With the advancement of transcriptome profiling by micro-arrays and high-throughput RNA-sequencing, transcriptome complexity and its dynamics are revealed at different levels in cardiovascular development and diseases. In this review, we will highlight the recent progress in our knowledge of cardiovascular transcriptome complexity contributed by RNA splicing, RNA editing and noncoding RNAs. The emerging importance of many of these previously under-explored aspects of gene regulation in cardiovascular development and pathology will be discussed.
Cardiovascular diseases; Genes; Molecular biology; Signal transduction
PPM1G is a nuclear localized serine/threonine phosphatase implicated to be a regulator of chromatin remodeling, mRNA splicing and DNA damage. However, its in vivo function is unknown.
Here we show that ppm1g expression is highly enriched in the central nervous system during mouse and zebrafish development. ppm1g−/− mice were embryonic lethal with incomplete penetrance after E12.5. Rostral defects, including neural tube and craniofacial defects were observed in ppm1g−/− embryos associated with increased cell death in the neural epithelium. In zebrafish, loss of ppm1g also led to neural defects with aberrant neural marker gene expression. Primary fibroblasts from ppm1g−/− embryos failed to grow without immortalization while immortalized ppm1g−/− fibroblasts had increased cell death upon oxidative and genotoxic stress when compared to wild type fibroblasts.
Our in vivo and in vitro studies revealed a critical role for PPM1G in normal development and cell survival.
PPM1G; neural tube; serine threonine phosphatase; PP2Cγ
MicroRNA-21 (miR-21) has been suggested to play a significant role in the prognosis of carcinoma. The recognition of novel biomarkers for the prediction of cancer outcomes is urgently required. However, the potential prognostic value of miR-21 in various types of human malignancy remains controversial. The present meta-analysis summarises and analyses the associations between miR-21 status and overall survival (OS) in a variety of tumours.
Eligible published studies were identified by searching the PubMed and Chinese Biomedicine databases. The patients’ clinical characteristics and survival results were pooled, and a pooled hazard ratio (HR) with 95% confidence intervals (95% CI) was used to calculate the strength of this association. A random-effects model was adopted, and then, meta-regression and subgroup analyses were performed. In addition, an analysis of publication bias was also conducted.
Twenty-seven eligible articles (including 31 studies) were identified that included survival data for 3273 patients. The pooled HR suggested that high miR-21 was clearly related to worse overall survival (HR = 2.27, 95% CI: 1.81-2.86), with a heterogeneity measure index of I2 = 76.0%, p = 0.001, showing that miR-21 might be a considerable prognostic factor for poor survival in cancer patients.
MiR-21 might be a potentially useful biomarker for predicting cancer prognosis in future clinical applications.
Electronic supplementary material
The online version of this article (doi:10.1186/1471-2407-14-819) contains supplementary material, which is available to authorized users.
miR-21; Cancer; Prognosis; Meta-analysis
Post-transplant lymphoproliferative disorder (PTLD) is a potentially fatal complication of solid organ transplantation. The current report presents the case of a 42-year-old male who developed PTLD within the first year following renal transplantation. The disorder manifested as a tumor adjacent to the lower pole of the renal allograft and resulted in urinary obstruction. Durable complete remission was achieved as a result of surgical resection followed by a reduction in immunosuppression and low-dose rituximab-based chemotherapy, indicating that this therapeutic strategy may be safe and effective for the treatment of specific cases of localized and resectable PTLD.
post-transplant lymphoproliferative disorder; Epstein-Barr virus; early-onset; kidney transplantation
Edge detection is a key step in medical image processing. It is widely used to extract features, perform segmentation, and further assist in diagnosis. A poor quality edge map can result in false alarms and misses in cancer detection algorithms. Therefore, it is necessary to have a reliable edge measure to assist in selecting the optimal edge map. Existing reference based edge measures require a ground truth edge map to evaluate the similarity between the generated edge map and the ground truth. However, the ground truth images are not available for medical images. Therefore, a nonreference edge measure is ideal for medical image processing applications. In this paper, a nonreference reconstruction based edge map evaluation (NREM) is proposed. The theoretical basis is that a good edge map keeps the structure and details of the original image thus would yield a good reconstructed image. The NREM is based on comparing the similarity between the reconstructed image with the original image using this concept. The edge measure is used for selecting the optimal edge detection algorithm and optimal parameters for the algorithm. Experimental results show that the quantitative evaluations given by the edge measure have good correlations with human visual analysis.
Recurrent pericarditis is a rare complication following renal transplantation. Colchicine, an inhibitor of microtubule polymerization, has been recommended for the treatment of recurrent acute pericarditis in non-transplant patients and is commonly used for the treatment of gout in transplant patients. However, the use of colchicine for the treatment of recurrent pericarditis in renal transplant patients has rarely been reported. In the present study, a rare case of recurrent pericarditis, manifested as large pericardial effusion and pericardial tamponade within the first year following renal transplantation, was successfully treated with colchicine. Therefore, low-dose colchicine may be a safe and effective option for the treatment of recurrent pericarditis in renal transplant patients.
kidney transplant; recurrent pericarditis; pericardial tamponade; colchicine
The novel adipokine chemerin plays a role in regulating lipid and carbohydrate metabolism, and recent reports of elevated chemerin levels in polycystic ovarian syndrome elevated chemerin levels with polycystic ovary syndrome and preeclampsia point to an emerging role for chemerin in reproduction. We hypothesized that chemerin, like other adipokines, may function to regulate male gonadal steroidogenesis. Here we show that chemerin and its three receptors chemokine-like receptor 1 (CMKLR1), G-protein coupled receptor 1 (GPR1) and chemokine (C-C motif) receptor-like 2 (CCRL2) were expressed in male reproductive tracts, liver and white adipose tissue. CMKLR1 and GPR1 protein were localized specifically in the Leydig cells of human and rat testes by immunohistochemistry. The expression of chemerin and its receptors in rat testes was developmentally regulated and highly expressed in Leydig cells. In vitro treatment with chemerin suppressed the human chorinoic gonadotropin (hCG)-induced testosterone production from primary Leydig cells, which was accompanied by the inhibition of 3beta-hydroxysteroid dehydrogenase (3beta-HSD) gene and protein expression. The hCG-activated p44/42 mitogen-activated-protein kinase (MAPK) (Erk1/2) pathway in Leydig cells was also inhibited by chemerin co-treatment. Together, these data suggest chemerin is a novel regulator of male gonadal steroidogenesis.
chemerin; steroidogenesis; testosterone; adipokine; Leydig cell
Here, we report a Chinese case of Creutzfeldt–Jakob disease (CJD) with a rare mutation in the prion protein gene (PRNP) leading to an exchange of amino acid from valine (Val) to isoleucine (I) at codon 203 (V203I). The 80-y-old male presented with sudden memory loss, rapid loss of vocabulary, inattention and slow responses, accompanied by dizziness, blurred vision and ataxia. Two weeks after admission, he exhibited tremor, myoclonus and bilateral Babinski signs. At the end of the clinical course, he developed severe akinetic mutism. The cerebrospinal fluid (CSF) was positive for 14-3-3 protein. Increased bilateral signal intensity in the frontal and parietal lobes was seen on diffusion-weighted imaging (DWI); periodic activity was recorded on an electroencephalogram (EEG). There was no family history of similar symptoms. The total clinical course was approximately two months.
14-3-3 protein; PRNP; Creutzfeldt–Jakob disease; V203I; mutation
Strain has been widely used to manipulate the properties of various kinds of materials, such as ferroelectrics, semiconductors, superconductors, magnetic materials, and “strain engineering” has become a very active field. For strain-based information storage, the non-volatile strain is very useful and highly desired. However, in most cases, the strain induced by converse piezoelectric effect is volatile. In this work, we report a non-volatile strain in the (001)-oriented Pb(Mg1/3Nb2/3)O3-PbTiO3 single crystals and demonstrate an approach to measure the non-volatile strain. A bipolar loop-like S-E curve is revealed and a mechanism involving 109° ferroelastic domain switching is proposed. The non-volatile high and low strain states should be significant for applications in information storage.
Wound-healing represents a major health burden, such as diabetes-induced skin ulcers and burning. Many works are being tried to find ideal clinical wound-healing biomaterials. Especially, small molecules with low cost and function to promote production of endogenous wound healing agents (i.e. transforming growth factor beta, TGF-β) are excellent candidates. In this study, a small peptide (tiger17, c[WCKPKPKPRCH-NH2]) containing only 11 amino acid residues was designed and proved to be a potent wound healer. It showed strong wound healing-promoting activity in a murine model of full thickness dermal wound. Tiger17 exerted significant effects on three stages of wound healing progresses including (1) the induction of macrophages recruitment to wound site at inflammatory reaction stage; (2) the promotion of the migration and proliferation both keratinocytes and fibroblasts, leading to reepithelialization and granulation tissue formation; and (3) tissue remodeling phase, by promoting the release of transforming TGF-β1 and interleukin 6 (IL-6) in murine macrophages and activating mitogen-activated protein kinases (MAPK) signaling pathways. Considering its easy production, store and transfer and function to promote production of endogenous wound healing agents (TGF-β), tiger17 might be an exciting biomaterial or template for the development of novel wound-healing agents.
SmCo5 based magnets with smaller size and larger maximum energy product have been long desired in various fields such as renewable energy technology, electronic industry and aerospace science. However, conventional relatively rough synthetic strategies will lead to either diminished magnetic properties or irregular morphology, which hindered their wide applications. In this article, we present a facile chemical approach to prepare 200 nm single domain SmCo5@Co core/shell magnets with coercivity of 20.7 kOe and saturation magnetization of 82 emu/g. We found that the incorporation of GO sheets is responsible for the generation of the unique structure. The single domain SmCo5 core contributes to the large coercivity of the magnets and the exchange-coupled Co shell enhances the magnetization. This method can be further utilized in the synthesis other Sm-Co based exchange-coupled magnets.
Nek9 (also known as Nercc1), a member of the NIMA (never in mitosis A) family of protein kinases, regulates spindle formation, chromosome alignment and segregation in mitosis. Here, we showed that Nek9 protein was expressed from germinal vesicle (GV) to metaphase II (MII) stages in mouse oocytes with no detectable changes. Confocal microscopy identified that Nek9 was localized to the spindle poles at the metaphase stages and associated with the midbody at anaphase or telophase stage in both meiotic oocytes and the first mitotic embyros. Depletion of Nek9 by specific morpholino injection resulted in severely defective spindles and misaligned chromosomes with significant pro-MI/MI arrest and failure of first polar body (PB1) extrusion. Knockdown of Nek9 also impaired the spindle-pole localization of γ-tubulin and resulted in retention of the spindle assembly checkpoint protein Bub3 at the kinetochores even after 10 h of culture. Live-cell imaging analysis also confirmed that knockdown of Nek9 resulted in oocyte arrest at the pro-MI/MI stage with abnormal spindles, misaligned chromosomes and failed polar body emission. Taken together, our results suggest that Nek9 may act as a MTOC-associated protein regulating microtubule nucleation, spindle organization and, thus, cell cycle progression during mouse oocyte meiotic maturation, fertilization and early embryo cleavage.
meiosis; microtubule-organizing center (MTOC) spindle; oocyte; γ-tubulin
Polychlorinated biphenyls (PCBs) are persistent organic pollutants damaging to human health and the environment. Techniques to indicate PCB contamination in planta are of great interest to phytoremediation. Monitoring of dioxin-like PCBs in transgenic plants carrying the mammalian aryl hydrocarbon receptor (AHR) has been reported previously. Herein, we report the biomonitoring of non-dioxin-like PCBs (NDL-PCBs) using the mammalian pregnane X receptor (PXR). In the transgenic Arabidopsis designated NDL-PCB Reporter, the EGFP-GUS reporter gene was driven by a promoter containing 18 repeats of the xenobiotic response elements, while PXR and its binding partner retinoid X receptor (RXR) were coexpressed. Results showed that, in live cells, the expression of reporter gene was insensitive to endogenous lignans, carotenoids and flavonoids, but responded to all tested NDL-PCBs in a dose- and time- dependent manner. Two types of putative PCB metabolites, hydroxy- PCBs and methoxy- PCBs, displayed different activation properties. The vascular tissues seemed unable to transport NDL-PCBs, whereas mutation in QUASIMODO1 encoding a 1,4-galacturonosyltransferase led to reduced PCB accumulation in Arabidopsis, revealing a role for pectin in the control of PCB translocation. Taken together, the reporter system may serve as a useful tool to biomonitor the uptake and metabolism of NDL-PCBs in plants.
Prion diseases are kinds of progressive, incurable neurodegenerative disorders. So far, survival time of the patients with these diseases in China is unclear.
Based upon the surveillance data from Chinese Creutzfeldt-Jakob disease (CJD) surveillance network from January 2008 to December 2011, a retrospective follow-up survey was performed. The survival times of Chinese patients with prion diseases and the possible influencing factors were analyzed.
Median survival time of 121 deceased patients was 7.1 months, while those for sporadic CJD (sCJD), familial CJD (fCJD) and fatal familial insomnia (FFI) cases were 6.1, 3.1 and 8.2 months, respectively. 74.0% of sCJD patients, 100% of fCJD cases and 91.7% FFI cases died within one year. The general socio-demographic factors, abnormalities in clinical examinations, clinical manifestations, and social factors did not significantly influence the survival times of Chinese prion patients.
Survival time of Chinese patients with prion diseases was comparable with that of many Western countries, but obviously shorter than that of Japan. Patients with acute onset and rapid progression had significantly short survival times.
Accurate and comprehensive de novo transcriptome profiling in heart is a central issue to better understand cardiac physiology and diseases. Although significant progress has been made in genome-wide profiling for quantitative changes in cardiac gene expression, current knowledge offers limited insights to the total complexity in cardiac transcriptome at individual exon level.
To develop more robust bioinformatic approaches to analyze high-throughput RNA sequencing (RNA-Seq) data, with the focus on the investigation of transcriptome complexity at individual exon and transcript levels.
Methods and Results
In addition to overall gene expression analysis, the methods developed in this study were used to analyze RNA-Seq data with respect to individual transcript isoforms, novel spliced exons, novel alternative terminal exons, novel transcript clusters (i.e., novel genes) and long non-coding RNA genes. We applied these approaches to RNA-Seq data obtained from mouse hearts following pressure-overload induced by trans-aortic constriction. Based on experimental validations, analyses of the features of the identified exons/transcripts, and expression analyses including previously published RNASeq data, we demonstrate that the methods are highly effective in detecting and quantifying individual exons and transcripts. Novel insights inferred from the examined aspects of the cardiac transcriptome open ways to further experimental investigations.
Our work provided a comprehensive set of methods to analyze mouse cardiac transcriptome complexity at individual exon and transcript levels. Applications of the methods may infer important new insights to gene regulation in normal and disease hearts in terms of exon utilization and potential involvement of novel components of cardiac transcriptome.
RNA-Seq; transcriptome profiling; hypertrophy; heart failure
Although the aggregation of PrPSc is thought to be crucial for the neuropathology of prion diseases, there is evidence in cultured cells and transgenic mice that neuronal death can be triggered by the accumulation of cytosolic PrPs, leading to the hypothesis that the accumulation of PrPs in the cytosol of neurons may be a primary neurotoxic culprit. Hsp70, a molecular chaperone involved in protein folding/refolding and degradation in the cytoplasm, has a protective effect in some models of neurodegenerative diseases, e.g., Alzheimer’s and Parkinson’s diseases, but its role in prion diseases remains unclear.
To study the role of Hsp70 in prion diseases, we used immunoprecipitation to first identify a molecular interaction between Hsp70 and PrPs. Using immunofluorescence, we found that Hsp70 colocalized with cytosolic PrPs in HEK293 cells transiently transfected with plasmids for Cyto-PrP and PG14-PrP but not with wild-type PG5-PrP or endoplasmic reticulum (ER)-retained PrPs (3AV-PrP and ER-PrP). Using western blot analysis and apoptosis assays of cultured cells, we found that the overexpression of Hsp70 by transfection or the activation of Hsp70 by geldanamycin selectively mediated the degradation of cytosolic PrPs and restored cytosolic PrP-induced cytotoxicity. Moreover, we found that Hsp70 levels were up-regulated in cells expressing Cyto-PrP and in hamster brains infected with the scrapie agent 263K.
These data imply that Hsp70 has central role in the metabolism of cytosolic PrPs
Hsp70; Cytosolic PrP; Apoptosis; Prion disease; Geldanamycin
In the eukaryotic transcriptome, both the numbers of genes and different RNA species produced by each gene contribute to the overall complexity. These RNA species are generated by the utilization of different transcriptional initiation or termination sites, or more commonly, from different messenger RNA (mRNA) splicing events. Among the 30 000+ genes in human genome, it is estimated that more than 95% of them can generate more than one gene product via alternative RNA splicing. The protein products generated from different RNA splicing variants can have different intracellular localization, activity, or tissue-distribution. Therefore, alternative RNA splicing is an important molecular process that contributes to the overall complexity of the genome and the functional specificity and diversity among different cell types. In this review, we will discuss current efforts to unravel the full complexity of the cardiac transcriptome using a deep-sequencing approach, and highlight the potential of this technology to uncover the global impact of RNA splicing on the transcriptome during development and diseases of the heart.
Alternative RNA splicing; Transcriptome; Gene regulation; Heart; RNA-seq
A case of Creutzfeldt-Jakob disease (CJD) with a rare mutation of the prion protein (PrP) gene (PRNP) at codon 208 (R208H), while the codon 129 was a methionine homozygous genotype is reported. The patient initial displayed hand tremor, dizziness and progressive cognitive dysfunction. Subsequently, other symptoms gradually appeared, including cerebellar ataxia and mental disorder. No periodic activity was recorded at electroencephalography (EEG) and 14-3-3 protein in cerebrospinal fluid was negative. Total clinical course was about four months. Retrospective investigation of this family across seven generations did not figure out clear family history. However, genetic analyses revealed six first-degree family members with the R208H allele.
creutzfeldt-Jakob disease; PRNP; R208H
Protein disulfide isomerase (PDI), is sorted to be enzymatic chaperone for reconstructing misfolded protein in endoplasmic reticulum lumen. Recently, PDI has been identified as a link between misfolded protein and neuron apoptosis. However, the potential for PDI to be involved in the pathogenesis of prion disease remains unknown. In this study, we propose that PDI may function as a pleiotropic regulator in the cytotoxicity induced by mutated prion proteins and in the pathogenesis of prion diseases.
To elucidate potential alterations of PDI in prion diseases, the levels of PDI and relevant apoptotic executors in 263K infected hamsters brain tissues were evaluated with the use of Western blots. Abnormal upregulation of PDI, Grp78 and Grp58 was detected. Dynamic assays of PDI alteration identified that the upregulation of PDI started at the early stage and persistently increased till later stage. Obvious increases of PDI and Grp78 levels were also observed in cultured cells transiently expressing PrP mutants, PrP-KDEL or PrP-PG15, accompanied by significant cytotoxicities. Excessive expression of PDI partially eased ER stress and cell apoptosis caused by accumulation of PrP-KDEL, but had less effect on cytotoxicity induced by PrP-PG15. Knockdown of endogenous PDI significantly amended cytotoxicity of PrP-PG15, but had little influence on that of PrP-KDEL. A series of membrane potential assays found that apoptosis induced by misfolded PrP proteins could be regulated by PDI via mitochondrial dysfunction. Moreover, biotin-switch assays demonstrated active S-nitrosylted modifications of PDI (SNO-PDI) both in the brains of scrapie-infected rodents and in the cells with misfolded PrP proteins.
Current data in this study highlight that PDI and its relevant executors may function as a pleiotropic regulator in the processes of different misfolded PrP proteins and at different stages during prion infection. SNO-PDI may feed as an accomplice for PDI apoptosis.
Multiple injuries have been highlighted as an important clinical dimension of the injury profile following earthquakes, but studies are scarce. We investigated the pattern and combination of injuries among patients with two injuries following the 2008 Wenchuan earthquake. We also described the general injury profile, causes of injury and socio-demographic characteristics of the injured patients.
A retrospective hospital-based analysis of 1,871 earthquake injured patients, totaling 3,177 injuries, admitted between 12 and 31 May 2008 to the People's Hospital of Deyang city (PHDC). An electronic, webserver-based database with International Classification of Diseases (ICD)-10-based classification of earthquake-related injury diagnoses (IDs), anatomical sites and additional background variables of the inpatients was used. We analyzed this dataset for injury profile and number of injuries per patient. We then included all patients (856) with two injuries for more in-depth analysis. Possible spatial anatomical associations were determined a priori. Cross-tabulation and more complex frequency matrices for combination analyses were used to investigate the injury profile.
Out of the 1,871 injured patients, 810 (43.3%) presented with a single injury. The rest had multiple injuries; 856 (45.8%) had two, 169 (9.0%) patients had three, 32 (1.7%) presented with four injuries, while only 4 (0.2%) were diagnosed with five injuries. The injury diagnoses of patients presenting with two-injuries showed important anatomical intra-site or neighboring clustering, which explained 49.1% of the combinations. For fractures, the result was even more marked as spatial clustering explained 57.9% of the association pattern. The most frequent combination of IDs was a double-fracture, affecting 20.7% of the two-injury patients (n = 177). Another 108 patients (12.6%) presented with fractures associated with crush injury and organ-soft tissue injury. Of the 3,177 injuries, 1,476 (46.5%) were fractures. Most injuries were located in the head (22.9%) and lower extremities (30.8%).
Multiple injuries are put forward as an important component of the injury profile after this earthquake. A pattern of injury combinations and spatial aggregation of injuries was also found. Clinical diagnosis and treatment should be adapted to care of these patients. More studies are needed to generalize these findings.
Genetic Creutzfeldt-Jakob disease (gCJD) is caused by a range of mutations in the prion protein gene (PRNP) and account for approximately 10–15% of overall human prion diseases worldwide. They are different with disease onset, disease duration, clinical signs and diagnostic findings. Here we reported a 71 year-old female with an E196K mutation in one PRNP allele, while the codon 129 was a methionine homozygous genotype. The patient started with non-specific symptoms, but displayed rapidly progressive disturbances of speech, memory, cognitive and physical movement. No periodic activity was recorded at electroencephalography (EEG) during the entire disease course. Retrospective investigation of her family members did not reveal similar neurological disorders. Total clinical course was about seven months.
Creutzfeldt-Jakob disease; PRNP; E196K
Transmissible spongiform encephalopathy (TSE) diseases are known to be zoonotic diseases that can infect different kinds of animals. The transmissibility of TSE, like that of other infectious diseases, shows marked species barrier, either being unable to infect heterologous species or difficult to form transmission experimentally. The similarity of the amino acid sequences of PrP among species is believed to be one of the elements in controlling the transmission TSE interspecies. Other factors, such as prion strains and host's microenvironment, may also participate in the process.
Two mouse-adapted strains 139A and ME7 were cerebrally inoculated to Golden hamsters. Presences of scrapie associate fibril (SAF) and PrPSc in brains of the infected animals were tested by TEM assays and Western blots dynamically during the incubation periods. The pathogenic features of the novel prions in hamsters, including electrophoretic patterns, glycosylating profiles, immunoreactivities, proteinase K-resistances and conformational stabilities were comparatively evaluated. TSE-related neuropathological changes were assayed by histological examinations.
After long incubation times, mouse-adapted agents 139A and ME7 induced experimental scrapie in hamsters, respectively, showing obvious spongiform degeneration and PrPSc deposits in brains, especially in cortex regions. SAF and PrPSc in brains were observed much earlier than the onset of clinical symptoms. The molecular characteristics of the newly-formed PrPSc in hamsters, 139A-ha and ME7-ha, were obviously distinct from the original mouse agents, however, greatly similar as that of a hamster-adapted scrapie strain 263 K. Although the incubation times and main disease signs of the hamsters of 139A-ha and ME7-ha were different, the pathogenic characteristics and neuropathological changes were highly similar.
This finding concludes that mouse-adapted agents 139A and ME7 change their pathogenic characteristics during the transmission to hamsters. The novel prions in hamsters' brains obtain new molecular properties with hamster-specificity.
The human papillomavirus (HPV) E2 protein is a multifunctional DNA-binding protein. The transcriptional activity of HPV E2 is mediated by binding to its specific binding sites in the upstream regulatory region of the HPV genomes. Previously we reported a HPV-2 variant from a verrucae vulgaris patient with huge extensive clustered cutaneous, which have five point mutations in its E2 ORF, L118S, S235P, Y287H, S293R and A338V. Under the control of HPV-2 LCR, co-expression of the mutated HPV E2 induced an increased activity on the viral early promoter. In the present study, a series of mammalian expression plasmids encoding E2 proteins with one to five amino acid (aa) substitutions for these mutations were constructed and transfected into HeLa, C33A and SiHa cells.
CAT expression assays indicated that the enhanced promoter activity was due to the co-expressions of the E2 constructs containing A338V mutation within the DNA-binding domain. Western blots analysis demonstrated that the transiently transfected E2 expressing plasmids, regardless of prototype or the A338V mutant, were continuously expressed in the cells. To study the effect of E2 mutations on its DNA-binding activity, a serial of recombinant E2 proteins with various lengths were expressed and purified. Electrophoresis mobility shift assays (EMSA) showed that the binding affinity of E2 protein with A338V mutation to both an artificial probe with two E2 binding sites or HPV-2 and HPV-16 promoter-proximal LCR sequences were significantly stronger than that of the HPV-2 prototype E2. Furthermore, co-expression of the construct containing A338V mutant exhibited increased activities on heterologous HPV-16 early promoter P97 than that of prototype E2.
These results suggest that the mutation from Ala to Val at aa 338 is critical for E2 DNA-binding and its transcriptional regulation.
HPV-2; E2; DNA-binding; Transcriptional regulation; Promoter
Creutzfeldt-Jakob disease (CJD) is a rare, rapidly progressive fatal central nervous system disorder, which consists of three main catalogues: sporadic, familial, and iatrogenic CJD.
In China, the surveillance for CJD started in 2006, covering 12 provincial Centers for Disease Control and Prevention (CDCs) and 15 hospitals. From 2006 to 2010, 624 suspected patients were referred to China CJD surveillance. The epidemiological, clinical and laboratory features of sporadic CJD (sCJD) were analysed. Both groups of probable and possible sCJD showed highest incidences in the population of 60 to 69 year-olds. The most common presenting symptoms were progressive dementia and mental-related symptoms (neurological symptoms including sleeping turbulence, depression, anxiety and stress). Among the four main clinical manifestations, myoclonus was more frequently observed in the probable sCJD patients. About 2/3 of probable sCJD cases showed positive 14-3-3 in CSF and/or periodic sharp wave complexes (PSWC) in electroencephalography (EEG). The presence of myoclonus was significantly closely related with the appearance of PSWC in EEG. Polymorphisms of codon 129 in PRNP of the notified cases revealed a highly predominant M129M genotype in Han Chinese. Among 23 genetic human prion diseases, ten were D178N/M129M Fatal familial insomnia (FFI) and five were T188K genetic CJD (gCJD), possibly indicating a special distribution of gCJD-related mutations in Han Chinese.
From the period of 2006 to 2010, 261 patients were diagnosed as sCJD and 23 patients were diagnosed as genetic human prion diseases in China. The epidemiological, clinical and laboratory analysis data were consistent with the characteristics of sporadic CJD, which provide insight into the features of CJD in China.
Tubulin polymerization promoting protein/p25 (TPPP/p25), known as a microtubule-associated protein (MAP), is a brain-specific unstructured protein with a physiological function of stabilizing cellular microtubular ultrastructures. Whether TPPP involves in the normal functions of PrP or the pathogenesis of prion disease remains unknown. Here, we proposed the data that TPPP formed molecular complex with PrP. We also investigated its influence on the aggregation of PrP and fibrillization of PrP106–126 in vitro, its antagonization against the disruption of microtubule structures and cytotoxicity of cytosolic PrP in cells, and its alternation in the brains of scrapie-infected experimental hamsters.
Using pull-down and immunoprecipitation assays, distinct molecular interaction between TPPP and PrP were identified and the segment of TPPP spanning residues 100–219 and the segment of PrP spanning residues 106–126 were mapped as the regions responsible for protein interaction. Sedimentation experiments found that TPPP increased the aggregation of full-length recombinant PrP (PrP23–231) in vitro. Transmission electron microscopy and Thioflavin T (ThT) assays showed that TPPP enhanced fibril formation of synthetic peptide PrP106–126 in vitro. Expression of TPPP in the cultured cells did not obviously change the microtubule networks observed by a tubulin-specific immunofluorescent assay and cell growth features measured by CCK8 tests, but significantly antagonized the disruption of microtubule structures and rescued the cytotoxicity caused by the accumulation of cytosolic PrP (CytoPrP). Furthermore, Western blots identified that the levels of the endogenous TPPP in the brains of scrapie-infected experimental hamsters were significantly reduced.
Those data highlight TPPP may work as a protective factor for cells against the damage effects of the accumulation of abnormal forms of PrPs, besides its function as an agent for dynamic stabilization of microtubular ultrastructures.