Ultrasound assisted phytochemical extraction of red cabbage by using deep eutectic solvent: Modelling using ANFIS and optimization by genetic algorithms.

The present investigation studied the effect of process parameters on the extraction of phytochemicals from red cabbage by the application of ultrasonication and temperature. The solvent selected for the study was deep eutectic solvent (DES) prepared by choline chloride and citric acid. The ultrasound assisted extraction process was modeled using adaptive neuro-fuzzy inference system (ANFIS) algorithm and integrated with the genetic algorithm for optimization purposes. The independent variables that influenced the responses (total phenolic content, antioxidant activity, total anthocyanin activity, and total flavonoid content) were ultrasonication power, temperature, molar ratio of DES, and water content of DES. Each ANFIS model was formed by the training of three Gaussian-type membership functions (MF) for each input, trained by a hybrid algorithm with 500 epochs and linear type MF for output MF. The ANFIS model predicted each response close to the experimental data which is evident by the statistical parameters (R2>0.953 and RMSE <1.165). The integrated hybrid ANFIS-GA algorithm predicted the optimized condition for the process parameters of ultrasound assisted extraction of phytochemicals from red cabbage was found to be 252.114 W for ultrasonication power, 52.715 °C of temperature, 2.0677:1 of molar ratio of DES and 25.947 % of water content in DES solvent with maximum extraction content of responses, with fitness value 3.352. The relative deviation between the experimental and ANFIS predicted values for total phenolic content, antioxidant activity, total anthocyanin activity, and total flavonoid content was found to be 1.849 %, 3.495 %, 2.801 %, and 4.661 % respectively.

Dragon fruit peel extract microcapsule incorporated pearl millet and dragon fruit pulp powder based functional pasta: formulation, characterization, and release kinetics study.

The pearl millet based functional pasta was formulated by incorporating freeze dried dragon fruit pulp powder and 2% (w/w) microcapsule containing dragon fruit peel extract. The control pasta consisted of 100% pearl millet flour. The other four functional pasta samples consisted of pearl millet and freeze-dried dragon fruit pulp powder (DFP) in the ratio of 95:5, 90:10, 85:15, and 80:20 (w/w), respectively. The inclusion of dragon fruit powder enhanced the swelling index, water absorption index, color, and functional properties of the pasta. The total phenolic content (0.24-0.43 mg GAE/100 g d.w.), antioxidant activity (17.76-30.67%), and betacyanin content (0.149-0.152 mg/g d.w.) of the pasta was increased with the increase of dragon fruit pulp level in the formulation. The release kinetics of phenolic compounds into the simulated gastric juice was modeled using Higuchi and Peppas- Sahlin models. Out of these two models Peppas- Sahlin model ( R 2 > 0.980 and R M S E < 1.527 ) found to predict the release of phenolics into simulated gastric juice with respect to time of release when compared with Higuchi model ( R 2 > 0.964 and R M S E < 6.126 ). The onset of transition temperature and enthalpy of gelatinization of pasta samples was found to be in the range of 66.321-74.681 °C and increased with the increase of dragon fruit level in the formulation.

Effect of intermittent microwave convective drying on physicochemical properties of dragon fruit.

The study was carried out to investigate the effect of Intermittent microwave convective drying (IMCD) on the overall quality of dried dragon fruit in terms of total phenolic content, color change, and rehydration ratio. Three levels of microwave power (200-600 W) and a temperature of 60 °C for hot air were applied alternately throughout the process with three levels of pulse ratio such as 1:10, 1:20, and 1:40, respectively. The total phenolic content of the dragon fruit slice obtained by IMCD was ranged between 5.750 and 6.575 mg GAE/g dry weight. Within the experimental range of process variables under IMCD conditions, the drying efficiency, color change, and rehydration ratio of the dried dragon fruit slices were 15.287-51.930%, 18.643-24.847, and 1.908-3.239, respectively. The Weibull model scale (α) parameter was found to vary between 27.512 - 498.174 , while the shape (ß) parameter was found to vary between 0.769 - 0.851 . The Weibull model parameters were shown to decrease with increasing microwave power at constant pulse ratio. The IMCD method produced a dried dragon fruit slices with reduced color changes and higher total phenolic content and rehydration ratio values. This investigation would contribute to the development of effective drying techniques for increased food quality and product consistency in the drying of diverse fruits and vegetables.

Comprehensive study on applications of artificial neural network in food process modeling.

Artificial neural network (ANN) is a simplified model of the biological nervous system consisting of nerve cells or neurons. The application of ANN to food process engineering is relatively novel. ANN had been employed in diverse applications like food safety and quality analyses, food image analysis, and modeling of various thermal and non-thermal food-processing operations. ANN has the ability to map nonlinear relationships without any prior knowledge and predicts responses even with incomplete information. Every neural network possesses data in the form of connection weights interconnecting lines between the input to hidden layer neurons and weights of hidden to output layer neurons, which has a significant role in predicting the output data. The applications of ANN in different unit operations in food processing were described that includes theoretical developments using intelligent characteristics for adaptability, automatic learning, classification, and prediction. The parallel architecture of ANN resulted in a fast response and low computational time making it suitable for application in real-time systems of different food process operations. The predicted responses obtained by the ANN model exhibited high accuracy due to lower relative deviation and root mean squared error and higher correlation coefficient. This paper presented the various applications of ANN for modeling nonlinear food engineering problems. The application of ANN in the modeling of the processes such as extraction, extrusion, drying, filtration, canning, fermentation, baking, dairy processing, and quality evaluation was reviewed.HIGHLIGHTS1. This paper discusses application of ANN in different emerging trends in food process.2. Application of ANN to develop non-linear multivariate modeling is illustrated.3. ANNs have been shown to be useful tool for prediction of outcomes with high accuracy.4. ANN resulted in fast response making it suitable for application in real time systems.

Ultrasound-assisted extraction of phytocompounds from dragon fruit peel: Optimization, kinetics and thermodynamic studies.

Ultrasound-assisted extraction method (UAE) was applied to recover phytocompounds from dragon fruit peel and the process was modelled and optimized using the combination of artificial neural network (ANN) and genetic algorithm (GA). The influence of ultrasonic temperature (30-70 °C), solvent to solid ratio (10:1-30:1 mL/g), solvent concentration (30-60%), and ultrasonic treatment time (5-25 min) on total polyphenolic content (ZT), antioxidant activity (ZD) and betacyanin content (ZB) was investigated. The ANN model successfully fitted to the experimental data and the output of ANN model was applied for genetic algorithm optimization. The optimal UAE conditions were obtained at ultrasonic temperature of 60 °C, solvent to solid ratio 25:1 mL/g, solvent concentration 60%, and ultrasonic treatment time of 20 min. The extraction kinetics and thermodynamic study for phytochemical compounds extracted from dragon fruit peel using UAE process was carried out at different combinations of temperature and time of extraction. The effective diffusion coefficient for total polyphenol content, antioxidant activity and betacyanin content were ranged from 2.99×10-11to4.84×10-11m2/s, 1.89×10-11to4.51×10-11m2/s and 2.55×10-11to5.40×10-11m2/s respectively and the corresponding mass transfer coefficient were varied between 2.00×10-06-2.81×10-06m/s, 1.53×10-06-2.66×10-06m/s and 1.81×10-06-3.05×10-06m/s respectively. The obtained information on effective diffusivity and mass transfer coefficient during extraction would allow the prediction of extraction rate and for estimation of operation conditions for industrial implementation.

Progesterone receptor-B enhances estrogen responsiveness of breast cancer cells via scaffolding PELP1- and estrogen receptor-containing transcription complexes.

Progesterone and estrogen are important drivers of breast cancer proliferation. Herein, we probed estrogen receptor-α (ER) and progesterone receptor (PR) cross-talk in breast cancer models. Stable expression of PR-B in PR-low/ER+ MCF7 cells increased cellular sensitivity to estradiol and insulin-like growth factor 1 (IGF1), as measured in growth assays performed in the absence of exogenous progestin; similar results were obtained in PR-null/ER+ T47D cells stably expressing PR-B. Genome-wide microarray analyses revealed that unliganded PR-B induced robust expression of a subset of estradiol-responsive ER target genes, including cathepsin-D (CTSD). Estradiol-treated MCF7 cells stably expressing PR-B exhibited enhanced ER Ser167 phosphorylation and recruitment of ER, PR and the proline-, glutamate- and leucine-rich protein 1 (PELP1) to an estrogen response element in the CTSD distal promoter; this complex co-immunoprecipitated with IGF1 receptor (IGFR1) in whole-cell lysates. Importantly, ER/PR/PELP1 complexes were also detected in human breast cancer samples. Inhibition of IGF1R or phosphoinositide 3-kinase blocked PR-B-dependent CTSD mRNA upregulation in response to estradiol. Similarly, inhibition of IGF1R or PR significantly reduced ER recruitment to the CTSD promoter. Stable knockdown of endogenous PR or onapristone treatment of multiple unmodified breast cancer cell lines blocked estradiol-mediated CTSD induction, inhibited growth in soft agar and partially restored tamoxifen sensitivity of resistant cells. Further, combination treatment of breast cancer cells with both onapristone and IGF1R tyrosine kinase inhibitor AEW541 was more effective than either agent alone. In summary, unliganded PR-B enhanced proliferative responses to estradiol and IGF1 via scaffolding of ER-α/PELP1/IGF1R-containing complexes. Our data provide a strong rationale for targeting PR in combination with ER and IGF1R in patients with luminal breast cancer.

Assessing treatment response after induction Bacillus Calmette-Guerin for carcinoma in situ of the urinary bladder: can post-induction random bladder biopsies be avoided?

OBJECTIVES: Patients diagnosed with bladder carcinoma in situ (CIS) and treated with intravesical Bacillus Calmette-Guerin (BCG) often undergo post-induction random bladder biopsies to assess treatment response. We sought to determine the correlation between post-induction urinary cytology/cystoscopy and histopathological findings obtained by random bladder biopsies. METHODS: Patients who were treated with BCG between 2006 and 2010 for CIS, had surveillance cystoscopy and cytology, and subsequently underwent random bladder biopsies were selected for analysis. Patients with a history of or concomitant urothelial cell carcinoma (UCC) stage T1 or higher were excluded. Cystoscopic finings were characterized as follows: negative - no mucosal erythema, raised lesions or papillary tumours; suspicious - mucosal erythema, but no raised lesions or papillary tumours; and positive - sessile or papillary tumours. The accuracy of cytology in predicting the results of subsequent random bladder biopsies was analysed. RESULTS: Of 21 patients included, surveillance cystoscopy findings were characterized as negative in nine, suspicious in seven and positive in five. Of 16 patients with negative/suspicious cystoscopy, 13 had agreement between cytology and biopsy, nine of whom were negative and four positive. Three of 16 patients had positive cytology, but negative biopsies; on further investigation of these three, one had CIS and two subsequent UCC. In the positive cystoscopy group, four of five patients had agreement between cytology and biopsy, two of whom were negative and two positive. One of the five patients had negative cytology, but a positive biopsy. CONCLUSION: Our data suggest foregoing random bladder biopsies in patients with negative urine cytology and no evidence of intravesical recurrence on cystoscopy following an induction course of BCG for CIS of the urinary bladder.

Management of small renal tumors: an overview.

The incidental finding of a small renal mass poses a therapeutic dilemma. The traditional treatment of clinically important masses has been radical nephrectomy. Recently, nephron-sparing surgery has emerged as a viable alternative; and experimental minimally invasive percutaneous tissue ablation techniques, including cryotherapy and radiofrequency ablation, are being evaluated. In this review, we discuss the dilemma posed by frequent renal imaging and the increased proportion of incidental tumors being detected, the limitations of needle biopsies for histologic diagnosis, nephron-conserving and minimally invasive surgery, and the possible merits of radiofrequency ablation and cryotherapy. We envision a defined role for minimally invasive percutaneous or extracorporeal ablation of small renal tumors.

The incidence of nephrolithiasis in patients with spinal neural tube defects.

PURPOSE: Bladder stones are common in patients with spinal neural tube defects but there are little data on the incidence of renal calculi in this population. We examined the incidence, nature and risk factors of nephrolithiasis in our clinic population of patients with neural tube defects. MATERIALS AND METHODS: We retrospectively reviewed the charts and radiological studies of 327 patients followed at our neural tube defects clinic with routine radiological imaging of the urinary tract. Additional confirmatory studies were performed when stones were noted. RESULTS: Renal calculi were identified in 20 patients with neural tube defects (6.1%). The incidence of nephrolithiasis increased with age. Renal stones were noted in 19 patients (10.7%) 12 years old or older. Management of the stones in these patients resulted in overall 53% stone-free and 87% recurrence rates after intervention. Major risk factors for new and/or recurrent renal stone formation were bacteriuria in 95% of the cases, lower urinary tract reconstruction in 80%, pelvicalicectasis in 70%, vesicoureteral reflux in 65%, a thoracic level spinal defect in 60% and renal scarring in 55%. CONCLUSIONS: These data suggest that there is a higher incidence of nephrolithiasis in patients with neural tube defects than in the general population and the risk of stone recurrence is also elevated. Most patients with stones had undergone lower urinary tract reconstruction. Other risk factors were bacteriuria, pelvicalicectasis, vesicoureteral reflux and a thoracic level neural tube defect.

T-antigen-dependent transcriptional initiation and its role in the regulation of human neurotropic JC virus late gene expression.

The multifunctional protein of papovaviruses, T-antigen, regulates the virus lytic cycle partly by exerting transcriptional control over viral and cellular gene expression. In this study, the ability of the T-antigen of human neurotropic JC virus (JCV) to enhance expression from the virus late promoter has been further examined. By deletion analysis, a T-antigen-responsive region was mapped within the JCV 98 bp enhancer/promoter between nucleotides 139 and 168. Interestingly, T-antigen appears to mediate transactivation by increasing expression from a basal transcriptional initiation site and through a novel T-antigen-dependent initiation site (TADI). The TADI element contains a region homologous to initiator (Inr) sequences and is sufficient to confer T-antigen responsiveness to a heterologous minimal promoter. Electrophoretic mobility shift and UV crosslinking analyses demonstrate that multiple cellular proteins interact with both single- and double-stranded forms of this sequence. Mutations within the TADI element which abolish T-antigen-mediated transcriptional activation also prevent the formation of specific nucleoprotein complexes. These data suggest that the ability of JCV T-antigen to regulate JCV late gene expression may be partly due to the formation of specific nucleoprotein complexes and transcriptional initiation from the TADI site on the viral promoter.

Utilization of polymerase chain reaction technology in the detection of solid tumors.

BACKGROUND: Most cancer detection tests currently performed are based on either antibody assays to a marker protein with altered expression in cancer patients or on imaging studies to identify characteristic lesions. Generally, for a positive result, these detection assays require that a tumor have a significant volume of cancer cells. Advances in diagnostic techniques and technology may allow for cancer detection at earlier stages, when the tumor burden is smaller and potentially more curable. The molecular techniques of polymerase chain reaction (PCR) and reverse transcriptase PCR (RT-PCR) are highly sensitive methods for detecting a small number of cancer cells. Over the past few years, numerous clinical studies have used PCR techniques to detect physical alterations of genes, such as mutations, deletions, translocations and amplification, the presence of oncogenic viruses, and the expression of genes specific to tissue, cancer, and metastasis. The current status of PCR as a method for detecting marker genes in the management of solid tumors is reviewed. METHODS: A review of the literature on the clinical utility of PCR and RT-PCR in the detection of solid tumor micrometastasis was conducted. RESULTS: Amplification by PCR is a highly sensitive method to determine gene expression. A single cell expressing a tumor marker among 10-100 million lymphocytes can be detected by the PCR assay. This approach has been used to detect tumor cells in approximately 18 different solid tumor types, with melanoma and carcinoma of the breast and prostate the most widely investigated to date. PCR-based assays have been used to detect cancer cells in biopsies of solid tissue, lymph nodes, bone marrow, peripheral blood, and other body fluids. Several studies have reported a high specificity and sensitivity of tumor marker detection and a high correlation between PCR results and the presence of metastatic disease. However, in a few studies, PCR assays have not consistently demonstrated a higher sensitivity and specificity of detection than traditional modalities for many types of cancer. There has been a wide range in sensitivity and specificity among the studies, which may be partly attributed to the lack of uniformity among the PCR protocols used in different studies. CONCLUSIONS: PCR can detect tumor marker-expressing cells that are otherwise undetectable by other means in patients with localized or metastatic cancer. Reports from various study groups have lacked uniformity in their protocols, and this has prevented adequate comparison. The clinical utility of this assay as a tool for the prognosis and management of cancer patients remains and area of active investigation. PCR is a powerful tool in the study of the biology of cancer metastasis and will likely serve as a useful adjunct to clinical decision-making in the future.

Reverse transcriptase polymerase chain reaction for prostate specific antigen in the management of prostate cancer.

PURPOSE: Polymerase chain reaction is a powerful tool for expanding minute quantities of deoxyribonucleic acid (DNA) for detailed study. Reverse transcriptase polymerase chain reaction (RT-PCR) involves the initial conversion of messenger ribonucleic acid (mRNA) to DNA, followed by the amplification of the DNA product for molecular analysis. The mRNA for prostate specific antigen (PSA) is expressed only by prostatic cells. RT-PCR offers a potentially more sensitive assay for the detection of cells expressing PSA mRNA in peripheral circulation or in extraprostatic tissues. The current status of RT-PCR in the amplification and detection of PSA gene expression in the management of prostate cancer is reviewed. MATERIALS AND METHODS: The literature was reviewed for available data using RT-PCR for detection of prostatic cells outside of the prostate. RESULTS: Amplification of mRNA by RT-PCR represents a highly sensitive method of detection of gene expression. A single cell expressing PSA among 10 to 100 million lymphocytes can be detected by the RT-PCR assay. This assay may detect extraprostatic or circulating prostatic cells in peripheral blood, lymph nodes and bone marrow in many patients with prostate cancer. Various studies have reported sensitivities of detection of PSA expressing cells in the peripheral blood ranging from 0 to 88%. This wide range in the sensitivity may be partly due to tremendous variation between the protocols used in each study. In patients with lymph node metastasis the RT-PCR assay appears more reliable than immunohistochemistry for identification of prostatic tissue in the lymph node. In some series analyses of radical prostatectomy specimens suggest a strong correlation between a positive RT-PCR result and capsular invasion by the tumor, while others do not support its use in determining pathological stage. In the majority of reports the RT-PCR assay was highly specific for detection of extraprostatic PSA expression in prostate cancer patients, and negative for detection in men with benign prostatic hyperplasia and in women. Sources of potential false-positive and false-negative results of this assay are identified and discussed. CONCLUSIONS: RT-PCR can detect PSA expressing cells that are otherwise undetectable by other means in patients with localized and metastatic cancers. This assay is highly specific, since PSA expressing cells were consistently undetectable in the peripheral circulation of patients without prostate cancer in most studies. Limited data to date suggest that this test may have a role in the staging of tumors before radical prostatectomy and in the serial followup of patients after treatment. RT-PCR may improve the detectability of lymph node metastasis over immunohistochemistry. Presently this test should remain a powerful research tool in the study of the biology and behavior of prostate cancer, and it should not be used to guide any clinical decision making. The use of this assay as a prognostic and management tool for prostate cancer is in the earliest stages.

The transcription factor E2F-1 modulates TGF-beta1 RNA expression in glial cells.

The cell type specificity of the regulation of expression of the potent growth inhibitory cytokine transforming growth factor-beta (TGF-beta), prompted our analyses of the regulation of TGF-beta1 gene expression in glial cells by viral and cellular oncoproteins. We have shown that SV40 T-antigen diminished TGF-beta1 expression in glial cells and this repression was dependent on the ability of T-antigen to interact with the tumor suppressor protein, pRb, and two structurally related proteins, p107 and p130. The cellular transcription factor E2F-1, which is a downstream effector of T-antigen, was unable to influence expression from the TGF-beta1 promoter by itself. Interestingly, E2F-1 could overcome viral T-antigen-mediated repression of the TGF-beta1 promoter, suggesting potential feedback loop between TGF-beta and E2F in virally transformed glial cells. Using deletion analyses, we have mapped two E2F-1-responsive regions on the TGF-beta1 promoter: a T-antigen-dependent negative regulatory sequence (TdNRS) between -323 and -175, and a T-antigen-independent positive regulatory sequence (TiPRS) between -34 and +10 on the TGF-beta1 promoter. Further examination of TiPRS revealed the presence of a functional E2F binding site. Interestingly, the amino terminus of E2F-1 was required for its activation of TGF-beta1 expression, as mutations in that domain abolished the ability of E2F-1 to increase TGF-beta1 expression. These data suggest that yet-uncharacterized interaction between the amino terminus of E2F-1 and cellular proteins regulates TGF-beta1 expression. The mechanism for E2F-1-mediated T-antigen-dependent regulation of TGF-beta1 expression from TdNRS awaits further characterization.

Soluble factors secreted by activated T-lymphocytes modulate the transcription of the immunosuppressive cytokine TGF-beta 2 in glial cells.

Coordination of the immune response to injury or disease in the brain is postulated to involve bi-directional discourse between the immune system and the central nervous system. This cross communication involves soluble mediators, including various growth factors, cytokines, and neuropeptides. In this report, we demonstrate that the supernatant from activated T-lymphocytes is able to induce the transcription of a potent cytokine, TGF-beta 2 in glial cells. The activating stimulus invokes signaling mechanisms distinct from known kinase or protease pathways. Activation of TGF-beta 2 transcription correlates with the loss of binding activity for an 80 kDA glial labile repressor protein, GLRP, to a responsive region within the TFG-beta 2 promoter. Although GLRP shares some characteristics with the inducible transcription factor AP-1, it appears to be distinct from known AP-1 family members. These data along with previous observations demonstrating the potent immunosuppressive activity of TGF-beta 2, support a model for a feedback mechanism between the activated T-lymphocytes and astrocytes via TGF-beta 2 to regulate the immune response.

Transcriptional regulation of human polyomavirus JC: evidence for a functional interaction between RelA (p65) and the Y-box-binding protein, YB-1.

The transcriptional control region of the human neurotropic polyomavirus JC virus contains a consensus NF-kappa B site which has been shown to enhance both basal and extracellular stimulus-induced levels of transcription of JC promoters. Here, we show that the expression of JC late promoter constructs containing the NF-kappa B site is decreased by cotransfection with the NF-kappa B/rel subunits, p50 and p52, but enhanced by the p65 subunit. However, JC promoter constructs lacking the NF-kappa B site were activated by p52 and p50 and repressed by p65. This antithetical response of the JC promoter mapped specifically to the D domain, which is a target site for the cellular transcription factor, YB-1. Band shift studies indicated that YB-1 and p65 modulate each other's binding to DNA: YB-1 augments the affinity of p65 for the NF-kappa B site, while p65 reduces the binding of YB-1 to the D domain. Results from coimmunoprecipitation followed by Western blot (immunoblot) analysis suggest an in vivo interaction between p65 and YB-1 in glial cells. Functionally, YB-1 appears to act synergistically with p65 to control transcription from the NF-kappa B site. A converse pattern is seen with the D domain, in which YB-1 acts synergistically with p50 and p52 to regulate transcription. p50 and p52 may function as transcriptional activators on the D domain by removing the repressive effect of p65 on YB-1 binding to the D domain. On the basis of these data, we propose a model in which NF-kappa B/rel subunits functionally interact with consensus NF-kappa B sites or YB-1-binding sites, with disparate effects on eukaryotic gene expression.

Evidence for GEAPS, novel Glial E2F1-Associated Proteins in hamster glioma cells induced by the human neurotropic polyomavirus, JCV.

Injection of the human neurotropic polyomavirus, JCV, into neonatal hamsters causes tumors of glial origin. Previously, a rapidly proliferating cell line, HJC-15, which expresses high levels of the viral T-antigen, had been established from JCV-induced hamster glial tumors. In our analyses of the mechanisms involved in the control of glial cell proliferation in these tumor cells, we have focused our attention on E2F1, a DNA-binding transcription factor which modulates the activity of genes involved in the S-phase of the cell cycle. Here, we report the identification of a novel nucleo-protein complex that forms between select E2F1-binding sites and nuclear proteins from HJC-15 and normal hamster glial cells. In comparison to the previously characterized E2F1 complexes, this complex exhibited distinct mobility, binding and biochemical characteristics. This slower migrating complex also contained several unique Glial E2F1-associated proteins, (GEAP), which have a distinct molecular mass. Of particular, unlike the classical E2F1 whose DNA binding activity is increased during S-phase, the level GEAPs remained constant throughout the cell cycle. GEAPs appeared to confer an increased basal transcriptional activity of promoters containing select E2F1 sites in HJC-15 cells. Interestingly, the increased transcriptional activity modulated by GEAPs in HJC-15 cells was overcome by overexpression of E2F1 in these cells. These data point to the presence of novel members of the E2F family in hamster glial cells with the potential to regulate expression of S-phase specific genes in glial tumors obtained upon intracerebral injection of JCV. The importance of these findings in the pathogenesis of viral-induced tumors and the role of cell cycle regulatory proteins in brain tumor formation is discussed.

Identification and characterization of a novel GGA/C-binding protein, GBP-i, that is rapidly inducible by cytokines.

Immunosuppressive states with accompanying alterations in cytokine profiles have been postulated to play a vital role in the reactivation of viruses from latency. Cytokines regulate gene expression by activating transcription factors via well-characterized signal transduction pathways. In this study, we report the identification of a novel inducible protein, GBP-i, that binds to a double-stranded GGA/C-rich region of the transcriptional control region of the human papovavirus JC virus (JCV), specifically within the origin of viral DNA replication. GBP-i is distinct from previously characterized GC-box-binding proteins with respect to both its sequence specificity and its electrophoretic mobility on native and denaturing gels. GBP-i responds within 90 min to phorbol myristate acetate stimulation; however, unlike typical phorbol myristate acetate-inducible factors, this rapid induction is regulated primarily at the transcriptional level. Further, the induction of GBP-i appears to be widespread and mediated by many inflammatory cytokines, including interleukin-1 beta, tumor necrosis factor alpha, gamma interferon, and transforming growth factor beta. Interestingly, the induced protein acts as a transcriptional repressor in its native context in the JCVL promoter. However, when its binding sequence is transposed to a heterologous promoter, GBP-i appears to function as a transcriptional activator. The data presented here suggest a role for GBP-i in cytokine-mediated induction of viral and cellular genes.

Central nervous system-derived cells express a kappa B-binding activity that enhances human immunodeficiency virus type 1 transcription in vitro and facilitates TAR-independent transactivation by Tat.

The Tat protein of human immunodeficiency virus type 1 (HIV-1) is a potent activator of long terminal repeat-directed transcription. While in most cell types, activation requires interaction of Tat with the unusual transcription element TAR, astrocytic glial cells support TAR-independent transactivation of HIV-1 transcription by Tat. This alternative pathway of Tat activation is mediated by the viral enhancer, a kappa B domain capable of binding the prototypical form of the transcription factor nuclear factor kappa B (NF-kappa B) present in many cell types, including T lymphocytes. Tat transactivation mediated by the kappa B domain is sufficient to allow replication of TAR-deleted mutant HIV-1 in astrocytes. The present study demonstrates the existence of kappa B-specific binding factors present in human glial astrocytes that differ from prototypical NF-kappa B. The novel astrocyte-derived kappa B-binding activity is retained on an HIV-1 Tat affinity column, while prototypical NF-kappa B from Jurkat T cells is not. In vitro transcription studies demonstrate that astrocyte-derived kappa B-binding factors activate transcription of the HIV-1 long terminal repeat and that this activation is dependent on the kappa B domain. Moreover, TAR-independent transactivation of HIV-1 transcription is reproduced in vitro in an astrocyte factor-dependent manner which correlates with kappa B-binding activity. The importance of the central nervous system-enriched kappa B transcription factor in the regulation of HIV-1 expression is discussed.