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At least 595 records · Page 33Linked to original sources
PubMed1997

Epstein-Barr virus: biology and disease.

Epstein-Barr virus is a human herpes virus which, whilst found as a widespread asymptomatic infection, is also associated with certain tumours of lymphoid and epithelial origin including Burkitt's lymphoma (BL), immunoblastic lymphoma (IBL), Hodgkin's Disease (HD) and nasopharyngeal carcinoma (NPC). A unique characteristic of EBV is its ability to infect and transform primary resting B lymphocytes in vitro into permanently growing lymphoblastoid cell lines (LCLs); this effect is associated with constitutive expression of a limited set of viral genes. Interestingly, the pattern of EBV gene expression observed in LCLs in vitro is also a feature of IBLs, a tumour associated with immunosuppression. The other EBV associated tumours display a more restricted pattern of EBV latent protein expression. B cell lines can be activated in vitro into the virus replicative cycle, where a large number of viral genes associated with EBV DNA replication and virus assembly are synthesised. Whilst EBV can be detected in throat washings from seropositive individuals, the only in vivo situation where full virus replication can be reliably observed in hairy leukoplakia (HL), a benign lesion of lingual epithelium frequently found in AIDS patients. Thus, the relative contribution of lymphoid cells and epithelial cells to latent EBV infection/persistence vs replication in vivo remains controversial. Recent studies suggest that HL represents a focus of EBV replication in the absence of a truly latent infection and this supports the contention that EBV persistence resides in the lymphoid compartment. These aspects together with the role of EBV in oral diseases and the effect of certain EBV genes on the control of epithelial cell growth and differentiation will be discussed.

Carcinoma↗
PubMed1999

Genetic interaction of flavivirus nonstructural proteins NS1 and NS4A as a determinant of replicase function.

Nonstructural protein 1 (NS1) of yellow fever virus (YF) is a glycoprotein localized to extracytoplasmic compartments within infected cells. We have previously shown that NS1 can be supplied in trans and is required for viral RNA replication, a process thought to occur in membrane-bound cytoplasmic complexes. Here we report that the NS1 gene from a related virus, dengue virus (DEN), is unable to function in the process of YF RNA replication. This virus-specific incompatibility leads to a lack of initial minus-strand accumulation, suggesting that DEN NS1 is unable to productively interact with the YF replicase. Based on a YF deletion mutant that requires NS1 in trans, a genetic screen for suppressor mutants was used to select virus variants able to utilize DEN NS1. In three independent selections, a single mutation was mapped to the NS4A gene, which encodes a putative transmembrane replicase component. This mutation, as well as several additional mutations, was engineered into the NS1-deficient genome and confirmed a genetic interaction between NS1 and NS4A. These findings suggest a potential mechanism for integrating NS1 into the cytoplasmic process of RNA replication.

Amino Acid Sequence↗
PubMed2000

Bcl-2 blocks a caspase-dependent pathway of apoptosis activated by herpes simplex virus 1 infection in HEp-2 cells.

Earlier reports have shown that herpes simplex virus 1 (HSV-1) mutants induce programmed cell death and that wild-type virus blocks the execution of the cell death program triggered by expression of viral genes, by the Fas and tumor necrosis factor pathways, or by nonspecific stress agents. In particular, an earlier report from this laboratory showed that the mutant virus d120 lacking the genes encoding infected cell protein 4 (ICP4), the major regulatory protein of the virus, induces a caspase-3-independent pathway of apoptosis in human SK-N-SH cells. Here we report that the pathway of apoptosis induced by the d120 mutant in human HEp-2 cells is caspase dependent. Specifically, in HEp-2 cells infected with d120, (i) a broad-range inhibitor of caspase activity, z-vad-FMK, efficiently blocked DNA fragmentation, (ii) cytochrome c was released into the cytoplasm, (iii) caspase-3 was activated inasmuch as poly(ADP-ribose) polymerase was cleaved, and (iv) chromatin condensation and fragmentation of cellular DNA were observed. In parallel studies, HEp-2 cells were transfected with a plasmid encoding human Bcl-2 and a clone (VAX-3) expressing high levels of Bcl-2 was selected. This report shows that Bcl-2 blocked all of the manifestations associated with programmed cell death caused by infection with the d120 mutant. Consistent with their resistance to programmed cell death, VAX-3 cells overproduced infected cell protein 0 (ICP0). An unexpected observation was that ICP0 encoded by the d120 mutant accumulated late in infection in small, quasi-uniform vesicle-like structures in all cell lines tested. Immunofluorescence-based colocalization studies indicated that these structures were not mitochondria or components of the endoplasmic reticulum or the late endosomal compartment. These studies affirm the conclusion that HSV can induce programmed cell death at multiple steps in the course of its replication, that the d120 mutant can induce both caspase-dependent and -independent pathways of programmed cell death, and that virus-induced stimuli of programmed cell death may differ with respect to the pathway that they activate.

Amino Acid Chloromethyl Ketones↗
PubMed2001

Environmental blue light prevents stress in the fish Nile tilapia.

The present study aimed to test the effects of blue, green or white light on the stress response of the Nile tilapia, Oreochromis niloticus (L.). Each color was tested on two groups of isolated adult Nile tilapia (8 replicates each): one being subjected to confinement stress, and the other not (control). A different environmental color was imposed on each compartment by covering the light source with cellophane of the respective color (green or blue; no cellophane was used for white light). The intensity of green, white and blue lights was 250, 590 and 250 lux, respectively. Basal plasma cortisol levels were determined for each fish prior to the experimental procedures. The fish were confined by being displaced toward one side of the aquarium using an opaque partition for 1 h both in the morning and the afternoon of the two consecutive days of the test. At the end of this 48-h period, plasma cortisol levels were measured again. Basal cortisol levels (ng/ml) were similar for each group (ANOVA, F(2;42) = 0.77, P = 0.47). Thus, plasma cortisol levels were analyzed in terms of variation from their respective basal level. After confinement, plasma cortisol levels were not increased in fish submitted to a blue light environment. Thus, blue light prevents the confinement-induced cortisol response, an effect not necessarily related to light intensity.

Analysis of Variance↗
PubMed1999

Murine stromal cells counteract the loss of long-term culture-initiating cell potential induced by cytokines in CD34(+)CD38(low/neg) human bone marrow cells.

Evidence has been provided recently that shows that high concentrations of cytokines can fulfill functions previously attributed to stromal cells, such as promote the survival of, and led to a net increase in human primitive progenitors initiating long-term cultures in vitro (LTC-IC) or engrafting NOD-SCID (nonobese diabetic severe-combined immunodeficient) recipients in vivo. These data prompted us to re-evaluate whether stromal cells will further alter the properties of primitive progenitor cells exposed to cytokines. Single CD34(+)CD38(low) and CD38(neg) cells were incubated 10 days in serum-containing or serum-free medium in the presence or in the absence of murine marrow-derived stromal cells (MS-5). Recombinant human cytokines stem cell factor (SCF), pegylated-megakaryocyte growth and differentiation factor (PEG-MGDF), FLT3-L, Interleukin (IL)-3, IL-6, and granulocyte-macrophage colony-stimulating factor (GM-CSF) were systematically added at various concentrations (10 to 300 ng/mL). Cell proliferation and LTC-IC potential were evaluated in each clone after 10 days. A striking and consistent observation was the retention of a high LTC-IC potential in clones exposed to cytokines in the presence of stromal feeders, whereas clones exposed to cytokines alone in the absence of stromal feeders rapidly lost their LTC-IC potential as they proliferated. This was reflected both by the higher proportion of wells containing LTC-IC and by the high numbers of CFC produced after 5 weeks in clones grown with MS-5 during the first 10 days. We further showed by analyzing multiple replicates of a single clone at day 10 that MS-5 cells promoted a net increase in the LTC-IC compartment through self-renewal divisions. Interestingly, these primitive LTC-IC were equally distributed among small and large clones, as counted at day 10, indicating that active proliferation and loss of LTC-IC potential could be dissociated. These observations show that, in primitive cells, stromal cells counteract differentiation events triggered by cytokines and promoted self-renewal divisions. Furthermore, the almost identical distribution of the size of the clones with or without MS-5 suggests that proliferation and function of human primitive cells may be independently regulated by external signals, and that the former is primarily under the control of cytokines.

ADP-ribosyl Cyclase↗
PubMed2003

Viruses as hijackers of PML nuclear bodies.

Promyelocytic leukemia nuclear bodies (PML-NBs) are discrete interchromosomal macromolecular structures. The integrity of this dynamic nuclear subcompartment critically depends on the presence of the name-giving PML protein. Among the permanent or transient residents of PML-NBs are various regulatory proteins, including Sp100, CBP, pRb, HIPK2, RAD51 and p53. PML-NBs are frequently targeted by viral infections, as a number of different RNA and DNA viruses, including herpesviruses, adenoviruses, papovaviruses, papillomaviruses and arenaviruses, cause changes in PML-NBs. Viruses interfere with PML-NB in two ways: 1) some viral proteins can associate with PML-NB proteins and/or lead to the destruction and lysis of this subnuclear compartment, thus aiding viral gene expression and disabling the host's innate immunity; 2) the parental genomes of some nuclear-replicating DNA viruses associate preferentially with PML-NBs, which presumably serves to assist in viral gene expression or replication. Here we feature the different viral strategies leading to the hijacking of PML-NBs and discuss the consequences for the immune response.

Animals↗
PubMed1982

Friend erythroleukemia virus complex: role of viral components in modifying erythroid differentiation in mice.

The present study was carried out to determine which component of the polycythemic strain of Friend erythroleukemia virus (FV-P)--Friend helper virus [Friend murine leukemia virus (F-MuLV)] of the replication-defective spleen focus-forming virus (SFFV)--is responsible for inducing erythropoietin-independent erythropoiesis and expansion of the erythroid compartment in infected mice. F-MuLV and SFFV were cloned in SC1 cells to give a cell line 643/22N, which released only F-MuLV and SC204, a nonproducer cell line carrying SFFV. On superinfection with specific helper viruses, SC204 yielded further cell lines, which released SFFV in conjunction with 1) Friend, 2) Moloney, or 3) Gross helper viruses. With the use of an in vitro colony formation assay to monitor erythropoiesis in the bone marrow of inbred DBA/2J mice infected with any of the virus preparations containing SFFV plus helper virus, the erythroid precursor cell population was found to become erythropoietin-independent and amplified; adult mice infected with any of the helper viruses alone did not develop any symptoms of Friend disease. Thus the biologic activity of the FV-P complex was unaffected by replacement of the F-MuLV with an unrelated helper virus (Moloney or Gross). These results indicated that the SFFV component of FV-P is responsible for modifying erythroid differentiation in adult mice.

Animals↗
PubMed1978

Erythropoietin responsiveness in vivo of Friend virus target cells for transformation and virus replication.

Techniques that allow the selective stimulation of the erythropoietin-responsive cell population in mice with suppressed multipotential hemopoietic stem cells were used to identify (1) the in vivo target cell transformed by Friend virus (FV) into a tumor colony-forming unit and (2) a target cell for FV replication in vivo. Plethoric mice with busulphan-induced reductions in stem cell populations (characterized as colony-forming units) and stimulated erythropoietin-responsive cell compartments were given FV; control groups, not receiving erythropoietin, also received FV. A comparison of the number of target cells transformed in each group provided evidence identifying the ERC as the in vivo compartment in which the target cell detected by tumor colony formation resides. Differences in plasma virus titers revealed that the erythropoietin-responsive cell is also predominantly responsible for the production of FV as measured by focus-forming activity.

Animals↗
PubMed1999

Human granulocytic ehrlichiosis agent and Ehrlichia chaffeensis reside in different cytoplasmic compartments in HL-60 cells.

The human granulocytic ehrlichiosis (HGE) agent resides and multiplies exclusively in cytoplasmic vacuoles of granulocytes. Double immunofluorescence labeling was used to characterize the nature of the HGE agent replicative inclusions and to compare them with inclusions containing the human monocytic ehrlichia, Ehrlichia chaffeensis, in HL-60 cells. Although both Ehrlichia spp. can coinfect HL-60 cells, they resided in separate inclusions. Inclusions of both Ehrlichia spp. were not labeled with either anti-lysosome-associated membrane protein 1 or anti-CD63. Accumulation of myeloperoxidase-positive granules were seen around HGE agent inclusions but not around E. chaffeensis inclusions. 3-(2, 4-Dinitroanilino)-3'-amino-N-methyldipropylamine and acridine orange were not localized to either inclusion type. Vacuolar-type H+-ATPase was not colocalized with HGE agent inclusions but was weakly colocalized with E. chaffeensis inclusions. E. chaffeensis inclusions were labeled with the transferrin receptor, early endosomal antigen 1, and rab5, but HGE agent inclusions were not. Some HGE agent and E. chaffeensis inclusions colocalized with major histocompatibility complex class I and II antigens. These two inclusions were not labeled for annexins I, II, IV, and VI; alpha-adaptin; clathrin heavy chain; or beta-coatomer protein. Vesicle-associated membrane protein 2 colocalized to both inclusions. The cation-independent mannose 6-phosphate receptor was not colocalized with either inclusion type. Endogenously synthesized sphingomyelin, from C6-NBD-ceramide, was not incorporated into either inclusion type. Brefeldin A did not affect the growth of either Ehrlichia sp. in HL-60 cells. These results suggest that the HGE agent resides in inclusions which are neither early nor late endosomes and does not fuse with lysosomes or Golgi-derived vesicles, while E. chaffeensis resides in an early endosomal compartment which accumulates the transferrin receptor.

Animals↗
PubMed2000

Uneven distribution of hepatitis C virus quasispecies in tissues from subjects with end-stage liver disease: confounding effect of viral adsorption and mounting evidence for the presence of low-level extrahepatic replication.

We have found differences among the populations of hepatitis C virus sequences in serum, peripheral blood mononuclear cells (PBMCs), and various tissues in patients with chronic hepatitis C. These results are compatible with the existence of independent viral compartments in the infected host. Our results also suggest that PBMCs, and probably various tissues, can selectively adsorb viral subpopulations differing in the E2 region.

5' Untranslated Regions↗
PubMed2004

Human cytomegalovirus UL76 encodes a novel virion-associated protein that is able to inhibit viral replication.

The human cytomegalovirus (HCMV) UL76 gene encodes a highly conserved herpesvirus protein, pUL76, which is able to modulate gene expression in either activation or repression. In this study, two specific transcripts were found to contain the reading frame of UL76, one a 4.5-kb and the other a 5.5-kb tricistronic mRNA encoding the UL76, UL77, and UL78 open reading frames. Both transcripts were expressed with true late kinetics, as revealed by data showing inhibition of production in the presence of phosphonoformic acid. Immediately after viral infection, pUL76 was found in the nuclear fraction and was detected in cells in the presence of the protein synthesis inhibitor cycloheximide. Subsequent virus particle purification and Western blot analysis revealed that two forms of pUL76 are associated within mature virions. The high-molecular-mass protein (H-pUL76) was verified as originating from a free form of pUL76 by cross-linking with an unknown protein(s). By performing a biochemical fractionation experiment with purified virions, we provide evidence that pUL76 and H-pUL76 are associated with the detergent-soluble (envelope) and -insoluble (tegument/capsid) fractions, respectively. Both results were consistent with the images exhibited by immunoelectron microscopy, which showed that the distribution of gold particles labeled by the anti-pUL76 antibody juxtaposed the compartments of the envelope and the tegument/capsid of the virion. Evidence indicated that expression of pUL76 at the immediate-early phase of the viral replication cycle leads to the inhibition of HCMV production. The viral constituent pUL76, with a dominant-negative effect on replication, may provide a novel mechanism for HCMV's resumption of latency.

Cell Line↗
PubMed1993

Patterns of frog virus 3 DNA methylation and DNA methyltransferase activity in nuclei of infected cells.

The iridovirus frog virus 3 (FV3) can replicate in culture in fat head minnow (FHM) fish cells or in BHK-21 hamster cells. Viral DNA replication commences about 3 h after infection of FHM cells with FV3. Between 3 and 6 h postinfection (p.i.), a portion of the intranuclear FV3 DNA is partly unmethylated. At later times, p.i., all of the viral DNA in the nuclear and cytoplasmic compartments is methylated at the 5'-CCGG-3' sequences. Cytoplasmic FV3 DNA has not been found unmethylated. We have cloned viral DNA fragments from methylated virion DNA. By using the genomic sequencing technique, it has been demonstrated for segments of the FV3 DNA replicated both in FHM fish and BHK21 hamster cells that in a stretch encompassing a total of 350 bp, all of the analyzed 5'-CG-3' dinucleotides are methylated. The modified nucleotide 5-methyldeoxycytidine is present exclusively in the 5'-CG-3' dinucleotide combination. In the cloned FV3 DNA fragment p21A, an open reading frame has been located. The 5' region of this presumptive viral gene is also methylated in all 5'-CG-3' positions. DNA methyltransferase activity has been detected in the nuclei of FV3-infected FHM cells at 4, 11, and 20 h p.i. In the cytoplasmic fraction, comparable activity has not been observed. These data are consistent with the interpretation that FV3 DNA is newly synthesized and de novo methylated in the nuclei of infected FHM cells and subsequently exported into the cytoplasm for viral assembly.

Amino Acid Sequence↗
PubMed1988

Proliferation and biosynthetic activities of myocytes from conductive system and working myocardium of the developing mouse heart. Light microscopic autoradiographic study.

Incorporation of 3H-thymidine, 3H-uridine and 3H-leucine into myocytes and their mitotic activity have been investigated in different compartments of the mouse heart conductive system (CS) and working myocardium (WM) at pre- and postnatal stages of development. On 15th and 18th d of embryogenesis, 3H-thymidine pulse and cumulative labelling indices (LI) of myocytes in the sinoatrial node (SAN), atrioventricular node (AVN) and His bundle (HB) were found to be 4 to 6 times lower than in WM (0.001 less than or equal to P less than or equal to 0.01). On 3rd to 9th d after birth LI remained decreased (0.001 less than or equal to P less than or equal to 0.05) in SAN while replicative activity of myocytes in AVN and HB approached that of both ventricular and atrial WM. Since 13th d after birth LI did not exceed 1% in all the compartments studied. In the prenatal period LI in SAN were higher than in AVN P less than or equal to 0.01), whereas in the postnatal one it was just the opposite (0.001 less than or equal to P less than or equal to 0.05). During cardiogenesis change of mitotic indices in WM and CS correlates with that of LI. Duration of cell cycle and its periods (G2, G2 + 1/2 M, S) estimated by curves of labeled mitoses, double-labelling technique with 14C-thymidine and 3H-thymidine and label "dilution" method differ slightly in myocytes from WM and CS. The latter incorporate 3H-uridine much less intensively at most stages of heart development compared with WM. At perinatal stages of cardiogenesis 3H-leucine labels CS myocytes somewhat weaker than WM ones until 8th d after birth when the labelling intensities in all cell types become similar. It can be concluded that on the whole CS myocytes from the developing mouse heart appear to be less active than those of WM concerning replication, transcription, and translation processes.

Aging↗
PubMed1999

Development of a general method of limited sampling for the determination of AUC for a drug that displays two-compartment pharmacokinetics.

OBJECTIVES: To develop a method of limited sampling that would enable accurate estimation of the area under the concentration time curve (AUC) when using the log trapezoidal method. METHODS: A series of datasets were simulated. Each dataset comprised 1000 subjects. Each subject was "administered" an intravenous bolus dose of a drug that displays two compartment pharmacokinetics. In the first series of simulations, a variety of combinations of the number of sampling times (K) and number of replicate measurements (R) at each of these times were tested, where K x R = 12 (i.e. N = 12). The times that each of the K samples were taken were chosen to be those that divided the AUC into K - 1 trapezoids of equal area. The concentration-time curves were estimated based on a priori estimates of the population parameters. The best combination of K and R was tested under various conditions of parameter variability and assay variability. The combinations were compared with a conventional sampling strategy, where N = 12, K = 12 (R = 1). RESULTS: The combination K = 4 and R = 3 proved to be the "best". It had similar accuracy to the conventional method. The best limited sampling combination was superior to the conventional method when assay variability was high (CV= 30%), was similar when assay variability was 15%, but the conventional method became statistically superior when assay variability was 7.5% or less. The accuracy of the best limited sampling combination was inversely related to the parameter variability. If K was set to 4 and R allowed to increase to 6 (i.e. N is not equal to 12), there was no further gain in accuracy. CONCLUSION: The proposed method of limited sampling is at least as accurate as the conventional intensive sampling technique, but more efficient in terms of sampling.

Area Under Curve↗
PubMed1992

Different pattern of activity of inhibitors of the human immunodeficiency virus in lymphocytes and monocyte/macrophages.

Monocyte/macrophages (M/M) are important targets for HIV in the body, and represent the majority of cells infected by the virus in some body compartments such as the central nervous system (CNS). M/M can be different from T-lymphocytes in terms of surface antigens, cell replication and drug metabolism. Thus, we evaluated, in M/M and in T-lymphocytes, the pattern of viral inhibition induced by various anti-HIV drugs, and assessed some of the mechanisms of action related to such antiviral activity. Inhibitors of HIV binding on CD4 receptors have similar activity in M/M and T-lymphocytes, while AZT and other dideoxynucleosides (ddN) are in general more active against HIV in M/M than in T-lymphocytes. This phenomenon can be related to the increased ratio in M/M of ddN-triphosphate/deoxynucleoside-triphosphate, and can at least in part explain the ability of zidovudine and didanosine in improving neurological dysfunctions in AIDS patients. Moreover, the antiviral activity of AZT (but not of other ddN- or HIV-binding inhibitors) is potently enhanced by cytokines like granulocyte-macrophage colony stimulating factor (GM-CSF) in M/M, while anti-HIV activity of TIBO compounds in M/M is not down-modulated by GM-CSF and other cytokines. Finally, non-toxic concentrations of adriamycin, an anticancer drug reported to be active against DNA viruses, can inhibit HIV replication in M/M (but not in T-lymphocytes). Taken together, these results suggest that M/M are selective targets for HIV with peculiarities different from those of T-lymphocytes. Thus, promising anti-HIV compounds should be evaluated both in T-cells and in M/M before reaching clinical trials. This may help in selecting drugs with good chances of being effective in patients with HIV-related disease.

Adenine↗
PubMed2005

Dynamic chromatin modifications characterise the first cell cycle in mouse embryos.

On fertilisation, gametes undergo epigenetic reorganisation and re-establish totipotency. Here, we investigate links between chromatin remodelling and asymmetric maintenance of DNA methylation in the early mouse embryo. Using antibodies for lysine specific H3 methylation reveals that the male pronucleus is negative for di- and trimethyl H3-K9 yet the female is positive for these residues. However, the male is positive for monomethyl H3-K9 and H3-K27 and these signals increase during pronuclear maturation. Non-histone chromatin proteins of the Polycomb group are found in the paternal compartment as early as sperm decondensation. However, trimethyl H3-K27 is not observed in the male until the completion of DNA replication. Heterochromatin protein 1 beta (HP1beta) is abundant in the male pronucleus, despite the absence of di- and trimethyl H3-K9, and co-localises with monomethyl H3-K9. Recent evidence identifies monomethyl H3-K9 as the preferred substrate of Suvar39h, the histone methyl transferase (HMT) responsible for heterochromatic H3-K9 trimethylation. The association of HP1beta with monomethyl H3-K9 may assist in preventing further modification of H3-K9. Association of dimethylation but not trimethylation of H3-K9 with DNA methylation, in the female pronucleus, suggests a mechanistically significant link. These differences begin to provide a chromatin based explanation for paternal-specific active DNA demethylation and maternal specific protection in the mouse.

Animals↗
PubMed2003

Development of bespoke bioluminescent reporters with the potential for in situ deployment within a phenolic-remediating wastewater treatment system.

A suite of ecologically relevant, site-specific bioreporters was constructed by transposon mutagenesis of microorganisms isolated from a polluted phenolic-remediating wastewater treatment system. Four Pseudomonad species were engineered to carry a stable chromosomal copy of the lux operon (luxCDABE) derived from Photorhabdus luminescens. These recombinant reporter microorganisms were tested for bioluminescence response to relevant phenol concentrations in the laboratory and to phenolic-containing effluents generated by an industrial wastewater treatment plant. The reporters displayed proportional responses of bioluminescence decay with increasing phenol concentrations up to 800 mg l(-1) of phenol. When deployed against samples from the treatment system, they showed superior operational range and sensing capabilities to that observed for industry standard microorganisms such as Vibrio fischeri. Specifically, the engineered strains accurately predicted toxicity shifts in all the treatment compartments under study (with phenolic concentrations ranging from approximately 10 to 600 mg l(-1)) with a low coefficient of variation of replicate determinations (between 1.16% and 8.32%). This work highlights the utility of genetic modification of native microorganisms from sites of interest to provide robust and ecologically relevant organism-based reagents for toxicity monitoring with the potential for in situ deployment.

Biodegradation, Environmental↗
PubMed2001

A photoregulated ligand for the nuclear import receptor karyopherin alpha.

The ability to orchestrate the transport of proteins between nucleus and cytoplasm provides cells with a powerful regulatory mechanism. Selective translocation between these compartments is often used to propagate cellular signals, and it is an intimate part of the processes that control cell division, viral replication, and other cellular events. Therefore, precise experimental control over protein localization, through the agency of light, would provide a powerful tool for the study and manipulation of these events. To this end, a prototype photoregulated nuclear localization signal (NLS) was derived from a native NLS. A library of 30 mutants of the bipartite NLS from Xenopus laevis nucleoplasmin containing a novel, photoisomerizable amino acid was prepared by parallel, solid-phase synthesis and screened in vitro for binding to the nuclear import receptor karyopherin alpha, which mediates the nuclear import of cellular proteins. A single peptide was identified in which the cis and trans photoisomers bind the receptor differentially. The strategy used to obtain this peptide is systematic and empirical; therefore, it is potentially applicable to any peptide-receptor system.

Amino Acid Sequence↗