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Topic:Molecular biology
Molecular biology in horses involves the study of molecular processes and genetic mechanisms that underpin equine physiology and health. This field encompasses the analysis of DNA, RNA, proteins, and other biomolecules to understand gene expression, genetic variation, and cellular functions in horses. Techniques such as genomic sequencing, gene expression profiling, and molecular diagnostics are employed to explore topics like hereditary diseases, performance traits, and immune responses in equines. This page assembles peer-reviewed research studies and scholarly articles that investigate the molecular biology of horses, focusing on genetic research, molecular techniques, and their applications in equine science.
Rotavirus infection in horses. Genome profile analysis of a rotavirus isolated from an infected foal. Electrophoretic analysis in polyacrylamide gel (PAGE) of the equine rotavirus 106/88/LI/EQ, isolated from the diarrhea of an 18 day old foal was compared to the bovine strain NCDV. There was a notable difference in the migration of some segments of the viral RNA. Bands 2 and 3 of the equine rotavirus comigrated while there was a clear separation of segments 7, 8 and 9. Moreover, the migration of segments 1, 4 and 5 revealed a lower molecular weight than the corresponding segments of NCDV.
Reduced endotoxin-induced production of tumor necrosis factor activity by equine peritoneal macrophages exposed to the dual inhibitor of arachidonic acid metabolism, SK & F 86002. The purpose of this study was to determine if a structurally novel dual inhibitor of arachidonic acid metabolism, SK & F 86002, would inhibit the endotoxin-induced production of tumor necrosis factor (TNF) activity by equine peritoneal macrophages. Equine peritoneal macrophages were variously pretreated for 0, 0.5 and 2 h with SK & F 86002 at 10(-9) to 10(-4) molar final concentrations or were left untreated. Then, the macrophages were cultured in vitro in the presence of endotoxin (5 ng/mL). Supernatant media were collected after 4 h and stored at -70 degrees C until assayed for TNF a...
Interconversion of E and S isoenzymes of horse liver alcohol dehydrogenase. Several residues contribute indirectly to catalysis. The E and S isoenzymes of horse liver alcohol dehydrogenase differ by 10 amino acid residues, but only the S isoenzyme is active on 3 beta-hydroxysteroids. This functional difference was correlated to the differences in structures of the isoenzymes by characterizing a series of chimeric enzymes, which could represent intermediates in the evolution of catalytic activity. Deletion of Asp-115 from the E isoenzyme created the E/D115 delta enzyme that is active on steroids. The deletion alters the substrate binding pocket by moving Leu-116, which sterically hinders binding of steroids in the E isoe...
The ‘natural’ hybrid haemoglobin from mule. Interrelationships with its parent haemoglobins from horse and donkey. The equilibrium O2-binding properties of the hybrid haemoglobin (Hb) present in vivo in erythrocytes from mule and of its parent Hbs from horse and donkey were compared with special reference to the effect of heterotropic ligands such as Cl-, D-glycerate 2,3-bisphosphate (DPG) and inositol hexakisphosphate. All these Hbs display a decreased effect by polyphosphates, confirming that what has been observed for horse Hb [Giardina, Brix, Clementi, Scatena, Nicoletti, Cicchetti, Argentin & Condò (1990) Biochem. J. 266, 897-900] is common to other equine species, at least from a qualitative sta...
Identification and partial purification of serum growth hormone binding protein in domestic animal species. The chemical nature and variations in serum concentrations of growth hormone binding protein (GHBP) from humans, rabbits, and rodents have been reported. To date little is known about the GHBP of domestic animals. Therefore, we initiated these studies to determine whether a serum GHBP was present in domestic animals and to purify the binding protein (BP) from serum of selected species. Using a dextran-coated charcoal separation assay, specific growth hormone (GH) binding was demonstrated in ovine, bovine, chicken, human, goose, porcine, and equine serum (listed in sequence from lowest to highe...
Characterization of a trypsin inhibitor from equine urine. A trypsin inhibitor was isolated from pregnant mares' urine by adsorption on bentonite and elution with aqueous pyridine followed by batch DEAE-cellulose treatment and column chromatography. Final purification to an electrophoretically homogenous glycoprotein was achieved by gel permeation chromatography. This equine urinary trypsin inhibitor (E-UTI) is acid- and heat-stable, has a molecular weight of 22 to 23 kDa, an isoelectric point of 4.55, forms a 1:1 molar complex with trypsin and has serine as its N-terminal amino acid. The N-terminal amino acid sequence of this protein is almost identi...
Isolation of a mucoid alginate-producing Pseudomonas aeruginosa strain from the equine guttural pouch. The isolation and characterization of a mucoid, alginate-producing strain of Pseudomonas aeruginosa from a nonhuman host, namely, in chondroids from an equine guttural pouch, is reported for the first time. Pure cultures of P. aeruginosa 12534 were isolated from a 17-month-old pony mare with a history of chronic bilateral mucopurulent nasal discharge from the right guttural pouch. Transmission electron microscopy of chondroids showed mucoid P. aeruginosa growing as microcolonies within a matrix of extracellular material. On the basis of expression of the mucoid phenotype under different growth...
Total synthesis of horse heart cytochrome C. A strategy based on complexation-assisted condensation of large synthetic protein fragments and mitochondria-mediated stereospecific heme insertion has been utilized to assemble a functional molecule corresponding to native horse heart holocytochrome c. This original approach offers the unique opportunity of selective modifications both in the C-terminal and in the N-terminal regions of the apoprotein and may represent an useful alternative to site-directed mutagenesis, particularly when D-amino acids, chemically and/or isotopically modified or other unnatural amino acids have to be introduced...
N-acetylation and C-terminal proteolysis of beta-endorphin in the anterior lobe of the horse pituitary. beta-Endorphin is post-translationally processed to both N-acetylated and C-terminally shortened derivatives in the anterior lobe of the horse pituitary, a processing pattern qualitatively different from that of the rat and virtually every other mammalian species. Thus, separation of the molecular forms of beta-endorphin using gel filtration and ion exchange chromatography showed that the horse anterior lobe primarily contains beta-endorphin-1-31 and N-acetyl-beta-endorphin-1-27 along with smaller amounts of beta-lipotropin, beta-endorphin-1-27, and N-acetyl-beta-endorphin-1-31 and -1-26, in c...
Analysis of immediate-early transcripts of equine cytomegalovirus. Equine cytomegalovirus (ECMV) contains a linear, double-stranded DNA genome composed of a 146-kbp unique region flanked by a pair of 18-kbp direct repeat (DR) sequences at the termini. Cycloheximide, actinomycin D, and phosphonoacetic acid were applied to infected cell cultures to divide viral transcription into immediate-early (IE), early, and late phases. Eight IE transcripts were identified and mapped to two regions (I and II) of the viral genome. Two of these IE RNAs (13.0 and 5.5 kb in size) were transcribed from region I, which is located within the DR regions; these IE genes are diploid...
Identification and characterization of the structural and nonstructural proteins of African horsesickness virus and determination of the genome coding assignments. Proteins present in purified African horsesickness virus (AHSV) and in infected cells were analyzed by SDS-polyacrylamide gel electrophoresis. Twelve viral proteins were identified, one minor and four major structural proteins, three major and two minor nonstructural proteins, as well as variable amounts of two additional structural proteins. Cell-free translation of total AHS virion RNA in a rabbit reticulocyte system resulted in the synthesis of proteins which were qualitatively and quantitatively similar to those found in infected cells. The in vivo and in vitro synthesized proteins were vi...
Characterization of the regulatory functions of the equine herpesvirus 1 immediate-early gene product. Use of the translation-inhibiting drug cycloheximide has indicated that the equine herpesvirus 1 (EHV-1) immediate-early (IE) gene, the sole EHV-1 IE gene, encodes a major viral regulatory protein since IE mRNA translation is a prerequisite for all further viral gene expression (W.L. Gray, R. P. Baumann, A. T. Robertson, G. B. Caughman, D. J. O'Callaghan, and J. Staczek, Virology 158:79-87, 1987). An EHV-1 IE gene expression vector (pSVIE) in combination with chimeric EHV-1 promoter-chloramphenicol acetyltransferase (CAT) reporter constructs was used in transient transfection assays to charact...
Methylprednisolone acetate induced release of cartilage proteoglycans: determination by high performance liquid chromatography. A high performance liquid chromatography (HPLC) procedure suitable for the simultaneous determination of the molecular size and concentration of macromolecular hyaluronate and proteoglycans in synovial fluid has been developed. Irrigation of the equine tarsocrural joint with 20 ml physiological saline (PSS) caused a mild inflammation with an increase of proteoglycans in the synovial fluid over the baseline arthrocentesis control sample. Proteoglycan and hyaluronate in the synovial fluid did not interact to form hyaluronate-proteoglycan aggregates, but separated as distinct chromatographic peak...
Sequence analysis of the equine H7 influenza virus haemagglutinin gene. The nucleotide sequences of ten haemagglutinin genes of representative H7N7 equine influenza viruses isolated between 1956 and 1977 have been determined by primer extension sequencing. Their nucleotide and deduced amino acid sequences demonstrate a high degree of homology. These equine viruses can be divided into two distinct subgroups, the prototype-like, and a group comprising the early American isolates and the remaining equine viruses. The equine H7 haemagglutinins form a quite distinct group compared to H7 haemagglutinins isolated from other species. Each of these equine H7 haemagglutinin...
Crystallographic studies of a calcium binding lysozyme from equine milk at 2.5 A resolution. The crystal structure of a calcium binding equine lysozyme has been determined at 2.5 A resolution by means of molecular replacement. The energy minimized equine lysozyme as the starting model, was refined with the molecular dynamics program, X-PLOR, and the R factor of the current model was found to be 24% without any water molecules. The conformation of the calcium binding loop is similar to that of alpha-lactalbumin. The profiles of backbone atomic displacements throughout the lysozyme and alpha-lactalbumin superfamilies are comparable as well as their homologous tertiary structures.
Putative fragile sites in the horse karyotype. After fluorouracil/5-bromodeoxyuridine synchronization and subsequent FPG-staining, the karyotype of 15 phenotypically normal horses displayed several breaks and gaps. Twelve bands 1q24, 4p12, 8q23, 11p12, 16q21, 17q21, 23q31, 23q32, Xp21, Xq22, Xq25 and Xq27 showed relatively frequent fragility. After thymidine/cytidine synchronization and subsequent GWL-banding the same horses display karyotypes without any fragility. Hence it is suggested that the above listed bands harbour folate and/or 5-bromodeoxyuridine sensitive fragile sites.
Diagnosis of equid herpesviruses -1 and -4 by polymerase chain reaction. The polymerase chain reaction (PCR) is a sensitive technique used to detect DNA of viral pathogens. We have applied the technique to the detection of Equid herpesviruses-1 and -4 (EHV-1 and EHV-4) DNA within nasopharyngeal swab samples from horses. Ninety-eight samples from suspected field cases and in-contact horses were analysed. The assays were conducted blind and later decoded and compared with virus isolation data. Our results indicate that PCR is a sensitive and rapid technique for the diagnosis of EHV-1 and EHV-4 infection.
Cloning of highly polymorphic microsatellites in the horse. We have isolated equine microsatellites by screening a genomic library with (TG)n and (TC)n probes. TG microsatellites were found to be more abundant than TC repeats, with an estimated frequency of one per 100,000bp. Sequence analysis of eight TG-positive clones revealed varying structures of the repeat regions; perfect stretches of TG repeats, imperfect stretches of TG repeats and compound regions of TG and TC repeats. Five loci were analysed by PCR and showed extensive polymorphism; three to seven alleles and heterozygosities of 0.40-0.76 were observed when screening 20-30 unrelated individu...
Identification of equine chromosomes in horse x mouse somatic cell hybrids. Giemsa-11 (G-11) staining and in situ hybridization were used to identify the equine chromosome complement of horse x mouse somatic cell hybrids. The presence of horse chromosomes in somatic cell hybrids was determined by differential G-11 staining. The slides were then destained and hybridized with biotinylated total horse (Equus caballus) genomic DNA without suppression. Fluorescence detection permitted rapid confirmation of horse chromosomal DNA in the hybrid cells.
Localization of the calcium release channel gene in cattle and horse by in situ hybridization: evidence of a conserved synteny with glucose phosphate isomerase. In situ hybridization techniques were used to localize regionally the calcium release channel (CRC) gene on cattle and horse chromosomes, using a porcine CRC cDNA probe. In cattle, the hybridization signal peaked on the 18q23-q26 bands and in horse on the 10pter region. Previous studies have shown that the glucose phosphate isomerase (GPI) gene localizes at the same site in both species, indicating that the two loci are syntenic. As CRC and GPI are syntenic in human, pig and mouse, the present results in cattle and horse represent another example of synteny conservation in the evolution of mam...
Stabilization of the structure of horse plasma vitamin D binding protein by disulfide bonds. Vitamin D binding protein (DBP) was isolated from horse plasma in a four-step procedure that involved Affi-Gel Blue affinity chromatography, gel filtration, hydroxylapatite chromatography, and anion exchange high-pressure liquid chromatography. The yield of DBP from 80 mL of plasma was 6-7 mg. Horse plasma DBP closely resembles other plasma DBPs, being a tryptophan-free protein of Mr 53,000. It is able to bind to and block the polymerization of monomeric actin. The secondary structure of DBP was calculated from circular dichroism measurements to be 39% alpha-helix, 42% beta-sheet, and 19% rand...
DNA fingerprinting in horses using a simple (TG)n probe and its application to population comparisons. A synthetic polynucleotide (TG)n was hybridized to equine DNA digested with HinfI and hypervariable hybridization patterns were obtained. Mendelian inheritance of these DNA fingerprinting patterns was confirmed by pedigree analysis. Estimates of the probabilities of identical band patterns in unrelated individuals of different breeds (Swedish Trotters, North Swedish Trotters, Thoroughbreds and Arabians) were in the range 1 x 10(-4) - 7 x 10(-6). The variability derived with the (TG)n probe in horses was higher than what we obtained with several other commonly used probes for DNA fingerprinting...
An inhibitor of tumor cell growth from normal horse serum. During our studies of cytostatic cytokines in the mixed leukocyte culture, we found that horse serum in the medium control contained a tumor cell growth-inhibitory factor. The fraction isolated by molecular sieving and ion exchange chromatography inhibited the growth and DNA synthesis of the primary culture and passaged cell line of the canine transmissible venereal sarcoma, murine T (L5178Y) and B (P3-X63-Ag8.653) lymphoid tumor cells, murine mammary tumor cells (RIII), bovine lymphoid tumor cells (BL3), and the nontransformed cell line of baby hamster kidney cells. Nontransformed cell lines ...
One-dimensional isoelectric focusing and immunoblotting of equine major histocompatibility complex class I antigens. The cells of 60 randomly selected Hannoveranian warm-blooded horses were subjected to one-dimensional isoelectric focusing and immunoblotting with a cross-reacting monoclonal antibody (Bo 1) recognizing bovine class I antigens. The banding patterns were correlated with the serologically defined specificities of the ELA-A locus. ELA-A2 was correlated with four bands, while ELA-A5, ELA-W18, ELA-A6, ELA-A14 and ELA-A9 were correlated with a single band each. The complexity of the pattern and additional polymorphic bands which could not be correlated to any of the known ELA specificities may indic...
Evolutionary pattern of the H 3 haemagglutinin of equine influenza viruses: multiple evolutionary lineages and frozen replication. The nucleotide and deduced amino acid sequences of the haemagglutinin genes coding for the HA 1 domain of H3N8 equine influenza viruses isolated over wide regions of the world were analyzed in detail to determine their evolutionary relationships. We have constructed a phylogenetic model tree by the neighbour-joining method using nucleotide sequences of 15 haemagglutinin genes, including those of five viruses determined in the present study. This gene tree revealed the existence of two major evolutionary pathways during a twenty five-year period between 1963 to 1988, and each pathway appeared t...
Linkage of hyperkalaemic periodic paralysis in quarter horses to the horse adult skeletal muscle sodium channel gene. A genetic disease observed in certain Quarter horses is hyperkalaemic periodic paralysis (HYPP). This disease causes attacks of paralysis which can be induced by ingestion of potassium. Recent studies have shown that HYPP in humans is due to single base changes within the adult skeletal muscle sodium channel gene. A large Quarter horse pedigree segregating dominant HYPP was studied to determine if mutations of the sodium channel gene are similarly responsible for HYPP in horses. We used cross-species, PCR-mediated, cDNA cloning and sequencing of the horse adult skeletal muscle sodium channel a...
Isolation, characterization, and quantitative analysis of ceruloplasmin from horses. Ceruloplasmin (Cp) was isolated from fresh equine plasma by precipitation, cellulose chromatography, and improved ion-exchange chromatography. Purified equine Cp is a glycoprotein having a molecular weight of approximately 115,000. In electrophoresis, equine Cp migrated to the alpha 1-globulin region, its isoelectric point was about 4.15 and consisted of about 890 amino acid residues. Serum Cp concentration was measured by use of the single radial immunodiffusion method. In clinically normal horses, the mean (+/- SD) serum Cp concentration of newborn foals was 2.87 +/- 0.40 mg/ml and that of 3...
A minimal lentivirus Tat. Transcriptional regulatory mechanisms found in lentiviruses employ RNA enhancer elements called trans-activation responsive (TAR) elements. These nascent RNA stem-loops are cis-acting targets of virally encoded Tat effectors. Interactions between Tat and TAR increase the processivity of transcription complexes and lead to efficient copying of viral genomes. To study essential elements of this trans activation, peptide motifs from Tats of two distantly related lentiviruses, equine infectious anemia virus (EIAV) and human immunodeficiency virus type 1 (HIV-1), were fused to the coat protein of b...
