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PubMed1993

Polymorphism in BoLA-DRB3 exon 2 correlates with resistance to persistent lymphocytosis caused by bovine leukemia virus.

The relationship between polymorphism of the bovine lymphocyte Ag (BoLA)-DRB3 gene and resistance and susceptibility to persistent lymphocytosis (PL) caused by bovine leukemia virus (BLV) was investigated. Exon 2 of the BoLA-DRB3 gene was cloned from animals with BoLA haplotypes previously found to be associated with resistance and susceptibility to PL. Sequence analysis revealed the presence of the amino acids Glu-Arg (ER) at putative Ag binding residues 70 and 71 only in BoLA haplotypes associated with resistance to PL. This correlation was confirmed in a case control study (n = 26) using an allele-specific polymerase chain reaction for the detection of ER at residues 70-71. These results provide a molecular basis for Ir gene control of resistance and susceptibility to PL and suggest that the cellular immune response is important in preventing the in vivo spread of BLV infection.

Alleles↗
PubMed1994

Evidence for the expression of three different BoLA-class II molecules on the bovine BL-3 cell line: determination of a non-DR non-DQ gene product.

Studies were conducted to determine the reactivity of six monoclonal antibodies specific for major histocompatibility complex class II molecules expressed on a bovine B cell line homozygous for BoLA-DR and BoLA-DQ alleles. Direct immunoprecipitation, serial immunodepletion experiments, and two-dimensional gel electrophoresis revealed that these antibodies reacted with three different molecules. Bovine orthologues of HLA-DR were recognized by three monoclonal antibodies--H34A, TH12A, and TH14B. Orthologues of HLA-DQ were characterized by two other monoclonal antibodies, TH22A and TH81A. A third BoLA class II antigen, neither DR nor DQ was revealed by the last monoclonal antibody H42A. The relation of this molecule to known molecules in humans and other species remains to be established. However, cumulative data suggest that the determinant is expressed on a molecule related to HLA-DP.

Alleles↗
PubMed1994

Both DQB genes are expressed in BoLA haplotypes carrying a duplicated DQ region.

The objective of this study was to determine whether more than one DQB gene is expressed in three BoLA haplotypes that have a duplicated DQ region. Leukocyte mRNA from three animals genotyped at the BoLA-A, DQB, and DRB3 loci was used as template for reverse transcription-polymerase chain reaction (RT-PCR), cloning, and DNA sequencing. Five DQB alleles were identified. All cDNA clones were 564 base pairs (bp) in length, including 507 bp of nonprimer-derived sequence that contained the coding sequence for the full length of exon 2 and 74 amino acids of exon 3. Three alleles were assigned to the DQB1 locus and two were assigned to DQB2 on the basis of sequence comparisons with previously reported alleles. The expression of DQB1 and DQB2 in individual animals was examined by RT-PCR followed by double digestion of the 564 bp PCR products with Eco O109 I and Dra III in order to discriminate the DQB alleles. Evidence that DQB1 and DQB2 are both transcribed was obtained for three different BoLA haplotypes with DQB duplications, DQB10, DQB11C, and DQB12. These results suggest that the duplication or deletion event that gave rise to the DQB1 and DQB2 genes is a relatively recent event in the evolution of the cattle MHC.

Alleles↗
PubMed2003

T cell epitope identification for bovine vaccines: an epitope mapping method for BoLA A-11.

T cell responses play an important role in immunity to parasites and other microbial agents of infectious diseases, therefore a number of T cell-directed vaccines are in development. Computer-driven algorithms that facilitate the discovery of T cell epitopes from protein and genome sequences are now being used to accelerate preclinical studies of human vaccines. Similar tools are not yet available for predicting T cell epitopes for animal vaccines, but there may be sufficient data available to begin the process of compiling the algorithms. We describe the construction of a novel mathematical 'matrix' that describes the properties of bovine major histocompatibility complex (BoLA) system antigen (BoLA) A-11 peptide ligands, developed for use with EpiMatrix, an existing T cell epitope-mapping algorithm. An alternative means of developing BoLA matrices, using the pocket profile method, is also discussed. Matrices such as the one described here may be used to develop T cell epitope-mapping tools for cattle and other ruminants. Epitope-mapping algorithms offer a significant advantage over other methods of epitope selection, such as the screening of synthetic overlapping peptides, because high throughput screening can be performed in silico, followed by ex vivo confirmatory studies. Furthermore, using epitope-mapping algorithms, putative T cell epitopes can be derived directly from genomic sequences, allowing researchers to circumvent labor-intensive cloning steps in the genome-to-vaccine discovery pathway.

Algorithms↗
PubMed2005

Self-assembly of hollow cones in a bola-amphiphile/hexadiamine salt solution.

Cones are a very rare form of supramolecular self-assembly. It is shown that mixing of an unsymmetrical bola-fatty acid and a diamine (catanionic system) in dilute solutions can produce aggregates having a conical shape. These assemblies are made of frozen unsymmetrical monolayers in which bolas are arranged on a regular hexagonal lattice. Cones are stabilized by the introduction of pentagonal or n-gonal defects in the bola-lattice which yields the quantification of the cone apex angle according to the Euler theorems. The formation of cones represents a novel route for lipid and surfactant systems to minimize the edge energy of a flat membrane.

Diamines↗
PubMed2003

Ion channels from linear and branched bola-amphiphiles.

The synthesis and characterization of the ion channel activity of three new bola-amphiphiles is described. These compounds are conceptually derived from a previously reported bis-cyclophane bola-amphiphile through opening of the cyclophanes to acyclic structures and were found to readily form ion channels in planar bilayer membranes as assessed by bilayer clamp single-channel analysis. All three compounds behaved very similarly: the dominant channels formed by all three are Ohmic with specific conductance of 10 +/- 1 pS (NaCl electrolyte) and 39 +/- 1 pS (CsCl electrolyte). Single-ion permeability ratios, determined from dissymmetric electrolyte experiments, showed the selectivity P(Cs(+)) > P(Na(+)) > P(Cl(-)). Less frequently, lower conductance channels were also observed to act independently of the dominant channels. The lifetimes of the dominant channels range from 70 to 280 ms for the three compounds with some very long-lived openings (20-40 s) observed for two of the three. The lower conductance states have shorter lifetimes. This study demonstrates that bis-macrocyclic compounds are not essential for channel formation by bola-amphiphiles, and opens a new class of channel-forming compounds for structure-activity optimization.

Algorithms↗
PubMed2004

Sex pheromone components of the bronzed cutworm, Nephelodes minians, a prey species of a bolas spider, Mastophora hutchinsoni.

The bolas spider, Mastophora hutchinsoni, attracts adult males of four species of moths by aggressively mimicking their sex pheromone. Here, we report the identification of two sex pheromone components of one of these species, the bronzed cutworm, Nephelodes minians. Simultaneous gas chromatographic (flame ionization detection) and electroantennographic detection (EAD) analysis of extracts of the sex pheromone glands of female N. minians indicated two components eliciting strong EAD responses from a conspecific male antenna that corresponded in retention time with (Z)-11-hexadecenal (Z11-16:Ald) and (Z)-11-hexadecenyl acetate (Z11-16:OAc), using both polar and nonpolar columns. Mass spectra of these two peaks were identical to those of synthetic Z11-16:Ald and Z11-16:OAc. The double-bond positions were confirmed by GC-MS analysis of dimethyl disulfide derivatives of the putative pheromone components. In a field test, a 5:1 blend of Z11-16:Ald to Z11-16:OAc attracted male N. minians. Overall, there is no overlap in the chemical constituents of the sex pheromones of three moth species preyed upon by this bolas spider for which pheromones have now been identified. These studies suggest that bolas spiders are versatile in their ability to synthesize semiochemicals for different prey species. This versatility may be advantageous to a predator that has an otherwise extremely specialized hunting tactic.

Acetates↗
PubMed2003

Characterization of naturally processed and presented peptides associated with bovine major histocompatibility complex (BoLA) class II DR molecules.

Differential regulation of genetic resistance to infectious disease may partially be explained by variation in the binding affinity and the repertoire of pathogen-derived antigenic peptides associated with major histocompatibility complex (MHC) molecules. In this study, we investigated characteristics of peptides that bind to the bovine MHC allele BoLA-DRB3*2703, which is associated with occurrence of clinical mastitis in Holstein dairy cattle, and assigned a putative peptide-binding motif to this allele. This was achieved by in vitro expression of allele *2703 as well as a control allele, BoLA-DRB3*1201 which is present at high frequency in Holsteins. Transfected cell lines alone (for allele *1201) or in combination with blood mononuclear cells from an animal homozygous for allele *2703 were used as the source of naturally processed and presented peptides. Subsequent to elution of peptides from BoLA-DR+ cells, their sequences were determined by electrospray ionization mass spectrometry. Eluted peptides were between 13 and 20 amino acids long and the majority were in sets of overlapping sequences. These peptides were derived from intra- and extracellular proteins, as well as foreign proteins present in the culture medium. Some peptides had originated from molecular chaperones present in the endoplasmic reticulum, such as ER-60 and GRP78, pointing to some degree of overlap and cross-sampling between MHC class I and class II antigen presentation pathways. Consistent with reports of human and mouse MHC class II-associated peptides, putative peptide-binding motifs could be assigned to alleles *2703 and *1201, comprising a hydrophobic or an aromatic residue at relative position 1, a hydrophobic residue at position 4 and a small residue at position 6 of the eluted peptides. These findings provide the foundation for future studies of molecular mechanisms of MHC-disease associations of cattle.

Amino Acid Sequence↗
PubMed1979

Analysis of alloantisera against bovine lymphocytes. Joint report of the 1st International Bovine Lymphocyte Antigen (BoLA) workshop.

The results and agreements of the 1 international BoLA workshop, held in Edinburgh, Scotland in August 1978, are reported. Most of these concern the results from a comparison test of 249 alloantisera to bovine lymphocytes, the antisera being contributed by 9 laboratories. These sera were compared directly in Edinburgh on a panel of lymphocytes from 130 cattle of 21 breeds. In the microlymphocytotoxicity test used 75% of the sera reacted. Sixty eight of these sera were grouped into clusters according to their reaction patterns against the lymphocyte panel. Eleven of these clusters were clearly defined and were given workshop BoLA designations. In addition 22 sera were assigned to subgroups of the agreed clusters. There was no evidence that the method of production of the sera had any effect on their specificity. Although genetic data was not available, the phenotypes of the test panel of lymphocytes are consistent with the clusters detecting antigens controlled by multiple alleles at a single autosomal locus. It was agreed to name the genetic region where this putative locus is located BoLA (bovine lymphocyte antigen).

Animals↗
PubMed1987

Serological relationships among antigens of the BoLA and the bovine M blood group systems.

Alloimmunizations with either lymphocytes or red cells from donor cows positive for BoLA w16 and blood group M' antigens into recipients negative for these antigens produced antisera reactive in the cytotoxic test with w16-positive lymphocytes and in the haemolytic test with M'-positive erythrocytes. Similarly, alloimmunizations of blood group M1-negative recipients with either lymphocytes or red cells from donor cows possessing the M1 blood group factor produced antisera specifically reactive with lymphocytes and erythrocytes from M1-positive cattle. Absorptions with either lymphocytes or erythrocytes from individual animals of the same M antigenic type as the donor removed all haemolytic and cytotoxic reactivity. The results indicate that blood group M' and BoLA w16 share a similar antigenic structure. Likewise, blood group M1 has an antigenically similar counterpart which is also part of the BoLA system.

Animals↗
PubMed1988

Relationship between the bovine major histocompatibility complex (BoLA), erythrocyte markers and susceptibility to mastitis in Icelandic cattle.

Milk and blood samples were obtained from three Icelandic dairy herds. The herds were monitored regularly for mastitis incidence. Cell counts, adenosine triphosphate (ATP) and antitrypsin levels of the milk samples were recorded. In addition, red cell and BoLA typing were performed on the blood. Although cell counts and ATP levels showed significant associations with mastitis, antitrypsin levels did not. Red blood cell antigens N'2 and S1 and the lymphocyte antigen detected by the monoclonal antibody M7 were associated with low cell counts, whilst BoLA w6 and w6.1 were associated with high cell counts. BoLA w6.2 and w11 showed significant association with high antitrypsin levels. Only ED116 showed a significant association with mastitis.

Animals↗
PubMed1988

Immunomagnetic isolation of cells for serological BoLA typing.

This paper describes a totally new immunomagnetic (IM) technique adapted to serological BoLA typing. The basic technique has recently been developed by Vartdal et al. (1986) for serological HLA typing. The main advantage is that bovine mononuclear cells (e.g. T-cells and possibly their subsets, B-cells and monocytes) can be quickly and specifically isolated with high yield and viability from whole blood in a one-step procedure. This is achieved by magnetic separation of rosettes formed between the cells and superparamagnetic monosized polystyrene microspheres (Dynabeads TM) coated with cross-species reactive monoclonal antibodies (MAbs) specific for various human T-cell antigens or for HLA class II monomorphic epitopes. The cells are isolated within 5 min after a 5-min incubation at 4 degrees C. Magnetic separation of rosettes with a strong cobalt-samarium magnet eliminates all the laborious centrifugation steps necessary with conventional procedures. The isolated cells, still attached to the particles, are available for microcytotoxic assay. This is carried out within 55 min, including a two-step application of alloantiserum and complement and addition of acridine orange/ethidium bromide for the staining of viable (green) and dead (red) cells. The high viability of isolated cells gives a very low background kill compared with the conventional cytotoxic assay. The IM typing technique is also superior in sensitivity to the conventional technique as standardized for the international BoLA comparison test. The IM technique is likely to have its greatest impact on class II typing; class II positive cells being separated very efficiently. Polymorphic HLA class II MAbs detected likely polymorphic BoLA class II epitopes.

Animals↗
PubMed1991

Production of alloantisera against class II bovine lymphocyte antigens (BoLA) by cross-immunization between class I matched cattle.

This paper describes the production of alloantisera directed against bovine major histocompatibility complex (MHC) (BoLA) class II antigens in animals whose MHC phenotypes had been defined by one dimensional isoelectric focusing. Animals of closely matched BoLA class I types were selected by serology and subsequently typed for class I and class II by 1D-IEF of immunoprecipitated antigens. Those with similar class I type by both methods, but differing at the class II locus, were chosen for reciprocal immunization. Cross-immunization was by two skin implantations 6 weeks apart. The resulting antisera showed low titre after the first immunization and elevated titre 3 weeks after the second immunization. The sera reacted strongly with cells expressing specific BoLA class II antigens. The pattern of reactivity correlated well with IEF class II typing on a panel of animals representing all of the class II IEF types present in the Friesian population.

Animals↗
PubMed1996

Detection of a common BoLA-DRB3 deletion by sequence-specific oligonucleotide typing.

The bovine MHC clas II allele BoLA-DRB3*2A has an amino acid deletion of unknown function at codon 65 in the second exon, which codes for the antigen-binding site. Sequence-specific oligonucleotides were designed based on published nucleotide sequences on BoLA-DRRB3 alleles, and used to detect this deletion in 51 Hereford cattle. Probes 65+ and 65- detect the presence or absence of codon 65 respectively. Oligonucleotide probes were labelled with Digoxigenin (DIG), hybridized to dot blots of BoLA-DRB3 exon 2 polymerase chain reaction (PCR) product, and detected by chemiluminescence. Of the 51 animals screened, two were homozygous and 11 were heterozygous for the deletion at codon 65. The methodology described here provides the necessary tools to screen rapidly for this deletion in a large number of animals in order to study its effect on antigen binding and immune response.

Alleles↗
PubMed1980

Bovine lymphocyte antigens (BoLA): a serologic, genetic and histocompatibility analysis.

Eleven lymphocyte antigens have been defined in cattle using 81 lymphocytotoxic sera. These sera (typing reagents) were selected from over 1,000 normal sera, 35 alloimmune sera, and 111 samples of colostrum whey. Absorptions revealed that nine of the 11 antigens detected by these reagents are serologically independent. Segregation of the genes controlling these antigens was observed among 470 calves from mating of 56 heterozygous sires and 360 dams, indicating that each of the 11 antigens is controlled by one of 11 alleles at a single autosomal co-dominant locus BoLA-A (Bovine Lymphocyte Antigen, locus A). To determine the relation between BoLA antigens and histocompatibility, skin allografts were exchanged among four unrelated adult cows. Incompatibilities for the SD antigens, but not MLR or blood group antigens, were predictive of early rejection of allografts. Furthermore, graft recipients produced antibodies against the SD antigens of incompatible donors. These results indicate that the BoLA-A locus is part of the major histocompatibility complex (MHC) of cattle.

Absorption↗
PubMed1981

Analysis of BoLA w6. Evidence for multiple subgroups.

BoLA w6 is one of 16 specificities agreed upon at the 2nd International BoLA Workshop and many laboratories have produced sera reacting with this specificity. This paper presents evidence for at least four sub-groups in w6. Three Edinburgh sera showing identical reaction patterns in the 1st and 2nd BoLA workshops have been studied by absorption. One serum contained a single antibody population reacting with an epitope common to all w6 positive cells. The other two contained antibodies against the same common epitope, antibodies to epitopes on four sub-groups and an antibody reacting only with one subgroup w6.1. Another Edinburgh serum contained two populations of antibodies. One reacting with the common epitope and one with an epitope on two or possibly three subgroups. Immunisation between w6 positive individuals produced antisera to two subgroups without producing antibodies to common epitopes. At least two additional subgroups are likely to exist. These results indicate the presence of specificities unique to individual subtypes coupled with epitopes common to some and all w6 subgroups.

Animals↗
PubMed1983

Monoclonal antibodies to HLA recognize monomorphic and polymorphic epitopes on BoLA.

Fourteen monoclonal antibodies recognizing monomorphic and polymorphic epitopes on class I and class II antigens of the human MHC have been assayed on lymphocytes of a panel of 20-150 BoLA typed bovine animals from 12 different breeds. Some monomorphic antibodies cross-reacted and others did not. Two polymorphic monoclonal antibodies in man recognize a polymorphism in cows that follows allospecificities (BoLA-w3, w9) already described. Immunoprecipitation experiments with monomorphic anti-B2m and anti-HLA-DR monoclonal antibodies have shown that these cross-reactions concern BoLA antigens. They also revealed that Ia-like antigens in cattle present the same two chain features characterized in other species.

Animals↗
PubMed1988

Identification, cloning, and expression of bolA, an ftsZ-dependent morphogene of Escherichia coli.

A newly found morphogene of Escherichia coli, bolA, mapping at min 10 of the genetic map, was cloned in a 7.2-kilobase BamHI fragment and identified by its ability to produce osmotically stable spherical cells when overexpressed. This gene codes for a polypeptide of 13 kilodaltons. Overexpression of bolA+ was achieved in low-copy-number vectors with operon fusions to the tet and lac promoters, indicating a clockwise direction of transcription. While no modification of any of the penicillin-binding proteins was observed, morphological effects due to overexpression of bolA+ were shown to be dependent on the presence of an active ftsZ gene product. Our results suggest the existence of a mechanism mediated by FtsZ for modifying the conformation of nascent murein in the early steps of septum formation.

Cell Division↗