MHC class I allele frequencies in pigtail macaques of diverse origin

Pigtail macaques (Macaca nemestrina) are an increasingly common primate model for the study of human AIDS. Major Histocompatibility complex (MHC) class I-restricted CD8⁺ T cell responses are a critical part of the adaptive immune response to HIV-1 in humans and simian immunodeficiency virus (SIV) in macaques; however, MHC class I alleles have not yet been comprehensively characterized in pigtail macaques. The frequencies of ten previously defined alleles (four Mane-A and six Mane-B) were investigated in detail in 109 pigtail macaques using reference strand-mediated conformational analysis (RSCA). The macaques were derived from three separate breeding colonies in the USA, Indonesia and Australia, and allele frequencies were analysed within and between these groups. Mane-A*10, an allele that restricts the immunodominant SIV Gag epitope KP9, was the most common allele, present in 32.1% of the animals overall, with similar frequencies across the three cohorts. Additionally, RSCA identified a new allele (Mane-A*17) common to three Indonesian pigtail macaques responding to the same Gag CD8⁺ T cell epitope. This broad characterization of common MHC class I alleles in more than 100 pigtail macaques further develops this animal model for the study of virus-specific CD8⁺ T cell responses.     

Genetic variation in the carbonic anhydrase isozymes of macaque monkeys. II. Inheritance of red cell carbonic anhydrase levels in different carbonic anhydrase I genotypes of the pig-tailed macaque,Macaca nemestrina

The inheritance of red blood cell levels of carbonic anhydrase isozymes (CA I and CA II) has been studied in different carbonic anhydrase I genotypes of the pig-tailed macaque, Macaca nemestrina. Quantitation of CA I isozymes in a series of animals indicates that the total CA I concentration is the sum of the average effects of each CA I structural allele and that the average effects are independent of the various allelic combinations. The relative average effects were 0.32:0.95:1.0 for the CA I ^a, CA I^b, and CA I ^c structural genes, respectively. It is also demonstrated that the level of CA II is related to the CA I genotypes. Multiple regression analysis demonstrated that each dose of CA I-deficiency gene present decreased the CA II concentration by approximately 30%, with this decrease in CA II level being solely related to the dose of CA I-deficiency gene and not to the level of CA I. The CA I-deficient animals produce CA I products that are similar to the common CA Ia, CA Ib, CA Ic electrophoretic types. Limited mating data indicate that the CA I components in CA I-deficient animals are inherited codominantly.Supported by U.S. Public Health Service Research Grant GM-15419.This report is a portion of a dissertation submitted to the University of Michigan in partial fulfillment of the requirements for the Doctor of Philosophy degree.U.S. Public Health Service Predoctoral Trainee (GM-71-14).     

Latitudinal,taxonomic, sexual,and insular determinants of size variation in pigtail macaques,Macaca nemestrina

Regression analysis demonstrates that body and skull size are correlated with latitude in Sundaic (Macaca nemestrina nemestrina)and Indochinese (M. n. leonina)pigtail macaques. These two contiguously distributed subspecies are unusual in that they exhibit opposite latitudinal gradients of increasing size— M. n. nemestrinabecomes larger in a southerly direction,while M. n. leoninabecomes larger in a northerly direction in accordance with the usual pattern of ecogeographic variation seen in other sympatric macaques and mammals (Bergmann ’s rule). The difference in clinal size gradients is one more of a series of characters which delineate these two macaques as natural biological populations. However, since the size gradients converge on a narrow zone along the Thai-Malay Peninsula, the use of size as a character to evaluate genetic and morphological intergradation is equivocal when population variation is considered. The third subspecies, the Mentawi Island pigtail (M. n. pagensis),is an endemic,insular isolate differentiated from the Sundaic pigtail from similar latitudes in terms of its small size. Thus, latitude, insularity, and taxonomic differentiation all affect size variation in the pigtail macaques;also,although the data are not definitive, there is the suggestion that the degree of sexual dimorphism may be an additional covariate of latitude and/or body size.     

Studies of esterase 6 in Drosophila melanogaster. I. The genetics of a posttranslational modification

A locus has been found, an allele of which causes a modification of some allozymes of the enzyme esterase 6 in Drosophila melanogaster. There are two alleles of this locus, one of which is dominant to the other and results in increased electrophoretic mobility of affected allozymes. The locus responsible has been mapped to 3-56.7 on the standard genetic map (Est-6 is at 3-36.8). Of 13 other enzyme systems analyzed, only leucine aminopeptidase is affected by the modifier locus. Neuraminidase incubations of homogenates altered the electrophoretic mobility of esterase 6 allozymes, but the mobility differences found are not large enough to conclude that esterase 6 is sialylated.This work was supported by NIH Grant No. GM23706 and PHS Grant SO7RR7031 to Rollin C. Richmond and by NIH Genetics Training Grant No. 82 to Indiana University.     

Human-type ABO,MN, and Lewis blood groups,and Gm and Inv factors in several species of macaques

More than 1,000 blood samples were collected from macaques of speciesM. fuscata, M. cyclopis, M. irus, M. mulatta, M. nemestrina, andM. speciosa, and all or a part of them were tested for human-type ABO, MN, and Lewis blood groups, and Gm and Inv factors. Differences between and/or within species analogous to racial differences in man were markedly noted in the distribution of the ABO and Lewis blood groups. Saliva samples from a small number ofM. fuscata were tested quantitatively for the presence of H and Lewis substances, and it was found that almost all the animals were secretors of H, Le^a, and Le^b, independently of the Lewis blood groups of their red cells. Red cells of all macaques tested contained M or M-like, but not N^v(V), antigens, and no polymorphism of MN blood groups was present. Selected plasma samples fromM. fuscata, M. cyclopis, M. irus, andM. nemestrina were found to be negative for all Gm(1), Gm(2), Gm(4), and Inv(1) factors tested.This study was supported in part by the Japan Society for Promotion of Science Grant B-54 and by National Science Foundation Grant FJ 4.11. 1 as part of the Japan-U.S. Cooperative Science Program.     

Assignment of six enzyme loci to multipoint linkage groups in Fishes of the genusPoeciliopsis (Poeciliidae): Designation of linkage groups III–V

Three new linkage groups of enzyme loci are described usingPoeciliopsis monacha × P. viriosa-derived interspecific backcross hybrids. Comparison to known linkage groups of the confamilial genusXiphophorus shows homology betweenXiphophorus linkage group I andPoeciliopsis linkage group III,Xiphophorus linkage group II andPoeciliopsis linkage group I, andXiphophorus linkage group IV andPoeciliopsis linkage group IV. Comparison of the gene content of other fish, amphibians, and mammal syntenic groups suggests retention of plesiomorphic vertebrate gene arrangements in at least two poeciliid linkage groups. Expansion of thePoeciliopsis gene map should be of utility in the identification of tumor regulatory genes through demonstration of linkage to biochemical markers.This work was supported by NSF Grants BSR 19355 and BSR 16569 and NIH Grants CA 44303 and CA 39729 to R. S. Nairn and D.C.M. and NIEHS Grant EHS 1P50ES 0384801A1 to R.J.S.     

Regulation of amylase activity in Drosophila melanogaster: Variation in the number of enzyme molecules produced by different amylase genotypes

Purified amylases from high- and low-activity variants of Drosophila melanogaster showed identical specific activities. Immunoelectrophoresis of crude larval homogenates showed severalfold differences between strains in the amounts of cross-reacting material. Control of amylase activity is trans-acting in heterozygotes between high- and low-activity variants. These results suggest the existence of polymorphic regulatory genes affecting the production levels of amylase protein in D. melanogaster.This work was supported by Grant GM-21279 from the Institute of General Medical Science of the NIH to R. C. Lewontin and by an Operating Grant from the Natural Sciences and Engineering Research Council Canada to D. A. Hickey.     

Chromobacterium violaceum infections in 13 non-human primates

Background Recently, an Indian‐origin macaque was found dead and Chromobacterium violaceum was isolated from the skin wound, and hepatic and pulmonary abscesses. Methods By searching the database, a total of thirteen cases of C. violaceum infection in pigtail macaques (n = 8), rhesus macaques (n = 4), and one baboon were identified from 2001 to 2010 at Tulane National Primate Research Center. Medical records were reviewed for breed, sex, age, clinical findings, treatment, outcome, bacteriology, and gross and histological findings. Results Seven pigtail macaques and one Indian‐origin rhesus macaque died of chromobacterial septicemia. All chromobacterial septicemic pigtail macaques were adult with higher incidence in female. Hepatic abscess and thrombosis were typical findings along with pulmonary abscess and thrombosis, renal venous thromboembolism, and necrosuppurative pleuritis, peritonitis, splenitis, myocarditis, pericarditis, and meningoencephalitis. Skin wound, uterine infection, and oral and respiratory exposure were considered the points of entry for these animals. Conclusions This represents the first report of chromobacteriosis in pigtail, rhesus macaque, and baboon. Our experience suggests that chromobacterial infections may be more common in non‐human primates than previously recognized.     

Ultrastructure of the corpus cardiacum and corpus allatum of the house cricket Acheta domesticus

Summary The ultrastructure of the corpus cardiacum (CC) and corpus allatum (CA) of the house cricket, Acheta domesticus, is described. Axon profiles within the CC contain neurosecretory granules 160–350 nm in diameter which are indistinguishable from those found in type I neurosecretory cells of the pars intercerebralis and in the nervus corporis cardiaci I. The CC itself contains two cell types: intrinsic neurosecretory cells and glial cells. Intrinsic NSC cytoplasm contains Golgi bodies and electron dense neurosecretory granules 160–350 nm in diameter. Synaptoid configurations with 20–50 nm diameter electron lucent vesicles were observed within axon profiles of the CC. The structure of the CA is relatively uniform with one cell type predominating. Typical CA cells possess large nucleoli, active Golgi complexes, numerous mitochondria, and occassional microtubules. Groups of dark staining cells scattered throughout the CA of some animals were interpreted as evidence of cellular death.This work was done while JTB was supported by USPHS Training Grant HD-0266 from NICHDI wish to express my thanks to Dr. Richard A. Cloney for sharing his expertise in electron microscopy     

Biochemical genetics of Fundulus heteroclitus (L.). IV. Spatial variation in gene frequencies of Idh-A,Idh-B, 6-Pgdh-A,and Est-S

The spatial variation in gene frequencies of four unlinked polymorphic loci was studied in the teleost Fundulus heteroclitus. Three loci (Idh-A, Idh-B, and Est-S) exhibit significant north-south clinal variation in allelic frequencies along the Atlantic Coast of North America, while a fourth locus (6-Pgdh-A) shows a modest clinal variation. These data, together with our previous data for Ldh-B, Mdh-A, Gpi-B, and Pgm-A, reveal a pattern of low gene diversity in the colder northern extremes of the species range and high gene diversity in warmer southern latitudes.This work was supported by Grants DEB76-19877 and DEB79-12216 from the National Science Foundation and by Grant P60-80-04 from the State of Maryland. REC and RVB were supported by NIH Training Grant GM07231 to the Department of Biology.Contribution No. 1104 from the Department of Biology, The Johns Hopkins University.     

Identification of MHC class I sequences in Chinese-origin rhesus macaques

The rhesus macaque (Macaca mulatta) is an excellent model for human disease and vaccine research. Two populations exhibiting distinctive morphological and physiological characteristics, Indian- and Chinese-origin rhesus macaques, are commonly used in research. Genetic analysis has focused on the Indian macaque population, but the accessibility of these animals for research is limited. Due to their greater availability, Chinese rhesus macaques are now being used more frequently, particularly in vaccine and biodefense studies, although relatively little is known about their immunogenetics. In this study, we discovered major histocompatibility complex (MHC) class I cDNAs in 12 Chinese rhesus macaques and detected 41 distinct Mamu-A and Mamu-B sequences. Twenty-seven of these class I cDNAs were novel, while six and eight of these sequences were previously reported in Chinese and Indian rhesus macaques, respectively. We then performed microsatellite analysis on DNA from these 12 animals, as well as an additional 18 animals, and developed sequence specific primer PCR (PCR-SSP) assays for eight cDNAs found in multiple animals. We also examined our cohort for potential admixture of Chinese and Indian origin animals using a recently developed panel of single nucleotide polymorphisms (SNPs). The discovery of 27 novel MHC class I sequences in this analysis underscores the genetic diversity of Chinese rhesus macaques and contributes reagents that will be valuable for studying cellular immunology in this population.     

Linkage of the locus for conversion of albumin (Acf-1)in the house mouse,Mus musculus

The linkage of the locus for conversion of albumin (Acf-1) has been established on chromosome 1 with the following gene order and recombination percentages: Id-1 19.3±5.2% Acf-1 4.2±1.7% Dip-1 18.4±4.2% Lp.This work was supported by NIH Postdoctoral Fellowship 1F32 GM0527701, Grant BMS75-03397 from the National Science Foundation, Grant ACS VC-17-R from the American Cancer Society, and Contract NO1-ES42159 from the National Institute of Environmental Health Sciences. The Jackson Laboratory is fully accredited by the American Association for the Accreditation of Laboratory Animal Care.     

Biochemical characterization of “LAP,” a polymorphic aminopeptidase from the blue mussel,Mytilus edulis

A genetically variable naphthylamidase enzyme, previously described as leucine aminopeptidase, was purified approximately fiftyfold, and its biochemical properties were investigated. The enzyme was renamed aminopeptidase I. Substrate affinities demonstrate that it is an -aminoacyl peptide hydrolase (E.C. 3.4.11.-). Aminopeptidase I had a monomer molecular weight of 65–68,000, average pI of pH 4.88, and broad pH optima between 6.5 and 8.0. The enzyme was inactivated rapidly between 40 and 50 C. Antibodies from purified enzyme did not cross-react with other naphthylamidases, but aminopeptidase I activity was inhibited by immune serum. The enzyme exhibited highest naphthylamidase activity for aromatic and hydrophobic aminoacyl naphthylamides. Aminopeptidase activity was highest for aromatic and hydrophobic N-terminal residues of tripeptides. Certain divalent metal cations, p-O H-mercuribenzoate, and N-ethylmaleimide were strongly inhibitory while chelating agents activated the enzyme.This work was supported by NIH Grant GM-21133, NSF Grant DEB 77-06074, USPHS Career Award GM-28963 to R. K. Koehn, and NSF Grant PCM 7513461A-01 to N. Arnheim.     

Biochemical genetics of Fundulus heteroclitus (L.). III. Inheritance of isocitrate dehydrogenase (Idh-A and Idh-B), 6-phosphogluconate dehydrogenase (6-Pgdh-A), and serum esterase (Est-S) polymorphisms

Starch gel electrophoresis has shown that natural populations of Fundulus heteroclitus have variants at four enzyme-coding loci: Idh-A, Idh-B, 6-Pgdh-A, and Est-S. Analysis of the phenotypic distribution of the F₁ generation suggests that each of the variants segregates as autosomally inherited codominant alleles. Tissue specificity and intracellular localization were also determined for the IDH and 6PGDH isozymes.This work was supported by Grants DEB 76-19877 and DEB 79-12216 from the National Science Foundation and by Grant P60-80-04 from the State of Maryland. RVB and REC were supported by NIH Training Grant GM07231 to the Department of Biology.Contribution No. 1103 from the Department of Biology, The Johns Hopkins University.     

The effect of insertion of the maize transposable elementMutator is dependent on genetic background

A secondary mutant, derived from an allele of maize alcohol dehydrogenase 1 (Adh1) carrying a Mutator transposable element (Mu1) in its first intron, was reported to exhibit a threefold decrease in ADH enzymatic activity and steady-state RNA levels compared to the original mutant. The original mutant,Adh1-S3034 (abbreviatedS3034), was previously characterized at the molecular level. The derivative, abbreviatedS3034b, has now been cloned; at the DNA sequence level the insertion and surroundingAdh1 sequences are indistinguishable fromS3034. Furthermore, in our lines there is no difference in relative ADH activities between products of the two putative alleles. A comparison of gene expression in heterozygotes obtained by crossing to different tester lines reveals a correlation between the measured decrease in levels of ADH polypeptide produced by the mutant allele and the background in which it is measured; this effect is distinct from any background-related variation in the expression of the progenitor allele. It does not appear to be attributable to alternative patterns of DNA modification. It appears to reflect a background-associated difference in the level of normalAdh1-RNA produced. Thus the previously reported distinction betweenS3034 andS3034b may be due to differences in the extent to which the mutant allele and a given genetic background interact to produce functionalAdh1-RNA.This research was supported by United States Public Health Service Grant GM38616 and United States Department of Agriculture Grant 87-CRCR-1-2500 to J.S. D.O. was supported by an NIH predoctoral training grant to the Department of Genetics.     

Characterization of 47 MHC class I sequences in Filipino cynomolgus macaques

Cynomolgus macaques (Macaca fascicularis) provide increasingly common models for infectious disease research. Several geographically distinct populations of these macaques from Southeast Asia and the Indian Ocean island of Mauritius are available for pathogenesis studies. Though host genetics may profoundly impact results of such studies, similarities and differences between populations are often overlooked. In this study we identified 47 full-length MHC class I nucleotide sequences in 16 cynomolgus macaques of Filipino origin. The majority of MHC class I sequences characterized (39 of 47) were unique to this regional population. However, we discovered eight sequences with perfect identity and six sequences with close similarity to previously defined MHC class I sequences from other macaque populations. We identified two ancestral MHC haplotypes that appear to be shared between Filipino and Mauritian cynomolgus macaques, notably a Mafa-B haplotype that has previously been shown to protect Mauritian cynomolgus macaques against challenge with a simian/human immunodeficiency virus, SHIV_89.6P. We also identified a Filipino cynomolgus macaque MHC class I sequence for which the predicted protein sequence differs from Mamu-B*17 by a single amino acid. This is important because Mamu-B*17 is strongly associated with protection against simian immunodeficiency virus (SIV) challenge in Indian rhesus macaques. These findings have implications for the evolutionary history of Filipino cynomolgus macaques as well as for the use of this model in SIV/SHIV research protocols. Kevin J. Campbell and Ann M. Detmer contributed equally to this work.     

Naturally occurring quantitative variants of acid phosphatase-1 in Drosophila melanogaster

We have examined 111 wild Drosophila melanogaster lines for cis-acting quantitative variants of the Acph-1 gene, which codes for acid phosphatase-1 (ACPH). Three variants with obvious, reproducible phenotypes were isolated. All variants acted equally on all tissues and developmental stages examined. No recombinants were detected between one quantitative variant and the site determining the electrophoretic mobility of Acph-1 among 3885 flies examined. Several enzymatic properties of the variant enzymes were tested, including the K _m values for two substrates, inhibition by three different inhibitors, and thermal stability; the variant enzymes behaved identically to the wild-type enzyme in all cases. Immunological titration experiments showed that the variant enzymes had the same enzyme activity per molecule of ACPH as the wild-type enzyme. These results suggest that the quantitative variants we have identified are altered in the regulatory portion of Acph-1 so as to produce altered numbers of normal ACPH molecules.This work was supported by NIH Grant 21548. MAJ was supported by NIH Predoctoral Training Grant GM07413.     

Alcohol-oxidizing enzymes in 13 Drosophila species

Starch and polyacrylamide gel electrophoresis were used to ascertain the substrate specificities of alcohol-oxidizing enzymes in 13 Drosophila species. The substrates used were a variety of long- and short-chain aliphatic alcohols, one aromatic alcohol, and benzaldehyde. Only one enzyme (product of a single-gene locus) showed significant NAD⁺-dependent alcohol dehydrogenase activity with short-chain aliphatic alcohols. The 13 species, belonging to four different Drosophila groups, all showed a similar complement of alcohol-oxidizing enzymes, although differences in electrophoretic mobility and in levels of activity existed from species to species. These findings are relevant to the adaptation of Drosophila to alcohol environments.This study was supported by NIH Grant 1 PO1 GM 22221 and by Contract PA 200-14 Mod #4 with ERDA.     

Macromolecular interaction and the electrophoretic mobility of esterase-5 from Drosophila pseudoobscura

Esterase-5 is one of the most polymorphic loci in Drosophila pseudoobscura. Some variants reportedly produce a dimeric enzyme, while a few produce a monomeric form. This paper reports the finding that during electrophoresis ESTERASE-5 exists in a dynamic equilibrium between monomers and dimers, an equilibrium that is dependent on the running temperature of the gels. This is shown by a series of analytical electrophoresis experiments in which the apparent molecular weights of several variants are determined at four different temperatures. Increasing temperatures result in a linear decrease in the logarithm of apparent molecular weights. Macromolecular interactions thus are a significant determinant of EST-5 electrophoretic mobility.G.K.C. was supported by NIH Grant Gm 21179 to R. C. Lewontin. E.Á. was supported on an Icelandic Science Foundation Grant (Vísindasjóur Íslands), a Fulbright-Hayes Grant, a NATO-Science Fellowship, and a Fogarty International Research Fellowship of the Fogarty International Center (NIH F05 TWO 3027-01).     

Effect of a single dose of inducers and inhibitors on the rate of synthesis of cytochromes and reductases in liver organelles

Summary Measurement of the effect of drugs on the in vivo rates of synthesis of rabbit liver organelle bound proteins were measured following individual treatments with the inducers phenobarbital, 3-methylcholanthrene and PCB (a mixture of polychlorinated biphenyls) and the inhibitors, cycloheximide, aflatoxin 13, chloramphenicol and actinomycin D. Following their isolation from a homogenate containing the combined livers of ¹⁴C-leucine injected experimental animals and 3H-leucine injected control animals, purified fractions of the following proteins were prepared: microsomal cytochrome b₅, cytochrome P-450, NADH-cytochrome b₅ reductase, NADPH-cytochrome P-450 reductase and proteolipids, outer mitochrondrial membrane cytochrome b5, NADH-cytochrome b₅ reductase and proteolipids, inner mitochrondrial membrane cytochrome c, NADH dehydrogenase and proteolipids, intermitochrondrial membrane cytochrome b₅ and circulating serum albumin.This research was supported by grants from the PSC-BHE Research Award Program of The City University of New York, US Public Health Service Grant SO 7 PRO7132-07 and the Alma Toorock Fund for Cancer Research.