The expression of antibody diversity in natural and laboratory-made polyploid individuals of the clawed toad Xenopus

Antibody diversity, as measured by isoelectric focusing of dinitrophenol-specific antibodies, was compared in different polyploid species of the clawed toad Xenopus. Antibody heterogeneity increased with chromosome number and DNA content from Xenopus tropicalis (2n=20 chromosomes) to Xenopus ruwenzoriensis (2n=108 chromosomes). Laboratory allopolyploids made by hybridization between two species showing different antibody diversities and different chromosome numbers gave antibody patterns intermediate between the two parents. On the other hand, autopolyploid individuals showed no increase in antibody diversity, showing that increased polyploidy alone cannot be responsible for increased heterogeneity. In contrast to the increase in antibody diversity following polyploidization, the number of expressed major histocompatibility complex alleles, as measured by a mixed lymphocyte reaction, did not increase. This locus appeared to be diploid or in the process of rediploidization in all the Xenopus species studied. Selection has thus operated differentially on the polyploid immunoglobulin and major histocompatibility loci. It apparently preserved the additional heterogeneity acquired for immunoglobulins favoring the expression of an expanded antibody repertoire in polyploid species.     

Histocompatibility antigens and immunoglobulin genes in the clawed toad: Espression and linkage studies in recombinant and hyperdiploidxenopus hybrids

Hybrids betweenXenopus laevis andXenopus gilli can express species specific antigens recognizable by immunological techniques. The expression of various markers such as immunoglobulins, cell surface antigens, and major histocompatibility complex antigens, has been studied in three different types of models that help in linkage study and in gene mapping: 1) recombinant diploid gynogenetic animals, 2) recombinant diploid backcross progeny, and 3) a panel of hyperdiploid animals having supernumerarygilli chromosomes.The results are as follows: The major histocompatibility complex and immunoglobulin heavy chain genes are not closely linked. Somegilli red blood cells antigen, detected by aX. laevis antiserum, andgilli heavy chain genes belong to different linkage groups. The major histocompatibility complex is far from the centromere, as evidenced by the high frequency of crossing over detected at this locus in the gynogenetic progeny of a hybrid female. A negative correlation was found between the expression of onegilli red cell antigen and the expression of particulargilli anti-DNP antibodies. The above mentionedgilli antigen seems to depend on chromosom 9. Chromosome 12 can be definitely excluded from participation in heavy chain synthesis. Chromosomes 8 and 9 are unlikely to have a direct role in heavy chain synthesis.     

Identification of class I major histocompatibility complex encoded molecules in the amphibian Xenopus

Class I-like molecules have been immunoprecipitated from Xenopus leukocytes and erythrocytes with alloantisera directed against major histocompatibility complex (MHC)-linked antigens. The heavy chains, depending on the allele examined, have molecular weights of 40 000–44 000 of which 3000 daltons are asparagine-linked carbohydrates, probably present as one N-linked glycan. The presumed analogue of ₂-microglobulin has a molecular weight of 13 000 and bears no asparagine-linked glycans. Family studies show that the heavy chains are encoded by genes residing in or closely linked to the MHC.Abbreviations used in this paper MHC major histocompatibility complex - CML cell-mediated lympholysis - MLR mixed leukocyte reaction - APBS amphibian phosphate-buffered saline - kd kilodalton - LG Xenopus laevis — Xenopus gilli species hybrids - IEF isoelectric focusing Founded and supported by F. Hoffmann-La Roche & Co., Limited Company, CH-4005 Basel, Switzerland.     

Genomic in situ hybridization inArachis (Fabaceae) identifies the diploid wild progenitors of cultivated (A. hypogaea) and related wild (A. monticola) peanut species

Genomic in situ hybridization offers a powerful tool for investigating genome organisation and evolution of taxa known, or suspected, to be allopolyploids. The question of the diploid progenitors of cultivated peanut (Arachis hypogaea, 2n=4x=40) has been the subject of numerous studies at cytogenetical, cytochemical, biochemical and molecular levels, but no definitive conclusions have been reached. The biotinylated total genomic DNA from potential diploidArachis species were separately hybridized in situ to root tip chromosomes ofA. hypogaea and wild speciesA. monticola (2n=4x=40) without or mixed with an excess of unlabelled DNA from the species not used as a probe. Among the range of different species combinations used, the strong and uniform signals given by labelledA. ipaensis DNA when hybridized toA. hypogaea andA. monticola in combination with unlabelledA. villosa DNA indicates that overall molecular composition of twenty chromosomes ofA. hypogaea andA. monticola is very similar toA. ipaensis chromosomes. ProbingA. hypogaea andA. monticola chromosomes with labelled genomic DNA fromA. villosa mixed with unlabelled DNA fromA. ipaensis likewise labelled strongly and uniformly the other twenty chromosomes. BarringA. ipaensis, all the diploidArachis species presently investigated had characteristic centromeric bands in the twenty chromosomes within the complement indicating a clear division ofA. ipaensis from other species. InA. hypogaea andA. monticola only twenty chromosomes showed centromeric bands. These results (i) confirm the allopolyploid nature ofA. hypogaea andA. monticola, (ii) strongly support the view that wildA. monticola and cultivatedA. hypogaea are very closely related, and (iii) indicate thatA. villosa andA. ipaensis are the diploid wild progenitors of the tetraploid species studied. The present results also reveal that the nucleolus organizing region (NOR) originating fromA. villosa alone is expressed in the two tetraploid species.     

EVOLUTIONARY INFERENCES FROM RESTRICTION MAPS OF MITOCHONDRIAL DNA FROM NINE TAXA OF XENOPUS FROGS

Restriction endonuclease cleavage maps were prepared by the double digestion method for mitochondrial DNAs (mtDNAs) purified from Xenopus borealis, X. clivii, X. fraseri, X. muelleri, X. ruwenzoriensis, X. vestitus, X. laevis victorianus, X. l. laevis, and a variant of X. laevis designated X. laevis “davis.” An average of 21 cleavage sites per genome were mapped with 11 restriction endonucleases. Among the four invariant sites found are three conserved not only among the Xenopus mtDNAs tested but also among nearly all vertebrate mtDNAs examined to date. Two of these are Sac II sites in the 12S and 16S ribosomal RNA genes, and one is a Hpa I site in the gene for asparagine transfer RNA. These three sites permit the alignment and comparison of mtDNAs from different vertebrate classes. Although most of the differences observed among the Xenopus maps are attributable to point mutations causing gain or loss of restriction sites, the maps also differ by three large length mutations in or near the displacement loop. Phylogenetic analysis of 30 informative sites suggests that those members of the laevis species-group that have 36 chromosomes per somatic cell can be divided into three subgroups: 1) X. borealis, X. clivii, and perhaps X. fraseri (the “borealis” subgroup), 2) X. muelleri, and 3) the subspecies of X, laevis. The mtDNA of the hexaploid (2n = 108) species, X. ruwenzoriensis, is most similar to that of taxa in the latter two subgroups, which contrasts with the morphological similarity of this species to X. fraseri. X. ruwenzoriensis may be an allopolyploid with a mother (the contributor of the cytoplasmic mtDNA genome) on the X. laevis or X. muelleri lineage and a father on the X. fraseri lineage. We present a model showing how mtDNA and nuclear genomes can yield contrasting phytogenies for species-groups that have undergone several rounds of interspecific hybridization. Comparison of mitochondrial and nuclear sequence divergences suggests that Xenopus mtDNA, like that of mammals and birds, evolves faster than nuclear DNA. Genetic distances among mtDNAs of Xenopus species are very large, generally approaching or exceeding one substitution per nucleotide.     

Discovery of a diploid population of theHemidactylus garnotii-vietnamensis complex (Reptilia: Gekkonidae)

A diploid member of the parthenogenetic gekkonid species complexHemidactylus garnotii-vietnamensis was discovered for the first time from Thailand. This gecko, seemingly unisexual and parthenogenetic, possesses 2n=2x=38 chromosomes, showing distinct heteromorphisms. The absence of bisexual congeneric species with a combination of karyomorphs to produce this karyotype indicates the occurrence of chromosomal rearrangements after the initial estabilishment of a diploid clonal lineage of hybrid origin. Results of karyotypic comparisons of the present sample and the three known triploid species belonging to theH. garnotii-vietnamensis complex suggest that a triploid karyomorph similar to that ofH. vietnamensis has first emerged through an insemination of the diploid parthenogen's egg by the sperm from a bisexual species having 44 chromosomes (all telocentric), and that the karyomorph subsequently experienced some minor chromosomal aberrations to produce the karyomorphs ofH. vietnamensis andH. garnotii. The origin of theH. stejnegeri karyotype still remains an open question for future studies.     

Globin evolution in the genusXenopus: Comparative analysis of cDNAs coding for adult globin polypeptides ofXenopus borealis andXenopus tropicalis

Summary Globin mRNAs ofXenopus borealis andXenopus tropicalis have been cloned and sequenced. The nucleotide and derived amino acid sequences were compared with each other and with already available data fromXenopus laevis. This analysis rendered clear evidence that the common ancestor ofX. laevis andX. borealis, but not ofX. tropicalis, had lost one amino acid of the -globins prior to a genome duplication event that preceded the segregation of the former two species. Replacement-site substitutions were used to calculate a rough time scale of genome duplication and species segregation. The results suggest an ancient separation between theX. laevis and theX. tropicalis groups occurring approximately 110–120 million years ago. Analysis of the amino acid chains demonstrated various alterations. However, some functional domains, like heme-binding sites and₁₂ contact sites, were subject to a high degree of conservation, indicating the existence of functional constraints on them also in the genusXenopus.     

A major histocompatibility complex in the toadxenopus laevis (Daudin)

The genetic relationship between mixed leukocyte reaction (MLR), skin graft rejection, and some red blood cell antigens has been studied in a sibship of the toadXenopus laevis. MLR typing was achieved using blood lymphocytes. The graft experiments were performed at 17–19°C. Grafts exchanged between MLR identical sibs were rejected in 30.9±5.1 days, grafts exchanged between MLR different sibs were rejected in 20.4±2.4 days when animals differed at one MLR haplotype, and in 18.6±1.9 days when they differed at two MLR haplotypes. Immunizations and absorptions following the MLR typing produced agglutinating antisera that recognize red blood cell antigens segregating with MLR haplotypes. The results parallel those obtained in various mammalian and avian species and suggest that the homology of the major histocompatibility complex (MHC), described in higher vertebrates, can be extended to amphibians.     

Taxonomic studies ofAsarum sensu lato

The karyotype and the C-banding pattern in two species ofHexastylis andAsarum epigynum were analysed in detail, and the results obtained were compared with those of the other species ofAsarum, Asiasarum andHeterotropa previously reported. The present results were partially different from the previous reports related to the karyotypes of these species. The karyotype observed in two species ofHexastylis (2n=26) was represented by ten pairs of metacentric chromosomes and three pairs of small subtelocentric chromosomes, which is very similar to that ofAsiasarum in eastern Asia. The C-banding patterns ofHexastylis andAsiasarum, however, were clearly different from each other. A striking difference was found in one of the three pairs of small subtelocentric chromosomes. A Formosan speciesAsarum epigynum had the somatic chromosome number 2n=12 and a highly asymmetrical karyotype composed of mainly subtelocentric chromosomes. These karyological features were remarkably different from those of the other groups inAsarum s.l.     

The effect of thymectomy on the mixed leukocyte reaction and phytohemagglutinin responsiveness in the clawed toadXenopus laevis

In the clawed toadXenopus laevis, thymectomy at 7 days of age abrogates the phytohemagglutinin response of leukocytes and the mixed leukocyte reaction.A project grant from the Medical Research Council to Dr. J.D. Horton is gratefully acknowledged.     

Karyotype analysis and heterochromatin differentiation with Giemsa C-banding and fluorescent counterstaining inCephalanthera (Orchidaceae)

Detailed studies of the chromosomes of the three Austrian species of the genusCephalanthera showed them all to have basically similar karyotypes. BothC. damasonium (2n = 36) andC. longifolia (2n = 32) have three large and several classes of smaller chromosome pairs. The karyotype ofC. rubra (2n = 44) is composed of four large and several groups of smaller pairs. The heterochromatin in these species amounts to about 10% of total karyotype length. All the chromosomes have Giemsa-positive centromeres, but only a few have intercalary or terminal bands. Using differential fluorescent staining with DAPI/actinomycin D, quinacrine/actinomycin D (both A-T specific), and chromomycin A₃/distamycin A (G-C specific) three different types of major heterochromatic bands can be characterized in respect of their satellite DNA composition: highly A-T rich, slightly A-T rich, and very G-C rich. The chromosomes ofC. longifolia contain more A-T rich C-bands than those ofC. damasonium, while the latter's have more G-C rich heterochromatin. In both species several C-bands appear as secondary constrictions or gaps in the Feulgen-stained chromosomes, but most likely, in each species there is only one pair of chromosomes where the secondary constrictions function as nucleolus organizing regions. No major intraspecific variation could be observed except on one small chromosome pair ofC. longifolia which had a heteromorphic C-band in most individuals. Possible pathways of karyotype evolution involving polyploidy and Robertsonian events are discussed.     

Duplication of the MHC-linked Xenopus complement factor B gene

We have previously reported the molecular cloning of the mammalian major histocompatibility complex (MHC) class III gene, complement factor B (Bf) from Xenopus laevis, and linkage of the gene to the frog MHC. Here, we estimated the copy number of the Xenopus Bf gene by genomic Southern blotting analysis and demonstrated that Xenopus laevis has two copies of the Bf gene. Both genes co-segregated with the MHC-linked HSP70 genes among 19 offspring of an f/r × f/r cross, indicating a close linkage of the two Bf genes to the frog MHC. Both genes are transcribed and contain open reading frames. When compared with the previously determined cDNA sequence (Xenopus Bf A), the predicted amino acid sequence of the second cDNA species (Xenopus Bf B) shows 82% overall identity. Polymerase chain reaction analysis indicated that all of the partially inbred frogs with the f, r, g, and j MHC haplotypes, as well as 12 outbred frogs tested have both Bf genes, suggesting that the duplicated Bf genes are stable genetic traits in Xenopus laevis.     

Notes on chromosomes of ten species of the genus chrysolina mots. (Coleoptera:Chrysomelidae)

SixChrysolina species from Catalonia and the Canary Islands (Spain), viz.americana, gemina, femoralis, cerealis, menthastri andpolita, have similar diploid karyotypes of 24 (sub)metacentric chromosomes, and show Xy sex-determining system.C. banksi andobsoleta have 2n ()=23; their karyotype is presumably derived from that of the former group by loss of the y chromosomes. InC. haemoptera andC. carnifex 40 elements appear in the diploid set. It seems that 2n=24 is the most frequent number in theChrysolina. Higher chromosome numbers have possibly originated through centric fissions, as the acrocentric shape ofC. carnifex chromosomes seems to suggest. The 2n=23 and 24 species feed onLabiatae, while the two higher chromosome number species are associated with plants belonging to other families.     

A three-locus model for the chicken major histocompatibility complex

The major histocompatibility complexes (B complexes) of chickens of various origins have been studied by serological and biochemical methods. TwoB complexes are of particular interest:B ^R1, a recombinant haplotype derived from theB complexes of the inbred CB and CC strains, andB ^G-B1 , theB haplotype of the G-B1 strain. TheB ^R1 haplotype differs detectably from theB ^CB haplotype only at a locus controlling the synthesis of an antigen, B-G, which (in peripheral blood) is present only on red cells. Anti-B-G sera precipitate, from¹²⁵I-labeled red cell lysates, two chains of apparent molecular weights 42,000 and 31,000 (measured under reducing conditions); the smaller is perhaps derived by proteolysis from the larger. TheB ^G-B1 haplotype differs detectably from theB ^CC haplotype only at a locus controlling the synthesis of an antigen whose tissue distribution and biochemical and biological properties are identical to those of B-G. The chicken major histocompatibility complex therefore contains at least three loci—those controlling synthesis of the B-G, and of the previously defined B and B-L antigens.The following abbreviations are used in this paper MHC major histocompatibility complex - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - MLR mixed leukocyte reaction - GVH graft-versus-host reaction     

A new pollen type,C-banded and fluorochrome counterstained chromosomes,and evolution inGuatteria and related genera (Annonaceae)

Guatteria, Guatteriopsis, Guatteriella andHeteropetalum share the same conspicuous pollen type which is new for theSpermatophyta. It is zonoaperturate with a folded aperture region and an extremely reduced exine. First chromosome counts and karyotype analyses forGuatteriopsis (4 species investigated) andGuatteriella (1 species) are identical with those ofGuatteria (19 species seen): 2n = 28. The genome is characterized by diploidization and partly telocentric chromosomes. Sequentially Giemsa C- and fluorochrome banded chromosomes and interphase nuclei are described. The cuticular folding pattern is distinct forHeteropetalum only. Growth forms and ecology are reported for many species. The evolutionary pattern of theGuatteria group is discussed and compared with other genera and families.     

Chromosome studies on mosses from Austria,Czechoslovakia and other parts of Central Europe

Summary Reports are made on chromosomes in forty-six species from sixteen families of mosses. The plants were collected from all Austrian provinces, except the Tyrol and Vorarlberg, from Bohemia, Moravia, Slovakia, Bavaria, and also from Hungary and Italy. In addition to number or numbers of chromosomes reported for each species, observations are made on chromosome morphology, staining characteristics and behavior in meiosis or mitosis. The lowest numbers yet found are reported forDicranum fuscescens, n=8, andTimmia bavarica, n=8+1 m. For the latter species the study was made from mitotic configurations, and offers evidence that m-chromosomes may be normal members of the gametophytic set. These investigations reveal for the first time m-chromosomes inDicranoweisia cirrata, Pottia lanceolata, Timmia bavarica, Orthotrichum pumilum, Homalothecium lutescens, Brachythecium velutinum, Rhyncostegium murale, Rhyncostegiella pumila, andHypnum cupressiforme. Although the generaBartramia andPlagiopus previously have been characterized by different numbers of chromosomes, n=8 and n=7 respectively, plants ofPlagiopus oederi from Carinthia and Moravia were found, however, to have n=8. One species,Isopterygium seligeri, and one variety,Orthotrichum anomalum var.saxatile, have not been studied previously.Grateful acknowledgement is made to (1) the United States National Science Foundation for partial support of this investigation by a grant of research funds (NSF GB 6725) to Duke University, and (2) the Botanisches Institut der Universität Wien for laboratory space, equipment and use of the herbarium and library.     

Introgressive hybridization and evolution of a novel protein phenotype: Glue protein profiles in thenasuta-albomicans complex ofDrosophila

Glue proteins are tissue-specific proteins synthesized by larval salivary gland cells ofDrosophila. InDrosophila nasuta nasuta andD. n. albomicans of thenasuta subgroup, the genes that encode the major glue protein fractions are X-linked. In the present study, these X-linked markers have been employed to trace the pattern of introgression ofD. n. nasuta andD. n. albomicans genomes with respect to the major glue protein fractions in their interracial hybrids, called cytoraces. These cytoraces have inherited the chromosomes of both parents and have been maintained in the laboratory for over 400–550 generations. The analysis has revealed that cytoraces withD. n. albomicans X chromosome show eitherD. n. nasuta pattern or a completely novel pattern of glue protein fractions. Further, quantitative analysis also shows lack of correlation between the chromosomal pattern of inheritance and overall quantity of the major glue protein fractions in the cytoraces. Thus, in cytoraces the parental chromosomes are not just differentially represented but there is evidence for introgression even at the gene level.     

Karyotype evolution by centromeric fission inZamia (Cycadales)

The chromosome numbers of several species ofZamia from Mexico are reported.Z. paucijuga, distributed from central Oaxaca to Nayarit, has been found to have 2n = 23, 25, 26, 27 and 28. 2n = 28 is the highest chromosome number yet found in the cycads. Karyotypes of this species differ principally in the number of telocentric and metacentric chromosomes present in each; 2n = 23, 25, 26, 27 and 28 were found to have 5, 3, 2, 1 and 0 metacentric and 8, 12, 14, 16 and 18 telocentric chromosomes, respectively.Z. fischeri has been found to be 2n = 16,Z. furfuracea andZ. loddigesii 2n = 18.Zamia paucijuga on the basis of morphological and ecological characteristics, is considered to be an advanced member of this genus. Chromosome and karyotype evolution inZ. paucijuga may have occurred by centromeric fission of metacentric chromosomes; the karyotypes ofZ. paucijuga are strongly asymmetrical, suggesting that they evolved recently.     

Chromosome banding and pairing behaviour inFestuca andVulpia (Poaceae,Pooideae)

Giemsa C-banding patterns of the grassesFestuca rubra (2n = 6x = 42),Vulpia fasciculata (2n = 4x = 28) and their wild F1 hybrid ×Festulpia hubbardii (2n = 5x = 35) show marked differences between the chromosomes of the two parents that enable unequivocal identification ofFestuca andVulpia chromosomes in the hybrid. Moreover, meiotic banding patterns reveal that both homogenetic (Festuca-Festuca andVulpia-Vulpia) and heterogenetic (Festuca-Vulpia) pairing occurs in the F1. The significance of this in relation to the formation of backcrosses to theFestuca parent and to the evolution of theFestuca polyploid complex in general is discussed.     

Karyotype studies in seven species of cyprinidae

The present report embodies karyotype studies of seven Indian cyprinids. Of these, the karyotypes of five species possess chromosomes all of which are acrocentric. These areLabeo calbasu (Ham.) 2n=54,Puntius sophore (Ham.),P. conchonius Ham.,P. stigma (Ham.) 2n=50 andChela bacaila (Ham.) 2n=52. The karyotype ofCrossocheilus latius punjabensis Mukerji has 48 chromosomes, 12 of which are metacentric (2n=12m+36r's). InAmblypharyngodon mola (Ham.), 52 chromosomes form the diploid set which include 8 metacentrics. In none of the seven species morphologically distinguishable sex chromosomes were found.Literature on cyprinid karyotypes has been reviewed in order to understand in how far the cytological characters are in agreement with the classification based on morphological studies. The validity ofMatthey's notion of nombre fondamental to establish kinship between cytologically known cyprinid species has also been discussed.