| 1. |
- Söderpalm-Berndes, Cecilia, 1956-
(författare)
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Does acetylcholin play a role in mitosis of V79 Chinese hamster cells?
- 1997
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The first stages of mitosis are governed by a cascade of phosphorylations brought about by the universal MPF kinase complex. From a regulatory point of view, less is known about late mitotic events. The present study demonstrates that, firstly, V79 Chinese hamster cells produce acetylcholine and, secondly, that a set of acetylcholine receptor ligands can affect progression throughout mitosis.This was suggested using different approaches. Carbaryl and a-naphthol were used to induce a particular mitotic population consisting of cells with dislocated, unseparated chromosomes. Five distinct cell types can be scored, each representing different mitotic stages. Acetylcholine, as well as other acetylcholine receptor ligands, were found to change the frequencies of the five cell types, implying that mitotic progression be altered.Mitotic cell elongation was studied in more detail, using the experimental system with carbaryl and a-naphthol. The results indicate that cell elongation likely depends on actin filament formation and regulatory proteins functionally coupled to actin, as well as a maintained energy status. Also, some acetylcholine receptor might be involved in this cytoskeletal rearrangement. This was suggested by the finding that the acetylcholine analogue carbachol speeds up the formation of elongated cells and, in addition, that two acetylcholine receptor antagonists can decrease the frequency of elongated cells. Recording cells with a video time-lapse equipment demonstrated that acetylcholine receptor ligands can accelerate or decelerate progression through metaphase as well as anaphase, in line with the previous findings.Three different approaches were used to assess and evaluate the effects of acetylcholine receptor ligands on cell cleavage. Firstly, in the experimental system with carbaryl or a-naphthol complex concentration response curves were obtained for cleavage. Secondly, binuclear cells were induced by a two-hour treatment with five different acetylcholine receptor antagonists. Thirdly, video time-lapse recordings demonstrated that two acetylcholine receptor agonists, carbachol and nicotine, accelerates onset and progression of cleavage, respectively.Since the tested ligands affect different parts of mitosis it might be that there are a number of acetylcholine receptor types or affinity states which elicit signals in a time sequence. The role of acetylcholine could be to fine-tune mitotic progression.
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| 2. |
- Tatarenkov, Andrey, et al.
(författare)
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Evidence of a reproductive barrier between two forms of the marine periwinkle Littorina fabalis (Gastropoda)
- 1998
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Ingår i: Biological Journal of the Linnean Society. - 0024-4066. ; 63:3, s. 349-365
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Tidskriftsartikel (refereegranskat)abstract
- Studies of allozyme variation may reveal unexpected patterns of genetic variation which challenge earlier conclusions of species delimitations based on morphological data. However, allozyme variation alone may not be sufficient to resolve this kind of problem. For example, populations of the marine intertidal snail Littorina fabalis (= Littorina mariae) from wave exposed parts and from protected parts of the same shores are distinguished by different alleles of arginine kinase (Ark) while indifferent, or very nearly so, in another 29 loci. Intermediate populations have large deficiencies of exposed/sheltered heterozygote classes of Ark and we have earlier suggested habitat-related selection in this locus as the explanation. In this study we estimated growth rate of individual snails of different Ark-genotypes in three different habitats (exposed, sheltered and intermediate). In all habitats the snails homozygous for alleles of 'exposed' type grew faster and matured at a larger size than did snails homozygous for alleles of 'sheltered' types. This relationship was indirectly confirmed in three additional sites of intermediate exposure where exposed Ark-genotypes dominated among large (>8 mm) snails while the sheltered genotypes dominated among small (<5 mm) snails of truly sympatric samples. We furthermore found small differences in allele frequencies of two other loci (Pgi and Pgm-2) and in shell colour frequencies, comparing sympatric snails of exposed and sheltered Ark-homozygotes. Although we found no signs of habitat-related selection among snails of different Ark-genotype, or selection against heterozygotes, we cannot reject selection in Ark, as our experiments only covered one island, one season and grown-up snails. The coupling between allozyme and phenotypic characters in strictly sympatric samples of snails suggests the presence of two gene pools. Perhaps the large and small forms of L. fabalis represent very closely related cryptic taxa. However, introgression between them seems a possible explanation for the striking similarities in the vast majority of morphological and allozyme characters. (C) 1998 The Linnean Society of London.
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| 3. |
- Bartosch-Härlid, Anna
(författare)
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A new perspective on avian phylogeny - a study based on mitochondrial genomes
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The Class Aves (birds) is generally divided into two main groups - the Palaeognathae and the Neognathae. Paleognathous birds are the flightless ratites and the tinamous, while Neognathae include all other extant birds (more than 9,000 species). This opinion has prevailed for more than a century and is primarily based on morphological studies of the avian bony palate. In this study, the relationships between Palaeognathae and Neognathae along with other avian divergences have been investigated in phylogenetic analyses of complete mitochondrial (mt) genomes as well as in analyses of the mt cytochrome b gene. The complete mt genomes of the ostrich (Struthioniformes; Struthio camelus), the greater rhea (Struthioniformes; Rhea americana) and the rook (Passeriformes; Corvus frugilegus) were sequenced. At the time when this study was initiated and conducted, only one complete avian mt genome had been presented, that of the chicken (Galliformes; Gallus gallus), but this year four other complete avian mt genomes (representing the orders Anseriformes, Falconiformes and Passeriformes) were released. The phylogenetic analyses were carried out on the concatenated nucleotide and amino acid sequences (9,597 and 3,199 characters respectively) of twelve mt protein-coding genes from avian, mammalian, reptilian and piscine taxa. The results of the phylogenetic analyses did not support the most commonly accepted view of a basal divergence between paleognaths and neognaths. Instead, the earliest divergence among the taxa included was between the Passeriformes and all other birds including the ratites which were reconstructed at a more apical position in the tree as a sister group of galliforms and anseriforms. The origin of the Passeriformes was estimated to the early Cretaceous, about 120 MYBP (million years before present). A basal position of the ratites has in fact been questioned before, suggesting that ratites are not "primitive" birds but that ratite morphology includes a number of juvenile or neotenous traits (for example the downy plumage and the paleognathous palate) which have been maintained in adulthood. Because of the lack of pre-Tertiary passerine fossils (> 65MYBP) the order Passeriformes is generally regarded as one of the most recent avian lineages. It has been proposed, however, that the order Passeriformes originated already in the Cretaceous and that a recent origin of passerines is due to a misinterpretation of the fossil record. It is probable that the discrepancies between this study and the previous ones are attributable to the considerable length of the present dataset. Most importantly, however, the inclusion of a non-avian outgroup permits a different rooting strategy than used in most studies of avian relationships to date.
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| 4. |
- Bettencourt, Raul, 1963-
(författare)
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Hemolin, a versatile immune protein from the Cecropia moth
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Insects have become useful models for the study of innate immune mechanisms, due to their lack of antibodies and receptors involved in adaptive immune response. However, the molecules and mechanisms involved in primordial immune recognition are still poorly understood. Hemolin, originally cloned from Hyalophora cecropia, is a soluble and membrane associated Ig-related molecule which meets the criteria for a pattern recognition molecule. It is constitutively expressed in the hemolymph and up-regulated after bacterial injection. It was shown to bind specifically to the Lipid A moiety of LPS from bacteria and to associate with aggregates formed by hemocytes and bacteria. The aim of the present studies was to characterize the binding of hemolin to insect cells and further investigate the mechanisms behind. Earlier results shown in Manduca sexta were confirmed and it was shown that soluble hemolin has a deaggregating effect on naive hemocytes of H. cecropia. Moreover, hemolin increases phagocytic activity of hemocytes, especially when combined with LPS. This enhanced phagocytosis was correlated to the activity of protein kinase C and tyrosine phosphorylation. We revealed a number of cell adhesion characteristics of hemolin 1) a membrane associated form, (2) Ca2+-dependent homophilic binding, (3) glycosylation (4) importance of Ca2+ and carbohydrates on the binding to hemocytes. Based on our present results and the hemolin crystal structure, we have proposed a model for the interactions between soluble hemolin and its membrane form. The close relatedness to NCAMs prompted us to investigate its expression in other tissues than those originally described. It was found that hemolin is present in the retinal eye discs of pupae, in developing follicles during oogenesis and in embryos of H. cecropia. It was inferred that hemolin has a role in developmental processes in addition to its putative immune functions in insects. From a phylogenetic point of view, hemolin function is consistent with the assumption that non-self recognition molecules of the IgSF arose from cell-adhesion molecules with multiple functions.
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| 5. |
- Castillejo-Lopez, Casimiro
(författare)
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Repetitive DNA in search of a function - a study of telomeric and centromeric sequences in Chironomus
- 1998
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Repetitive DNA is quantitatively the main component of telomeres and centromeres, structures responsible for maintenance of the eukaryotic chromosome. The telomere is the specialized nucleoprotein complex that terminates linear chromosomes. In most species the DNA component consists of short repeats which are generated by the enzyme complex telomerase. However, there are important exceptions such as the Drosophila melanogaster telomeres which are elongated by retrotransposons and, as documented in the present thesis, a third telomeric system in Chironomus pallidivittatus in the form of arrays of 340 bp long complex tandem repeats which extend to the end of their chromosomes. Complex repeats are not elongated by telomerase and one aim of my work has therefore been to elucidate possible regeneration mechanisms for telomeres with such repeats. I have obtained evidence for DNA increase through DNA sequences of nontelomeric origin being inserted into the telomeric repeat array via gene conversion. Immunolocalization of reverse transcriptase related proteins in the telomeric puff of the related species C. thummi revealed, on the other hand, a possible link between the regeneration of Chironomus telomeric complex repeats and mechanisms used by other eukaryotes. Only seven of the eight pairs of chromosome termini in C. pallidivittatus have 340 bp repeats. The remaining telocentric end contains another repeat, the centromeric 155 bp unit, probably extending to the chromosome end. In the arrays of this repeat, a putative homologue of the mammalian centromeric CENP-B box has been found and characterized. Its interspersed distribution and its surrounding direct repeats suggest a mobile origin. It is present in different recombined forms, which could be related to a role in recombination as has been suggested for the human CENP-B box.
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| 6. |
- Halldén, Christer
(författare)
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Characterization and Use of a Multiplex PCR-based System: Random amplified Polymorphic DNA
- 1998
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Random amplified polymorhic DNA (RAPD) is a PCR-based marker system that makes use of universal sets of short oligonucleotides that amplify random DNA fragments under low-stringency conditions. The RAPD marker system has found widespread use and possesses a number of very positive features, being a multiplex and universal system. The present studies characterize and evaluate the properties of the RAPD marker system in a number of different applications. RAPD markers were compared with RFLP markers in their ability to determine genetic relationships between lines of oilseed rape and were found to have the same resolving power as RFLP markers have. A high density genetic linkage map was constructed in sugar beet using both RAPD and RFLP markers. Both types of markers show a high degree of clustering over the genetic map, compatiple with a strong distal localization of recombination. In regions of high marker density, dominant RAPD markers present in either linkage phase and codominant RFLP markers subclustered relative to each other. The subclustering could be attributed both to the mapping procedure and to genuine differences in the way RAPD and RFLP markers are recruited. DNA mixtures were used to investigate how reliably RAPD markers function as genetic markers. Competition occurs in the amplification of all RAPD fragments and is a major source of genotyping errors in RAPD analysis. Mixtures of multiple primers were used to analyse the properties of the RAPD system. Competition makes mixtures of more than to primers inefficient. Two-primer mixtures produces xn(n-1)/2 different products, of which at least xn2/4 are unique, and can generate the extremly large numbers of markers needed in bulked segregant analysis (BSA) and chromosome landing. BSA were used to isolate markers linked to a beet cystnematode resistence locus in sugar beet and fertility restorer gene for the Ogura cytoplasmic male sterility in oliseed rape. Graphical genotype information was shown to be useful in the selection of individuals used in BSA experiments and a set of dominant RAPD markers were used succesfully in a marker-assisted selection experiment
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| 7. |
- He, Hong
(författare)
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Organization of centromere associated DNA in Chironomus pallidivittatus.
- 1997
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Centromeres have not yet been functionally defined in higher eu- karyotes, largely due to the huge amounts of repeated DNA in these regions of most species and difficulties of delimiting centromeric from pericentric regions. Also the latter have large quantities of mostly noncoding DNA, expressed cytologically as pericentric heterochromatin. The dipteran Chironomus pallidivittatus is advantageous in having narrow centromeric/ pericentric regions, in thin chromosome bands containing about 50 kb of centromere specific tandem 155 bp repeats. We have described a mobile element characterized by a combination of an RNA polymerase III transcrip- tion unit and a large palindrome surrounding about 100 bp, some of which is partially palindromic, and the rest of which is highly AT-rich. Similar features also characterize the functionally defined centromeres in yeast. I have also defined the border between the block of 155 bp repeats in the centromere 2 region and adjoining pericentric repetitive DNA, which I have described up to the border towards unique DNA. There are several classes of pericentric repeats with different patterns of extracentromeric distri- butions. Except for the 155 bp repeats most sequences are absent from other centromeric/pericentric regions, speaking against a role related to centromeric function for these. Finally, I have identified a second centromere specific tandem repeat, 380 bp long, restricted to one of the four centromeres, which may be a functional equivalent of the 155 bp repeat.
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| 8. |
- Isaksson, Anders
(författare)
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Regulation of Drosophila Jun by phosphorylation and the ubiquitin system during eye development
- 1997
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The goal of these studies has been to elucidate the biological relevance of post-translational modifications in the context of a living organism. The model system used is photoreceptor differentiation during development of the compound eye in the fruit fly Drosophila melanogaster. Important signal transduction pathways in this process consist of receptor tyrosine kinases (RTKs) coupled to a mitogen activated protein kinase cascade. In the nucleus several transcription factors have been implicated as mediators of the pathway. The first part of the work presented here focuses on the role of signal-dependent phosporylation of D-Jun in photoreceptor differentiation. It appears that Rolled is the only kinase phosphorylating D-Jun during eye development. Interestingly, Rolled is a kinase of the extracellular signal reulated (ERK) sub-type of kinases, in contrast to the c-Jun N-terminal kiases (JNKs), which phosphorylate c-Jun in many cell culture systems. Phosphorylation by Rolled appears to be important for D-Jun function, since a D-Jun protein with the phosphorylation sites mutated to the non-phosphorylatable amino acid alanine can suppress formation of ectopic photoreceptors. Another study addresses the role of ubiquitin- dependent proteolysis in photoreceptor differentiation. A loss-of-function mutation in a gene involved in ubuquitin-dependent proteolysis (fat facets) causes a zygotic phenotype with 1-2 ectopic photoreceptors per unit eye. Genetic interactions with components of the neurogenic signal transduction pathway suggest that the pathway is hyperactivated in the faf mutants. This idea is consistent with the observed increased level and prolonged expression of D-Jun protein in faf eye imaginal discs. Since reducing D-Jun activity suppresses the formation of ectopic photoreceptors caused by faf, it is likely that the increased expression of D-Jun contributes to the phenotype.
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| 9. |
- Kaj, I, et al.
(författare)
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Probability of identity by descent in metapopulations
- 1999
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Ingår i: Genetics. ; 152:3, s. 1217-1228
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Tidskriftsartikel (refereegranskat)abstract
- Equilibrium probabilities of identity by descent (IBD), for pairs of genes within individuals, for genes between individuals within subpopulations, and for genes between subpopulations are calculated in metapopulation models with fixed or varying colony sizes. A continuous-time analog to the Moran model was used in either case. For fixed-colony size both propagule and migrant pool models were considered. The varying population size model is based on a birth-death-immigration (BDI) process, to which migration between colonies is added. Wright's F statistics are calculated and compared to previous results. Adding between-island migration to the BDI model can have an important effect on the equilibrium probabilities of IBD and on Wright's index.
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| 10. |
- Kamnert, Iréne
(författare)
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Classes of DNA associated with telomeres in the chironomids C. pallidivittatus and C. tentans
- 1997
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Telomeres in Chironomus consist of long complex tandem DNA repeats, which in C. pallidivittatus have been shown to extend to the extreme end of chromosomes. In C. pallidivittatus the telomeric repeats are present in large blocks, 50-200kb in size, most of them belonging to four major subfamilies. The main issue of this thesis has been to elucidate possible functional roles for these different subfamilies of telomeric repeats. Therefore, both intra- and intertelomeric mapping of the subfamilies were performed. One subfamily was found always to be located distally to the others, and have properties that could indicate a role in the protection of the chromosome ends. The subfamily identity of the most proximally situated repeat was identified in five of the seven non-telocentric chromosome ends. The junction between telomeric repeats and subterminal sequences was also characterized. In this region species and junction specific mutations were found, which were highly conserved between different chromosome ends within the same species. The homogenization pattern of these mutations indicated that gene conversion can act also between heterologous telomeres. The telomeric repeats exhibit a remarkable efficient homogenization whether they are located in the same or in different non-homologous chromosomes, an unusual feature since intrachromosomal homogenization usually are more efficient than homogenization between heterologues. Results from several earlier investigations had suggested transcriptional activity in the telomeric repeats in Chironomus. This issue was now addressed both in relation to possible roles for the different subfamilies in this transcription, and if this could be a connection to a process active in the elongation of the telomeres.
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