-effects odds ratio (OR) for the G versus A allele was 1.11 (P=0.02, 95% confidence interval (CI) 1.02-1.21), with no between-study heterogeneity. Subgroup and meta-regression analysis according to the ethnicity (European or Asian) demonstrated different scenarios concerning the CTLA-4 exon-1 +49A/G polymorphism's role in RA susceptibility for the two different subgroups. No effect of G on susceptibility was seen in European descent (five comparisons; OR=1.04, P=0.30, 95% CI 0.95-1.19; no significant between-study heterogeneity). However, there is a significant association in Asian descent under both fixed [OR=1.21, 95% CI (1.06-1.39), P=0.005] and random-effect models [OR=1.19, 95% CI (1.01-1.42), P=0.04]. Meta-regression analysis also supports the heterogeneity between the two subgroups (P=0.082). We also explored the role of this polymorphism on RA risk under other various interested genetic contrasts. These results further support that this polymorphism could not be a risk factor for Europeans. Interestingly, we find that in Asians the G allele has a greater tendency to cause RA in a recessive genetic model. However, sensitivity analysis showed that the combined result of Asian populations was unstable. In conclusion, our meta-analysis results suggest that CTLA-4 exon-1 +49G allele would not be a risk factor for RA in Europeans but might play a role in RA susceptibility for Asians.

XRIP alpha was identified as an adapter protein involved in RAP nuclear import. Several homologs were reported in mammal EST analysis, but the expression pattern and genomic organization of hRIP isoforms were not clarified yet. We isolated nine isoforms of hRIP from a premade human fetal brain library. hRIP alpha is the longest isoform with 219 residues, containing a N-terminal arginine-rich basic region, followed by an acidic region and two C-terminal Zn finger-like structures. hRIP beta deletes one Zn-finger-like structure. Three hRIP alpha isoforms and four hRIP sigma isoforms express truncated proteins due to frame shift. hRIP gamma isoforms lost the C-terminal Zn-finger-like structure. hRIP delta isoforms only contain the N-terminal arginine-rich basic region and the core sequence of the acidic region. The genomic organization of hRIP was identified by bioinformatic analysis. hRIP, containing seven exons, is located at human chromosome 17p13. hRIP was expressed in all 16 detected human tissues with a similar pattern. All EGFP-hRIP fusion proteins were located at the nucleus in the HEK293 cell. The two-polar molecular structure of hRIP might be involved in the basic cell function, and plays a role in the alternative nuclear ingress.

Mouse U26 has been defined as a 2-aminoadipic 6-semialdehyde dehydrogenase. It was speculated to be a PQQ-dependent AAS dehydrogenase due to the research of demonstrating PQQ as a new B vitamin. We isolated a novel human cDNA from the human fetal brain cDNA library we constructed. Its deduced protein was most related to mouse U26. Thus, we termed it human U26. This putative protein contains an AMP-binding domain, a Phosphopantetheine-binding domain and six PQQ-binding motifs. Human U26 mRNA is ubiquitously expressed in adult tissues and is highly expressed in colon adenocarcinoma (CX-1) and colon adenocarcinoma (GI-112) cell lines. Further study should be made to clarify the precise function of human U26.

The rapid growth of different kinds of biological information allows a good opportunity to analyze the co-evolutionary characteristics in endocrine regulatory pathways. Data ranging from kinds of species' genome, gene sequence, protein structure, and expression profile of different organisms can reveal the inner co-evolutionary relationship of ligands, receptors, and other related molecules. In return, these co-evolutionary characteristics can help us determine uncharacterized ligands and receptors, annotate gene functions, highlight amino acid residues with biochemical significance, and identify regulated genes in the endocrine process. Encouraging examples in this field, although at their starting stage, have emerged. Here we focus on recent progress in endocrine-related co-evolution research from a methodological approach.

Members of the large family of WD-repeat proteins are involved in diverse functions such as RNA-procession, signal transduction, vesicular trafficking, cytoskeletal assembly, and cell cycle control. By large-scale-sequencing analysis of a human fetal brain cDNA library, we isolated a novel human cDNA encoding a 7-WD40-repeat protein. This cDNA is 2004 bp in length and it codes for a 544-amino-acid protein. We term it human WD40-repeat-containing gene 25 (WDR25) and this gene shows significant similarity with human pre-mRNA splicing factor 17. The WDR25 gene is mapped to chromosome 14q32 and contains seven exons. RT-PCR analysis shows that the WDR25 gene is widely expressed in human tissues and the expression levers in heart, muscle, testis, ovary, uterus, and prostate are relatively high.

We have isolated a novel cDNA from the human fetal brain cDNA library with homology to the Mg2+ -dependent serine/threonine protein phosphatase 2C (PP2C) family. The cDNA is 3055 bp in length, and the predicted coding region encodes a 360-amino-acid protein, which shows 99% identity to the PP2C epsilon from rat and mouse. Then we term it human PP2C epsilon gene. The gene is mapped to chromosome 3q26.1 and contains 4 exons. RT-PCR analysis shows that the PP2C epsilon is widely expressed in human tissues and the expression levels in heart, placenta, lung, liver, kidney, and pancreas are relatively high.

Glycosyltransferases constitute a large group of enzymes that are involved in a wide range of functions in all living organisms. By large-scale sequencing analysis of a human fetal brain cDNA library, we isolated a novel human putative glycosyltransferase gene named beta3GnTL1. Its cDNA is 1372 base pair in length, encoding a predicted protein with 361 amino acid residues. The human beta3GnTL1 is located to chromosome 17q25.3 by comparison of its cDNA with human gemome database. RT-PCR result shows the beta3GnTL1 is expressed at various levels in most of tissues examined.

The broad-complex, tramtrack (ttk) and bric-a-brac/poxvirus and zinc finger proteins (BTB/POZ) domain is highly conserved in a large family of eukaryotic proteins and is crucial for the latter's diverse roles in mediating interactions among proteins that are involved in transcription regulation and chromatin structures. From a fetal brain cDNA library, we isolated a cDNA of 2489 base pairs (bp) encoding a novel human BTB domain-containing protein named BTBD10. The cDNA contained an open-reading frame (ORF) of 1428 bp encoding a putative 475-amino acid (aa) protein. The BTBD10 gene was located on human chromosome 11p15.2 and consisted of nine exons spanning about 75.2 kilobase pairs (kb) of the human genome. The cDNA microarray analysis showed that BTBD10 was down-regulated in all 18 glioma samples. The expression pattern of BTBD10 gene was examined by multiple tissue cDNA (MTC) panels (Clontech), which showed a ubiquitous expression pattern in the 16 tissues examined with high expression in adult brain, testis and small intestine and weak expression in the heart, lung, liver, kidney, pancreas, spleen, thymus, prostate, ovary and colon. The subcellular localization result revealed that BTBD10 was located specifically in the nucleus of HEK293 and COS7 cell lines, suggesting that it may function in transcriptional regulation. The different expression patterns of BTBD10 in different grades of glioma versus normal brain were also examined by RT-PCR and Northern blot. We also investigated the expression of BTBD10 in hepatocellular carcinoma, ovary cancer and lung cancer, and the results revealed no significant difference in these three tumors. All these data suggested that BTBD10 might play a role in glioma.

TGF-beta activated kinase (TAK1) plays a critical role in the TGF-beta signaling transduction pathway. By screening a human 18-week fetal brain library, we isolated a novel human TAK1-like (TAKL) gene. The gene encoded a putative protein of 242 amino acids, which shared a homology with human, mouse, and Xenopus TAK1. The TAKL gene was located in chromosome 21q21. Northern blot analysis revealed that the TAKL mRNA was expressed predominantly in peripheral blood leukocytes and ubiquitously in human adult and fetal tissues. TAKL was also expressed strongly in breast carcinoma GI-101, colon adenocarcinoma GI-112, and prostatic adenocarcinoma PC3.

Supernatant protein factor (SPF) and alpha-tocopherol-associated protein (TAP) both belong to a widespread lipid-binding Sec 14-like protein family. All the members of the family have the lipid-binding motif called CRAL_TRIO. SPF is showed to stimulate the conversion of squalene to lanosterol and enhance cholesterol biosynthesis. TAP is identified to be involved in the intracellular distribution of alpha-tocopherol. Recently TAP is identified as SPF though they have very different functions. Here we report a human SPF/TAP homology SEC14L3 with 2082 base pairs in length and contains an open reading frame encoding a 400 amino acids protein. Analysis shows that SEC14L3 is mapped to 22q12 and expresses only in the liver among the used sixteen tissues in the test.

Two novel splice variants of CDK5RAP1, named CDK5RAP1_v3 and CDK5RAP1_v4, were isolated through the large-scale sequencing analysis of a human fetal brain cDNA library. The CDK5RAP1_v3 and CDK5RAP1_v4 cDNAs are 1923bp and 1792bp in length, respectively. Sequence analysis revealed that CDK5RAP1_v4 lacked 1 exon, which was present in CDK5RAP1_v3, with the result that these cDNAs encoded different putative proteins. The deduced proteins were 574 amino acids (designated as CDK5RAP1_v3) and 426 amino acids (CDK5RAP1_v4) in length, and shared the 420 N-terminal amino acids. RT-PCR analysis showed that human CDK5RAP1_v3 was widely expressed in human tissues. The expression level of CDK5RAP1_v3 was relatively high in placenta and lung, whereas low levels of expression were detected in heart, brain, liver, skeletal muscle, pancreas, spleen, thymus, small intestine and peripheral blood leukocytes. In contrast, human CDK5RAP1_v4 was mainly expressed in brain, placenta and testis.