D77, one benzoic acid derivative, functions as a novel anti-HIV-1 inhibitor targeting the interaction between integrase and cellular LEDGF/p75

Integration of viral-DNA into host chromosome mediated by the viral protein HIV-1 integrase (IN) is an essential step in the HIV-1 life cycle. In this process, Lens epithelium-derived growth factor (LEDGF/p75) is discovered to function as a cellular co-fa

Expression pattern, cellular localization and promoter activity analysis of ovarian aromatase (Cyp19a1a) in protogynous hermaphrodite red-spotted grouper

Aromatase plays a key role in sex differentiation of gonads. In this study, we cloned the full-length cDNA of ovarian aromatase from protogynous hermaphrodite red-spotted grouper (Epinephelus akaara), and prepared the corresponding anti-EaCyp19a1a antiserum. Western blot and immunofluorescence studies revealed ovary-specific expression pattern of EaCyp19a1a in adults and its dynamic expression change during artificial sex reversal. EaCyp19a1a was expressed by follicular cells of follicular layer around oocytes because strong EaCyp19a1a immunofluorescence was observed in the cells of ovaries. During artificial sex reversal, EaCyp19a1a expression dropped significantly from female to male, and almost no any positive EaCyp19a1a signal was observed in testicular tissues. Then, we cloned and sequenced a total of 1967 bp T-flanking sequence of EaCyp19a1a promoter, and showed a number of potential binding sites for some transcriptional factors, such as SOX5, GATA gene family, CREB, AP1, FOXL1, C/EBP, ARE and SF-1. Moreover, we prepared a series of 5' deletion promoter constructs and performed in vitro luciferase assays of EaCyp19a1a promoter activities. The data indicated that the CREB regulation region from -1010 to -898 might be a major cis-acting element to EaCyp19a1a promoter, whereas the elements GATA and SOX5 in the region from -1216 to -1010 might be suppression elements. Significantly, we found a common conserved sequence region in the fish ovary-type aromatase promoters with identities from 93% to 34%. And, the motifs of TATA box, SF-1, SOX5, and CREB existed in the region and were conserved among the most of fish species. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Expression pattern of cellular nucleic acid-binding protein (CNBP) during embryogenesis and spermatogenesis of gibel carp

By differential screening, we cloned the CagCNBP, demonstrated its predominant expression in ovary and testis, and reported its development behavior during folliculogenesis and oogenesis by immunofluorescence localization (Liu and Gui, Gene 365:181-192, 2005), but its developmental behavior during spermatogenesis and its transcript distribution during embryogenesis are not revealed. In the present study, by in situ hybridization, we analyze CagCNBP expression pattern during gibel carp embryogenesis. The CagCNBP transcripts ubiquitously distributed in all embryonic cells in early developmental stage embryos, and peak in midbrain, hindbrain and somites of gibel carp larva during organogenesis. By antibody detection, we reveal CagCNBP protein distribution change during spermatogenesis. The cell-specific distribution of CagCNBP is revealed by immunofluorescence staining, and predominant CagCNBP expression in testis somatic cells and spermatogonia is demonstrated in this paper. For the first time, the CNBP distribution during spermatogenesis in vertebrate has been revealed.

Molecular characterization of Chinese sturgeon gonadotropins and cellular distribution in pituitaries of mature and immature individuals

Chinese sturgeon (Acipenser sinensis) is a rare and endangered species, and also an important resource for the sturgeon aquaculture industry. To understand molecular characterization of Chinese sturgeon gonadotropins (GTHs), we cloned the full-length cDNAs of gonadotropin subunits common alpha (GTH-alpha), follicle-stimulating hormone (FSH) and luteinizing hormone (LH) from a pituitary cDNA library of mature female. Two subtypes of GTH-alpha were identified. The nucleotide sequences of A. sinensis common alpha I (AsGTH-alpha I), common alpha II (AsGTH-alpha II), FSH beta (AsFSH beta) and LH beta (AsLH beta) subunit cDNAs are 345, 363, 387 and 414 bp in length, and encode mature peptides of 115, 121, 129 and 138 aa, respectively. Then, three polyclonal antibodies were prepared from the in vitro expressed AsGTH-alpha I, AsFSH beta and AsLH beta mature proteins, respectively. Significant expression differences were revealed between immature and mature sturgeon pituitaries. Western blot detection and immunofluoresence localization revealed the existence of three-gonadotropin subunits (AsGTH-alpha, AsFSH beta and AsLH beta) in mature sturgeon pituitaries, but only AsFSH beta was detected in immature individual pituitaries during early stages in the sturgeon life, and obvious difference was observed between males and females. In males, AsFSH beta was expressed in 4-year-old individuals, whereas in females, AsFSH beta was just expressed in 5-year-old individuals. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Predominant expression and cellular distribution of fish Agr2 in renal collecting system

Anterior gradient 2 (Agr2) genes encode secretory proteins, and play significant roles in anterior-posterior patterning and tumor metastasis. Agr2 transcripts were shown to display quite diverse tissue distribution in different species, and little was known about the cellular localization of Agr2 proteins. In this study, we identified an Agr2 homologue from gibe[ carp (Carassius auratus gibelio), and revealed the expression patterns and cellular localization during embryogenesis and in adult tissues. The full-length cDNA of CagAgr2 is 803 nucleotides (nt) with an open reading frame of 510 nt encoding 169 amino acids. The Agr2 C-terminus matches to the class I PDZ-interacting motif, suggesting that it might be a PDZ-binding protein. During embryogenesis, CagAgr2 was found to be transcribed in the mucus-secreting hatching gland from tailbud stage and later in the pharynx region, swim bladder and pronephric duct as revealed by RT-PCR and whole mount in situ hybridization. In the adult fish, its transcription was predominantly confined to the kidney, and lower transcription levels were also found in the intestine, ovary and gills. To further localize the Agr2 protein, the anti-CagAgr2 polyclonal antibody was produced and used for immunofluorescence observation. In agreement with mRNA expression data, the Agr2 protein was localized in the pronephric duct of 3dph larvae. In adult fish, Agr2 protein expression is confined to the renal collecting system with asymmetric distribution along the apical-basolateral axis. The data provided suggestive evidence that fish Agr2 might be involved in differentiation and secretory functions of kidney epithelium. (C) 2009 Elsevier Inc. All rights reserved.

Optimization by orthogonal array design and humoral immunity of the bivalent vaccine against Aeromonas hydrophila and Vibrio fluvialis infection in crucian carp (Carassius auratus L.)

Aeromonas hydrophila and Vibrio fluvialis are the causative agents of a serious haemorrhagic septicaemia that affects a wide range of freshwater fish in China. In order to develop a bivalent anti-A. hydrophila and anti-V. fluvialis formalin-killed vaccine to prevent this disease, an orthogonal array design (OAD) method was used to optimize the production conditions, using three factors, each having three levels. The effects of these factors and levels on the relative per cent survival for crucian carp were quantitatively evaluated by analysis of variance. The final optimized formulation was established. The data showed that inactivation temperature had a significant effect on the potency of vaccine, but formalin concentration did not. The bivalent vaccine could elicit a strong humoral response in crucian carp (Carassius auratus L.) against both A. hydrophila and V. fluvialis simultaneously, which peaked at 3 or 5 weeks respectively. Antibody titres remained high until week 12, the end of the experiment, after a single intraperitoneal injection. The verification experiment confirmed that an optimized preparation could provide protection for fish at least against A. hydrophila infection, and did perform better than the non-optimized vaccine judged by the antibody levels and protection rate, suggesting that OAD is of value in the development of improved vaccine formulations.

Expression pattern and developmental behaviour of cellular nucleic acid-binding protein (CNBP) during folliculogenesis and oogenesis in fish

In vertebrates, folliculogeneis establishes an intricate system for somatic cell-oocyte interaction, and ultimately leads to the acquisition of their respective competences. Although the formation process and corresponding interactions are strikingly similar in diverse organisms, knowledge of genes and signaling pathways involved in follicle formation is very incomplete and the underlying molecular mechanisms remain enigmatic. CNBP has been identified for more than ten years, and the highest level of CNBP transcripts has been observed in adult zebrafish ovary, but little is known about its functional significance during folliculogeneis and oogenesis. In this study, we clone CNBP cDNA from gibel carp (Carassius auratus gibelio), and demonstrate its predominant expression in gibel carp ovary and testis not only by RTPCR but also by Western blot. Its full-length cDNA is 1402 bp, and has an ORF of 489 nt for encoding a peptide of 163 aa. And its complete amino acid sequence shared 68.5%-96.8% identity with CNBPs from other vertebrates. Based on the expression characterization, we further analyze its expression pattern and developmental behaviour during folliculogeneis and oogenesis. Following these studies, we reveal an unexpected discovery that the CagCNBP is associated with follicular cells and oocytes, and significant distribution changes have occurred in degenerating and regenerating follicles. More interestingly, the CagCNBP is more highly expressed in some clusters of interconnected cells within ovarian cysts, no matter whether the cell clusters are formed from the original primordial germ cells or from the newly formed cells from follicular cells that invaded into the atretic oocytes. It is the first time to reveal CNBP relevance to folliculogeneis and oogenesis. Moreover, a similar stage-specific and cell-specific expression pattern has also been observed in the gibel carp testis. Therefore, further studies on CNBP expression pattern and developmental behaviour will be of significance for understanding functional roles of CNBP during gametogenests. (c) 2005 Elsevier B.V. All rights reserved.

Studies on morphogenesis and cellular interactions of Rana grylio virus in an infected fish cell line

A pathogenic virus (RGV), isolated from diseased pig frog Rana grylio with lethal syndrome, was investigated with regard to morphogenesis and cellular interactions in EPC cells, a cell Line from fish. Different stages of virus amplification, maturation and assembly were observed at nucleus, cytoplasm and cellular membranes. The matured virus particles, were not only distributed diffusely in nucleus, cytoplasm and cellular surface, but also aggregated as pseudocrystalline arrays in the cytoplasm. Virions were released by budding from the plasma membranes, or following cell lysis. Various types of cell damage, such as small vacuoles, spherical inclusions, and swollen and empty mitochondria, were also found. Some typical characteristics of RGV, such as the symmetrical shape of the virions, replication process involving both nuclear and cytoplasmic phases, budding release from cellular membrane and intracellular membrane, viromatrix and paracrystalline aggregation in cytoplasm, and its acute pathogenic effects, were observed to be similar to that of other iridoviruses. Therefore, the RGV appears to be a member of the Iridoviridae based on these studies. (C) 1999 Elsevier Science B.V. All rights reserved.

Single-electron tunneling depressing synapse for cellular neural networks

In this paper, a cellular neural network with depressing synapses for contrast-invariant pattern classification and synchrony detection is presented, starting from the impulse model of the single-electron tunneling junction. The results of the impulse model and the network are simulated using simulation program with integrated circuit emphasis (SPICE). It is demonstrated that depressing synapses should be an important candidate of robust systems since they exhibit a rapid depression of excitatory postsynaptic potentials for successive presynaptic spikes.

Detector-induced dephasing in quantum-dot cellular automata qubit

We investigate the quantum dynamics of the quantum-dot cellular automata qubit in the presence of a quantum point contact detector by modified rate equations. It is demonstrated that the qubit information can be resolved by measuring the detector current variation. Furthermore, we show that this oscillating current and the electron occupation probabilities in states \b> and \c> decay drastically as the dephasing rate increases, clearly revealing the influence of the dephasing induced by the detector. Moreover, it is shown that the operation speed of the quantum-dot cellular automata qubit may be adjusted by varying the interdot coupling strength. (C) 2003 American Institute of Physics.

Potential role of DNA-dependent protein kinase in cellular resistance to ionizing radiation

In this paper, we study the ability of DNA-PK-deficient (M059J) and -proficient (M059K) cells to undergo the rate of cellular proliferation, cell cycle distribution and apoptosis after 10 Gy X-ray irradiation, and the role of DNA-PK in radiosensitivity. The results showed that M059J cells exhibited hyper-radiosensitivity compared with M059K cells. A strong G2 phase arrest was observed in M059J cells post irradiation. Significant accumulation in the G2 phase in M059J cells was accompanied by apoptosis at 12 h. Altogether, the data suggested that DNA-PK may have two roles in mammalian cells after DNA damage, a role in DNA DSB repair and a second role in DNA-damaged cells to traverse a G2 checkpoint, by which DNA-PK may affect cellular sensitivity to ionizing radiation. 地址: [Li Ning; Zhang Hong; Wang Yanling; Hao Jifang] Chinese Acad Sci, Inst Modern Phys, Lanzhou 730000, Peoples R China; [Li Ning; Zhang Hong; Wang Yanling; Hao Jifang] Key Lab Heavy Ion Radiat Med Gansu Prov, Lanzhou 730000, Peoples R China; [Li Ning; Wang Yanling] Chinese Acad Sci, Grad Sch, Beijing 100039, Peoples R China; [Wang Xiaohu] Gansu Tumor Hosp, Dept Radiotherapy, Lanzhou 730050, Peoples R China

Effects of Ionizing Radiation on Three-Dimensional Human Vessel Models: Differential Effects According to Radiation Quality and Cellular Development

Little is known about the effects of space radiation on the human body. There are a number of potential chronic and acute effects, and one major target for noncarcinogenic effects is the human vasculature. Cellular stress, inflammatory response, and other radiation effects on endothelial cells may affect vascular function. This study was aimed at understanding the effects of space ionizing radiation on the formation and maintenance of capillary-like blood vessels. We used a 3D human vessel model created with human endothelial cells in a gel matrix to assess the effects of low-LET protons and high-LET iron ions. Iron ions were more damaging and caused significant reduction in the length of intact vessels in both developing and mature vessels at a dose of 80 cGy. Protons had no effect on mature vessels up to a dose of 3.2 Gy but did inhibit vessel formation at 80 cGy. Comparison with gamma radiation showed that photons had even less effect, although, as with protons, developing vessels were more sensitive. Apoptosis assays showed that inhibition of vessel development or deterioration of mature vessels was not due to cell death by apoptosis even in the case of iron ions. These are the first data to show the effects of radiation with varying linear energy transfer on a human vessel model. (C) 2011 In Radiation Research Society

Modeling of Hillslope Runoff and Soil Erosion at Rainfall Events Using Cellular Automata Approach

National Science Fund for Distinguished Young Scholars of China [40225004]; National Natural Science Foundation of China [40471048]