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Biomedical subjects

A Burny

Publications and source records attributed to A Burny.

At least 55 records · Page 3Linked to original sources
PubMed1998

C-C chemokines, pivotal in protection against HIV type 1 infection.

Exposure to HIV type 1 (HIV-1) does not usually lead to infection. Although this could be because of insufficient virus titer, there is now abundant evidence that some individuals resist infection even when directly exposed to a high titer of HIV. This protection recently has been correlated with homozygous mutations of an HIV-1 coreceptor, namely CCR5, the receptor for the beta-chemokines. Moreover, earlier results already had shown that the same chemokines markedly suppress the nonsyncitial inducing variants of HIV-1, the chief virus type transmitted from person to person. CCR5 mutation, as a unique mechanism of protection, is, however, suspect because HIV-1 variants can use other chemokine receptors as their coreceptor. Moreover, recent results have established that infection can indeed sometimes occur with such mutations. Here, we report on transient natural resistance over time of most of 128 hemophiliacs who were inoculated repeatedly with HIV-1-contaminated Factor VIII concentrate from plasma during 1980-1985 before the development of the HIV blood test. Furthermore, and remarkably, 14 subjects remain uninfected to this date, and in these subjects we found homozygous CCR5 mutations in none but in most of them overproduction of beta chemokines. In vitro experiments confirmed the potent anti-HIV suppressive effect of these chemokines.

Cells, Cultured↗
PubMed1998

Apple messenger RNAs related to bacterial lignostilbene dioxygenase and plant SAUR genes are preferentially expressed in flowers.

In an attempt to use a differential display procedure to identify organ-specific genes in apple, cDNA fragments of two transcripts preferentially expressed in flowers were isolated and corresponding full-length cDNA inserts were subsequently obtained. One of these clones, Md-FS1, belongs to the SAUR gene family, originally identified as a set of auxin-inducible genes in soybean. The second one, Md-FS2, encodes a polypeptide with sequence similarities to bacterial lignostilbene-alpha,beta-dioxygenase isozymes, which are thought to be involved in lignin biodegradation. Northern blot analysis confirmed that both genes are preferentially expressed in floral organs at full bloom, while being expressed at lower or undetectable levels in vegetative organs (leaves, shoots or roots) as well as in immature, green and unopened blossoms. Furthermore, Md-FS1 transcripts also appeared to accumulate in vegetative tissues after auxin treatment of micropropagated apple shoots.

Amino Acid Sequence↗
PubMed1998

Evaluation of the genetic variability of 23 bovine microsatellite markers in four Belgian cattle breeds.

The polymorphism of 23 microsatellites in the four main cattle breeds in Belgium (Holstein Friesian, Belgian Blue, Belgian Red Pied and East Flemish) was analysed. Heterozygosity, polymorphism information content, the effective number of alleles, exclusion probability and the probability of genotypic identity for two random individuals were calculated for all microsatellites and all breeds. The Belgian Blue breed is generally a little less polymorphic in comparison with the other three breeds. Estimates of the genetic distances between these breeds confirmed the widely accepted proposition that the Belgian Blue is the most genetically distinct of these breeds. The three other breeds are likely to become one population, given current breeding strategies. Exclusion probabilities in parentage control cases are > 0.9999 in all four breeds when all 23 microsatellites are used and > 0.98 with only the two most polymorphic multiplexes.

Animals↗
PubMed1998

In vitro and in vivo oncogenic potential of bovine leukemia virus G4 protein.

In addition to the genes involved in the structure of the viral particle, the bovine leukemia virus (BLV) genome contains a region called X which contains at least four genes. Among them, the tax and rex genes, respectively, are involved in transcriptional and posttranscriptional regulation of viral transcription. Two other genes, R3 and G4, were identified after cloning of the corresponding mRNAs from BLV-infected lymphocytes. Although the function of the two latter genes is still unknown, they appear to have important roles, since deletion of them restricts viral propagation in vivo. In order to assess the oncogenic potential of the R3 and G4 proteins, we first analyzed their ability to immortalize and/or transform primary rat embryo fibroblasts (Refs). In this assay, the G4 but not the R3 protein cooperated with the Ha-ras oncogene to induce tumors in nude mice. It thus appears that G4 exhibited oncogenic potential in vitro. To extend these observations in vivo, the pathology induced by recombinant viruses with mutations in G4 and in R3 and G4 was next evaluated with the sheep animal model. Viral propagation, as measured by semiquantitative PCR, appeared to be reduced when the R3 and G4 genes were deleted. These observations confirm and extend our previous data underlining the biological function of these genes. In addition, we present the results of a clinical survey that involves 39 sheep infected with six different BLV recombinants. Over a period of 40 months, 83% of the sheep infected with a wild-type virus developed leukemias and/or lymphosarcomas. In contrast, none out of 13 sheep infected with viruses with mutations in G4 or in R3 and G4 developed disease. We conclude that in addition to its oncogenic potential in vitro, G4 is required for pathogenesis in vivo. These observations should help us gain insight into the process of leukemogenesis induced by the related human T-cell leukemia virus type 1.

Animals↗
PubMed1998

An interferon regulatory factor binding site in the U5 region of the bovine leukemia virus long terminal repeat stimulates Tax-independent gene expression.

Bovine leukemia virus (BLV) replication is controlled by both cis- and trans-acting elements. The virus-encoded transactivator, Tax, is necessary for efficient transcription from the BLV promoter, although it is not present during the early stages of infection. Therefore, sequences that control Tax-independent transcription must play an important role in the initiation of viral gene expression. This study demonstrates that the R-U5 sequence of BLV stimulates Tax-independent reporter gene expression directed by the BLV promoter. R-U5 was also stimulatory when inserted immediately downstream from the transcription initiation site of a heterologous promoter. Progressive deletion analysis of this region revealed that a 46-bp element corresponding to the 5' half of U5 is principally responsible for the stimulation. This element exhibited enhancer activity when inserted upstream or downstream from the herpes simplex virus thymidine kinase promoter. This enhancer contains a binding site for the interferon regulatory factors IRF-1 and IRF-2. A 3-bp mutation that destroys the IRF recognition site caused a twofold decrease in Tax-independent BLV long terminal repeat-driven gene expression. These observations suggest that the IRF binding site in the U5 region of BLV plays a role in the initiation of virus replication.

Animals↗
PubMed1998

Differential transcriptional activation by human T-cell leukemia virus type 1 Tax mutants is mediated by distinct interactions with CREB binding protein and p300.

The human T-cell leukemia virus type 1 Tax protein transforms human T lymphocytes, which can lead to the development of adult T-cell leukemia. Tax transformation is related to its ability to activate gene expression via the ATF/CREB and the NF-kappaB pathways. Transcriptional activation of these pathways is mediated by the actions of the related coactivators CREB binding protein (CBP) and p300. In this study, immunocytochemistry and confocal microscopy were used to localize CBP and p300 in cells expressing wild-type Tax or Tax mutants that are able to selectively activate gene expression from either the NF-kappaB or ATF/CREB pathway. Wild-type Tax colocalized with both CBP and p300 in nuclear bodies which also contained ATF-1 and the RelA subunit of NF-kappaB. However, a Tax mutant that selectively activates gene expression from only the ATF/CREB pathway colocalized with CBP but not p300, while a Tax mutant that selectively activates gene expression from only the NF-kappaB pathway colocalized with p300 but not CBP. In vitro and in vivo protein interaction studies indicated that the integrity of two independent domains of Tax delineated by these mutants was involved in the direct interaction of Tax with either CBP or p300. These studies are consistent with a model in which activation of either the NF-kappaB or the ATF/CREB pathway by specific Tax mutants is mediated by distinct interactions with related coactivator proteins.

Activating Transcription Factor 1↗
PubMed1997

Comparative processing of bovine leukemia virus envelope glycoprotein gp72 by subtilisin/kexin-like mammalian convertases.

Intracellular proteolytic processing of bovine leukemia virus (BLV) envelope glycoprotein precursor (gp72) at the C-terminal end of the RVRR268 / site is an essential step for virus infectivity. Subtilisin/kexin-like convertases cleave proproteins at preferred RX(K/R)R / sites, including those commonly found in viral envelope glycoprotein precursors. We first demonstrated that gp72 is processed into gp51/gp30 in both CV1 cells and the furin-deficient LoVo cells, leading us to compare the ability of mammalian convertases to cleave BLV gp72 in vitro. In contrast to the inability of the neuroendocrine PC1 to cleave gp72, the convertases furin, PACE4, PC5-A and PC5-B, which process constitutively secreted precursors, can effectively cleave gp72 into gp51/gp30. N-terminal sequence analysis of the convertase-generated gp30 demonstrated that cleavage occurs at the in vivo-utilized RVRR / SPV site. Such furin-, PACE4- and PC5-mediated processing was completely inhibited by the alpha1-antitrypsin variant alpha1-PDX. Mutagenesis of the gp72 cleavage site into RVRG-TPV resulted in complete abrogation of gp72 processing by endogenous CV-1 cells and by convertases in vitro. Since our in vitro data suggest a redundancy in the ability of the convertases to cleave gp72, RT-PCR analysis was used to define the convertases expressed in B-lymphocytes, representing one of the major targets of BLV infection. Our data revealed that only furin and the newly discovered PC7 mRNAs are expressed in Raji, B-Jab and LG2 cell lines.

Amino Acid Sequence↗
PubMed1997

Lack of mutation in the WAF1/CIP1 gene during bovine leukemia virus-induced leukemogenesis.

As a cyclin-dependent kinases (Cdks) inhibitor (CDI), the protein p21WAF1/CIP1 is able to interfere with cell cycle progression. Its expression is upregulated by wild-type p53, and the p21WAF1/CIP1 protein appears to be a potent effector of the p53-dependent cell cycle regulatory pathway. We have previously reported that p53 mutations frequently occur during bovine leukemia virus (BLV)-induced leukemogenesis in cattle but not in sheep. Therefore, we have investigated the involvement of p21WAF1/CIP1 mutations in the tumorigenic process associated with BLV. We first cloned the bovine and ovine WAF1 genes and determined the complete nucleotide sequences of their second coding exons. These sequences share respectively 79% and 80% homology with those of the human counterpart exon. In order to screen for mutations that could be associated with BLV-induced pathogenicity, we performed single strand conformation polymorphism (SSCP) assays on the WAF1 genes from BLV-induced tumors. No WAF1 mutations were detected in any of the ten BLV-induced bovine tumor samples. Among eleven sheep tumors and three ovine cell lines, only one sample revealed a single mutation in the WAF1 coding sequence, but this mutation was silent at the translational level. We concluded that mutations of the WAF1 gene are not involved in the development of the tumors during BLV-induced leukemogenesis.

Amino Acid Sequence↗
PubMed1997

Relationships of growth hormone gene and milk protein polymorphisms to milk production traits in Simmental cattle.

The importance of milk proteins and the positive effect of administration of growth hormone (GH) on milk production, and the presence in some dairy cattle lines of greater GH concentrations prompted us to examine the presence of restriction fragment length polymorphism at the GH gene using the restriction enzyme TaqI and to investigate associations between this polymorphism in Simmental cows and bulls, as well as milk protein variants in Simmental cows, and milk production traits. Blood and milk were sampled from 279 Italian Simmental cows and semen was collected from 148 bulls of the same breed. Two fragment bands, denoted A and B, of 6200 and 5200 bp respectively, were examined and three patterns, AA, AB and BB, were found in both animal samples. All variants previously reported in other studies, for kappa, beta, and alpha s1-caseins, and beta-lactoglobulin, were found in the cows' samples. For the cows' samples, a BLUP (Best Linear Unbiased Predictor) analysis of results was performed using a REML (Restricted Maximum Likelihood) program and known heritabilities, whereas for bulls we have performed a General Linear Model analysis. The effect of GH gene polymorphism, using TaqI restriction enzyme, on milk production traits was not significant, but bulls of BB pattern had a higher breeding value for milk yield than AA bulls (P < 0.05). For the kappa-casein genotypic effects, cows of AB genotype gave milk with 1.53 +/- 0.70 g/kg less fat than cows of AA genotype. In addition, breeding values for milk protein content were significantly higher in BB bulls, with 0.87 +/- 0.32 and 0.71 +/- 0.34 g/kg more milk protein than AA and AB bulls respectively. Thus, our results revealed a GH gene polymorphism and indicated significant effects of milk protein polymorphisms on milk production traits in the Italian Simmental breed.

Animals↗
PubMed1997

Knotted1-like homeobox genes are expressed during apple tree (Malus domestica [L.] Borkh) growth and development.

Apple (Malus domestica [L.] Borkh) cDNA clones encoding two distinct but very similar kn1-like homeobox gene class 1 homologues (KNAP1 and KNAP2) were isolated using a fragment amplified with degenerated primers as a probe. A fragment corresponding to the homeodomain region of KNAP1 was used to isolate a cDNA (KNAP3) belonging to the kn1-like homeobox gene class 2. These probes were used to detect corresponding gene copies in apple genomic DNA digests, together with other putative members of an apple kn1-like homeobox gene family. The kn1-like homeobox gene probes were also used to detect corresponding messenger accumulation levels in various organs of an apple tree. Transcripts corresponding to KNAP1 and KNAP2 genes appeared to be absent from leaves or floral organs, but they accumulate in tissue samples from elongated parts of the stem. In contrast, KNAP3 mRNAs accumulate at detectable levels in a wider range of both vegetative and reproductive organs.

Amino Acid Sequence↗
PubMed1997

Cloning of human IL-12 p40 and p35 DNA into the Semliki Forest virus vector: expression of IL-12 in human tumor cells.

IL-12 can enhance the development of effective immune responses against tumors as well as against certain infectious agents. It is therefore a potential candidate for therapeutic use in cancer therapy and in the design of vaccines against several infectious diseases. Several studies have demonstrated that IL-12 could efficiently induce tumor regression in animal models. To investigate the antitumor effect of direct gene transfer of human IL-12 into tumors, human IL-12 p35 and p40 cDNAs were cloned into the Semliki Forest virus (SFV) vector pSFV1. In order to express the two subunits from the same vector, the p35 and the p40 cDNAs were cloned into pSFV1, each under the control of a subgenomic SFV promoter. Recombinant RNA produced by in vitro transcription of SFV-IL-12 construct, was packaged into SFV viral particles with the use of a non-packageable helper RNA. We show that human tumor cell lines infected in vitro in vivo with recombinant SFV-IL-12 viral particles secrete high levels of biologically active heterodimeric p35/p40 IL-12, as demonstrated using ELISA and biological assays.

Gene Expression↗
PubMed1997

The major homology region of bovine leukaemia virus p24gag is required for virus infectivity in vivo.

In order to gain insight into the role of the major homology region (MHR) in the infectious potential of bovine leukaemia virus (BLV), mutations were introduced into the capsid gene of an infectious molecular clone. A provirus that was designed to contain only a slightly modified version of the MHR (substitution of phenylalanine 147 with a tyrosine) was still infectious in vivo. Furthermore, the provirus loads were not significantly different from those obtained with a wild-type virus. A second mutant was designed to analyse a mild modification of the MHR at the level of arginine 150. The substitution of this residue with a lysine completely destroyed the infectious potential of the recombinant virus. Finally, a third mutant that was deleted in the MHR region was unable to infect the host. Thus it appears that the integrity of the MHR domain is essential for BLV infectivity in vivo.

Amino Acid Sequence↗
PubMed1997

Both wild-type and strongly attenuated bovine leukemia viruses protect peripheral blood mononuclear cells from apoptosis.

Bovine leukemia virus (BLV) and the human T-cell leukemia viruses belong to the same subfamily of oncoviruses. Although much attention has focused on the mechanisms of cell proliferation and transformation by these viruses, experiments on the apoptotic process have yielded conflicting data in in vitro cell culture. Experimental infection of sheep with BLV proviruses offers the opportunity to analyze apoptosis in vivo. Here, we show that BLV-infected peripheral mononuclear cells, cultivated ex vivo, are protected from spontaneous programmed cell death. Moreover, the virus is able to specifically interfere with the apoptotic program of infected B lymphocytes. Strongly attenuated mutant proviruses that harbor deletions in the G4 and/or R3 genes also decrease the global susceptibility to apoptosis at levels similar to those obtained with the wild-type virus. In addition, cell culture supernatants from wild-type and mutant viruses can prevent uninfected cells from undergoing programmed cell death. These observations demonstrate that the R3 and G4 genes are not required to maintain both direct and indirect protection against apoptosis. They also imply that the level of programmed cell death observed ex vivo is independent of the amounts of proviruses in the animals. The failure of these cells to undergo apoptosis might be related to the pathogenesis induced by BLV.

Animals↗
PubMed1997

Fusion of bovine leukemia virus with target cells monitored by R18 fluorescence and PCR assays.

PCR and R18 fluorescence dequenching assays have been combined to monitor the kinetics of fusion of bovine leukemia virus with target cells (CC81, OVK, or Raji). Antibodies raised against gp51 allow us to demonstrate that not only the hydrophobic N-terminal domain of the transmembrane glycoprotein gp30 but also specific domains of gp51 (amino acids 39 to 103) are involved in bovine leukemia virus-cell fusion.

Animals↗
PubMed1997

The human T-cell leukemia virus type 1 transactivator protein Tax colocalizes in unique nuclear structures with NF-kappaB proteins.

The Tax protein of human T-cell leukemia virus type 1 (HTLV-1) is a potent activator of viral transcription. Tax also activates the expression of specific cellular genes involved in the control of T-lymphocyte growth via effects on cellular transcription factors, including members of the NF-kappaB/cRel family. Immunocytochemistry and electron microscopy were used to characterize the intracellular localization of Tax and identify cellular factors which are the potential targets for its transcriptional activity. These studies indicated that Tax localizes in discrete nuclear foci in T lymphocytes transformed by HTLV-1 and in cells transduced with Tax expression vectors. The Tax-containing foci are complex nuclear structures comprising a central core in which Tax colocalizes with splicing factor Sm. In addition to splicing factors Sm and SC-35, the Tax-containing nuclear structures also contain transcriptional components, including the largest subunit of RNA polymerase II and cyclin-dependent kinase CDK8. The inclusion of the two subunits of NF-kappaB, p50 and RelA, and the presence of the mRNA from a gene specifically activated by Tax through NF-kappaB binding sites suggest that these unique nuclear structures participate in Tax-mediated activation of gene expression via the NF-kappaB pathway.

Animals↗
PubMed1997

The cloning and sequencing of an ovine c-myc cDNA.

We report the cloning of an ovine c-myc cDNA. The clone was isolated from a bovine leukemia virus-infected cell line (YR2) cDNA library cloned in the lambda gt10 vector. The clone encodes the full length c-Myc protein made of 439 amino-acids with 93, 96, 92 and 93% similarity with human, feline, murine and rat c-Myc proteins, respectively.

Amino Acid Sequence↗
PubMed1997

Pit-1 gene polymorphism, milk yield, and conformation traits for Italian Holstein-Friesian bulls.

The growth hormone factor-1/pituitary-specific transcription factor Pit-1 is responsible for the expression of growth hormone in mammals. Mutations in Pit-1 have been found in growth hormone disorders of mice and humans. We studied the eventual association between Pit-1 polymorphism using the HinfI enzyme and the milk yield and conformation traits of 89 Italian Holstein-Friesian bulls. A strategy employing polymerase chain reaction was used to amplify a 451-bp fragment from semen DNA. Digestion of polymerase chain reaction products with HinfI revealed two alleles: allele A was not digested (451-bp fragment), and allele B was cut at one restriction site, generating two fragments of 244 and 207 bp. Three patterns were observed; frequencies were 2.2, 31.5, and 66.3% for AA, AB, and BB, respectively. Fixed and mixed linear models were fitted on daughter yield deviations for milk yields and on deregressed proofs for conformation traits. Predictions were weighted using the inverse of the estimated variance of records. The models used contained mean and gene substitution effects for Pit-1 A allele as fixed effects and random sire effect for the mixed model. The A allele was found to be superior for milk and protein yields, inferior for fat percentage, and superior for body depth, angularity, and rear leg set, which is difficult to explain. A canonical transformation revealed that Pit-1 had three actions, one linked to milk yield traits and angularity, a second linked to body depth and rear leg set, and a third linked to lower fat yields and to higher angularity.

Animals↗