Thesis--University of Oklahoma. / Includes bibliographical references.

Thesis (Ph. D.) -- University of Texas Southwestern Medical Center at Dallas, 2005. / Vita. Bibliography: 169-215.

Thesis (Ph. D.) -- University of Texas Southwestern Medical Center at Dallas, 2005. / Vita. Bibliography: 169-215.

Thesis (Ph. D.) -- University of Texas Southwestern Medical Center at Dallas, 2005. / Vita. Bibliography: 169-215.

Thesis (Ph. D.) -- University of Texas Southwestern Medical Center at Dallas, 2005. / Vita. Bibliography: 169-215.

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Στα πλαίσια της παρούσας διδακτορικής διατριβής μελετήθηκαν οι φυλογενετικές σχέσεις των χρωμοσωματικών φυλών του είδους M. thomasi με κυτταρογενετικές και μοριακές προσεγγίσεις. Συλλέχθηκαν συνολικά 408 άτομα από 65 τοποθεσίες της Ελλάδας, της Αλβανίας και του Μαυροβουνίου, που μελετήθηκαν με κλασικές κυτταρογενετικές μεθόδους (πρότυπο G & C ζώνωσης), συγκριτική μοριακή κυτταρογενετική ανάλυση με δείκτες τελομερικές και δορυφορικές αλληλουχίες, μοριακή φυλογένεση με δείκτες μιτοχονδριακούς γενετικούς τόπους (cytb, D-loop) και εργαστηριακές διασταυρώσεις ανάμεσα σε άτομα από διαφορετικές χρωμοσωματικές φυλές. Τα αποτελέσματα της παρούσας διατριβής δείχνουν ότι οι σκαπτοποντικοί από την Αττική και την Εύβοια παρουσιάζουν σημαντικές διαφορές σε σχέση με τους υπόλοιπους πληθυσμούς, σε όλες τις προσεγγίσεις που χρησιμοποιήθηκαν και προτείνεται να θεωρηθούν ως διακριτό είδος με το όνομα Microtus atticus, όπως είχε αρχικά περιγραφεί από τον Miller το 1910. Τα δύο αδελφά είδη M. thomasi και M. atticus έχουν παρόμοιο πρότυπο C και G ζώνωσης στα αυτοσωματικά χρωμοσώματα, αλλά διαφέρουν ως προς την σύσταση της ετεροχρωματίνης των Χ χρωμοσωμάτων (στο Μ. atticus αποτελείται κυρίως από τη δορυφορική αλληλουχία Mth-Alu900, ενώ στο M. thomasi από τελομερικές αλληλουχίες), παρουσιάζουν αναπαραγωγική απομόνωση (σε εργαστηριακές συνθήκες δίνουν στείρα F1 υβρίδια) και παρουσιάζουν γενετικές αποστάσεις, παραπλήσιες με άλλα στενά συγγενικά είδη Microtus ως προς τους μιτοχονδριακούς γενετικούς τόπους (cytb: 2,7% και D-loop: 2,6%). Τα δύο παραπάνω είδη παρουσιάζουν εκτεταμένους χρωμοσωματικούς πολυμορφισμούς που ομαδοποιούνται σε εννέα χρωμοσωματικές φυλές στο είδος M. thomasi: “thomasi” (2n=44, FN=44), “peloponnesiacus" (2n=44, FN=46), “Tichio” (2n=42, FN=44), “Rb-subalpine”(2n=40, FN=42), “subalpine” (2n=42, FN=42), “Preveza” (2n=40, FN=42), “Kali” (2n=40, FN=42), “Aridea” (2n=38, FN=42), “Edessa”(2n=38, FN=40) και δύο φυλές στο είδος M. atticus: "atticus" (2n=44, FN=46), "Evia" (2n=44, FN=44). Με βάση τα αποτελέσματα των εργαστηριακών διασταυρώσεων, τη φυλογενετική και την κυτταρογενετική μελέτη, πραγματοποιήθηκε μία εκτεταμένη αναθεώρηση της προέλευσης των ήδη γνωστών χρωμοσωματικών φυλών, ενώ βρέθηκαν και περιγράφηκαν νέες χρωμοσωματικές φυλές. Οι πληθυσμοί σκαπτοποντικών από την Αττική και την Πελοπόννησο, που σύμφωνα με παλαιότερες εργασίες, είχαν περιγραφεί ως "atticus" και άνηκαν στο είδος M. thomasi, στην παρούσα διδακτορική διατριβή διαχωρίζονται σε δύο χρωμοσωματικές φυλές, που κατατάσσονται στα δύο διαφορετικά είδη. Οι πληθυσμοί από την Αττική ανήκουν στη φυλή "atticus" και κατατάσσονται στο είδος M. atticus, ενώ οι πληθυσμοί σκαπτοποντικών από την Πελοπόννησο θεωρούνται ως μία διακριτή φυλή, την "peloponnesiacus" και κατατάσσονται στο είδος M. thomasi. Οι χρωμοσωματικές φυλές του είδους M. thomasi: “Tichio”, “Preveza”, “Kali” και "Edessa" που είχαν αναφερθεί ως πολυμορφισμοί σε παλαιότερες εργασίες, περιγράφονται πια ως διακριτές χρωμοσωματικές φυλές. Επίσης, περιγράφονται για πρώτη φορά η χρωμοσωματική φυλή “Aridea” (2n=38, FN=42) του είδους M. thomasi και η χρωμοσωματική φυλή "Evia" (2n=44, FN=44) του είδους M. atticus. Οι χρωμοσωματικές φυλές "atticus" του είδους M. atticus και "peloponnesiacus", "subalpine", "Kali" και "Edessa" του είδους M. thomasi διασταυρώθηκαν στο εργαστήριο με την θεωρούμενη ως πιο πρωτόγονη φυλή "thomasi" του είδους M. thomasi. Η χρωμοσωματική φυλή "atticus", που ανήκει στο είδος M. atticus, έδωσε στείρους απογόνους με τις φυλές "peloponnesiacus" και "thomasi", που ανήκουν στο είδος M. thomasi. Οι σκαπτοποντικοί της φυλής "subalpine" έδωσαν γόνιμους απογόνους στην F1 γενεά, στην F2 γενεά και στις ανάδρομες διασταυρώσεις, όταν διασταυρώνονται με σκαπτοποντικούς της φυλής "thomasi". Αντίθετα, οι σκαπτοποντικοί της φυλής "Kali" έδωσαν στείρους απογόνους στην F2 γενεά, ενώ οι σκαπτοποντικοί της φυλής "Edessa" έδωσαν στείρους απογόνους στην F1 γενεά, όταν διασταυρώνονται με σκαπτοποντικούς της φυλής "thomasi", αν και ανήκουν στο ίδιο είδος. Η αναπαραγωγική απομόνωση των φυλών "Kali" και "Edessa", από τη φυλή "thomasi" οφείλεται σε χρωμοσωματικές ασυμβατότητες και αποτελεί ένδειξη ότι η γενετική διαφοροποίηση των χρωμοσωματικών φυλών μπορεί να οδηγήσει υπό προϋποθέσεις σε ειδογένεση. Με βάση την παλαιογεωγραφία του ελληνικού χώρου και την συγκριτική ερμηνεία των αποτελεσμάτων που προέκυψαν από όλες τις προσεγγίσεις, παρουσιάζεται ένα πιθανό σενάριο της εξελικτικής ιστορίας των χρωμοσωματικών φυλών των ειδών M. thomasi και M. atticus. Υποθέτουμε ότι η προγονική μορφή των δύο ειδών βρισκόταν σε παγετώδες καταφύγιο της Νότιας Ελλάδας κατά την τελευταία παγετώδη περίοδο (πριν από 10.000-125.000 χρόνια). Εξαιτίας οικολογικών ή/και παλαιοκλιματολογικών παραγόντων, ο αρχικός πληθυσμός διασπάστηκε σε δύο υποπληθυσμούς, που σταδιακά διαφοροποιήθηκαν σε διακριτά είδη. Με το τέλος της πρόσφατης παγετώδους περιόδου, η σταδιακή βελτίωση των κλιματολογικών συνθηκών, επέτρεψε στο είδος M. thomasi να αποικίσει την Κεντρική και Βόρεια Ελλάδα, την Αλβανία και το Μαυροβούνιο, όπου διαμορφώθηκαν σταδιακά όλες οι γνωστές χρωμοσωματικές φυλές. Αντίθετα, οι αλλαγές στο ηπειρωτικό ανάγλυφο, όπως η άνοδος της στάθμης της θάλασσας και ο αποχωρισμός της Εύβοιας από την Στερεά Ελλάδα, απομόνωσαν το είδος M. atticus στην περιοχή της Αττικής και της Εύβοιας αποτρέποντας την περαιτέρω εξάπλωσή του. / During the current PhD project, the phylogenetic relationships of the chromosomal races of the underground vole Microtus thomasi were examined through cytogenetic and molecular approaches. In total, 408 voles were collected from 65 localities of Greece, Albania and Montenegro, in order to clarify their phylogenetic relationships with classical cytogenetic techniques (G and C banding), comparative cytogenetic analysis with telomeric and satellite markers, molecular phylogeny with markers mtDNA markers (cytb, D-loop) and hybridization experiments between voles from distinct chromosomal races. Our results indicate that the voles from Attiki and Evia seem to be differentiated from other populations, based in all applied approaches. In this context, we propose that the populations from Attiki and Evia should be considered as a distinct species, validating the species name Microtus atticus, as it has been originally described by Miller at 1910. The two sibling species, Microtus thomasi and Microtus atticus have karyotypes with similar G and C banding pattern, but differ in the content of the X chromosome heterochromatin (mainly Mth-Alu900 satellite sequence in M. atticus, telomeric repeats in M. thomasi), appear reproductively isolated (produce sterile F1 hybrids when crossed in laboratory conditions) and the between species genetic distances were calculated similar to other closely related Microtus species (2,7% for cytb and 2,6% for D-loop). The two species demonstrate an extensive chromosomal polymorphism, resulting in the description of nine chromosomal races in M. thomasi: “thomasi” (2n=44, FN=44), “peloponnesiacus" (2n=44, FN=46), “Tichio” (2n=42, FN=44), “Rb-subalpine”(2n=40, FN=42), “subalpine” (2n=42, FN=42), “Preveza” (2n=40, FN=42), “Kali” (2n=40, FN=42), “Aridea” (2n=38, FN=42), “Edessa”(2n=38, FN=40) and two chromosomal races in M. atticus: "atticus" (2n=44, FN=46), "Evia" (2n=44, FN=44). Based on the results from the hybridization experiments, the molecular phylogenetic and the cytogenetic analysis, we proceed to the re-evaluation of the known chromosomal races and the description of new chromosomal races. The voles from Attiki and Peloponnesus, which were described to the "atticus" chromosomal race of the species M. thomasi according to previous publications, are distinguished in two races, which are attributed to distinct species. In fact, the voles from Attiki are attributed to "atticus" race and classified in M. atticus, while the voles from Peloponnesus are attributed to a new race, named "peloponnesiacus" and classified to M. thomasi. The race "atticus", consisting of the populations from Attiki and Peloponnesus according to previous studies, was distinguished in two distinct, reproductively isolated races: "atticus" (2n=44, FN=46) (populations from Attiki) attributed to M. atticus, and "peloponnesiacus" (2n=44, FN=46) (populations from Peloponnesus), attributed to M. thomasi. Furthermore, we described four chromosomal races (“Tichio”, “Preveza”, “Kali” και "Edessa") in the species M. thomasi, which were referred in previous works as chromosomal polymorphisms. In the same context, we described for first time the chromosomal race "Aridea" (2n=38, FN=42) in the species M. thomasi and the chromosomal race "Evia" (2n=44, FN=44) in the species M. atticus. The chromosome races "atticus" of the species M. atticus and "peloponnesiacus", "subalpine", "Kali" and "Edessa" of the species M. thomasi, were crossed in laboratory conditions with the most primitive form "thomasi" of the species M. thomasi. The voles from the chromosomal race "atticus" (M. atticus) produced sterile F1 hybrids, when crossed with voles from the chromosomal races "thomasi" and "peloponnesiacus" (M. thomasi). The voles from the chromosomal race "subalpine" produced fully fertile hybrids in F1, F2 and backcrosses generations, when crossed with the voles from the chromosomal race "thomasi". On the contrary, the voles from the chromosomal races "Kali" and "Edessa" produced sterile hybrids in F2 and F1 generations, when crossed with the voles from the chromosomal race "thomasi", despite the fact that all three races belong to the same species. The reproductive isolation of the chromosomal races "Kali" and "Edessa" could be attributed to chromosomal incompatibilities and indicates that the genetic differentiation of the chromosomal races could eventually lead to speciation. Based on the geographical traits of the Southern Balkans and the comparative interpretation of our results, we are able to present a possible scenario of the evolutionary history of the chromosomal races of the species M. thomasi and M. atticus. We assume that the primitive form was distributed in a glacial refugium of Southern Greece, during the last Glacial period (10.000-125.000 BP). Due to ecological or/and climatological factors, the ancestral population was separated in two subpopulations, which evolved to distinct species. At the end of the last glacial period, the global warming allowed the voles of M. thomasi to expand northwards to the Balkans, colonizing unoccupied areas of Greece, Albania and Montenegro and establishing all known chromosomal races. On the contrary, the changes of the landscape, due to the rise of the sea level and the separation of Evia Island from the Greek mainland, further isolated the M. atticus populations in Attiki and Evia, prohibiting the northwards colonization.

Submitted by ALLISON KLEITON DOS ANJOS null (allison__anjos@hotmail.com) on 2018-01-15T10:50:34Z No. of bitstreams: 1 TESE FINAL ALLISON.pdf: 3595248 bytes, checksum: 33a447746191e365878b383ffd929cc8 (MD5) / Rejected by Adriana Aparecida Puerta null (dripuerta@rc.unesp.br), reason: Prezado Allison Kleiton dos Anjos, Solicitamos que realize uma nova submissão seguindo as orientações abaixo: - Resumo em língua estrangeira (inglês) - Faltou o Abstract no documento enviado. Este item é elemento obrigatório de acordo com as normas de trabalhos do seu Programa de Pós Graduação e deve vir logo após o Resumo em Português. Agradecemos a compreensão e aguardamos o envio do novo arquivo. Atenciosamente, Biblioteca Campus Rio Claro Repositório Institucional UNESP on 2018-01-15T16:52:52Z (GMT) / Submitted by ALLISON KLEITON DOS ANJOS null (allison__anjos@hotmail.com) on 2018-01-15T18:30:08Z No. of bitstreams: 1 TESE_FINAL_ALLISON.pdf: 4473839 bytes, checksum: ec9f453166f94072eff2f68fc43791e7 (MD5) / Approved for entry into archive by Adriana Aparecida Puerta null (dripuerta@rc.unesp.br) on 2018-01-15T18:37:51Z (GMT) No. of bitstreams: 1 anjos_ak_dr_rcla.pdf: 4115054 bytes, checksum: 5a5c208184ffdb419015062384640e33 (MD5) / Made available in DSpace on 2018-01-15T18:37:51Z (GMT). No. of bitstreams: 1 anjos_ak_dr_rcla.pdf: 4115054 bytes, checksum: 5a5c208184ffdb419015062384640e33 (MD5) Previous issue date: 2017-12-15 / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Os hemipteros da infraordem Cicadomorpha são representados por aproximadamente 30.000 espécies de insetos sugadores distribuídos mundialmente. Apesar de se destacarem por causarem muitos prejuízos na agricultura e pecuária pouco se sabe sobre a variabilidade genética e cromossômica desses animais que apresentam cromossomos holocentricos. Os DNAs repetitivos são uma ferramenta útil em estudos de diversificação cariotípica, organização e evolução dos genomas. Deste modo, este trabalho teve como objetivo contribuir com o conhecimento sobre a dinâmica dos cariótipos de representantes de Cicadomorpha e a organização de DNAs repetitivos, especificamente: I. analisar o cariótipo de espécies pertencentes a quatro famílias de Cicadomorpha, bem como caracterizar a heterocromatina constitutiva quanto a riqueza de pares de base; II. inferir a respeito da dinâmica evolutiva de famílias gênicas por meio do mapeamento cromossômico; III. analisar a organização cromossômica e testar a conservação interespecífica da sequência telomérica TTAGGn do pool de DNAs repetitivos obtidos (fração C0t) de espécies do gênero Mahanarva e das sequências teloméricas; IV. Analisar o satelitoma de Mahanarva quadripunctata e comparar os diferentes DNAs satélites de seu genoma com o de outras espécies do gênero visando entender a organização e evolução dos satDNAs neste grupo. Os dados obtidos revelam ampla variabilidade cromossômica entre as distintas famílias, causada principalmente por fusões cromossômicas, entretanto dentro de uma mesma família os cariótipos tendem a apresentar menos variações ao nível macrocromossômico. Embora os Cicadomorpha apresentem variabilidade em números diplóides a organização dos DNAs repetitivos é bastante conservada, mesmo em famílias distantes filogeneticamente, sugerindo estabilidade. Alguns dados foram analisados baseados na filogenia da infraordem, sendo sugerido os possíveis padrões ancestrais. Além disso, as possíveis causas da variabilidade em relação aos padrões modais são sugeridos. Os dados apresentados são um avanço no conhecimento da organização de DNAs repetitivos em Cicadomorpha, sendo para algumas sequências a primeira vez que se realiza algum estudo. / The hemipterans of the Cicadomorpha infraorder are represented by approximately 30.000 species of sucking insects distributed worldwide. Although they stand out by cause many damages in agriculture and livestock, they are poorly studied regarding the genetic and chromosomal variability of these animals that present holocentric chromosomes. Repetitive DNAs are a useful tools in studies of karyotypic diversification, organization and evolution of genomes. The aim of this work was to contribute with knowledge about the dynamics of the karyotypes of Cicadomorpha and the organization of repetitive DNAs, specifically: I. Analyze the karyotype of species belonging to four families of Cicadomorpha, as well as to characterize the constitutive heterochromatin regarding base pairs richess; II. Infer about the evolutionary dynamics of multigene families through the chromosomal mapping; III. Analyze the chromosomal organization and test the interspecific conservation of the telemetric repeat TTAGGn from the pool of repetitive DNA fraction (C0t fraction) of Mahanarva species and telomeric sequences; IV. Analyze the satellitome of Mahanarva quadripunctata and compare the different satellite DNAs of its genome with that from other species of the genus in order to understand the organization and evolution of satDNAs in this group. The data obtained reveal a wide chromosomal variability among the different families, caused mainly by chromosomal fusions, however within a same family the karyotypes tend to present less variations at the macro-chromosomal level. Although Cicadomorpha exhibit variability in diploid numbers, the organization of repetitive DNAs is highly conserved, even in phylogenetically distant families, suggesting stability. Some data were analyzed based on the phylogeny of the infraorder, suggesting possible ancestral patterns. In addition, the possible causes of variability relative to modal patterns are suggested. The data presented is an advance in the knowledge of the organization of repetitive DNAs in Cicadomorpha, being for some sequences the first time that some study is done. / FAPESP: 2014/06226-3.

The mammalian Y chromosome contains genes important for male sexual maturity and reproduction. The mouse Y chromosome long arm harbours a number of multi-copy genes whose absence or reduced representation has been linked to sperm defects and offspring sex ratio distortion in favour of females. Understanding the biological mechanisms of sex ratio distortion and related sperm aberrations could not only result in benefits for fertility research, but also in the development of methods for large scale animal breeding pre-implantation sex selection. The distortion has been linked to an intragenomic conflict between the X and Y chromosomes that impacts spermiogenesis. Since the proportion of X- and Y-bearing sperm does not differ in affected animals, and there is no selective destruction of male embryos post-fertilisation, a functional difference must exist between the X- and Y-bearing sperm. This thesis describes the investigation into the physiological and molecular mechanisms of a large Y-chromosome long arm deletion in the mouse model MF1XYRIIIqdel. The examination of physiological characteristics revealed a distinct sperm morphology within the deletion model. Detailed characterisation of sperm shape demonstrated that aberrations consistently occur within specific regions of the sperm head, linking the distorted morphology to particular maturation stages in the sperm cycle. Using sperm fluorescence in situ hybridisation, a novel and detailed comparison of X- and Y- bearing sperm has shown that a subtle distinction in shape also exists between the X- and Y- bearing sperm in the deletion model. Breeding data were examined and showed a skew towards female offspring and a slightly reduced litter size. Sperm enzyme activity assays did not reveal altered hyaluronidase activity in MF1XYRIIIqdel sperm. Physiological differences between X- and Y- bearing sperm must result from differential gene expression, complicated by the syncitial nature of sperm development. To explore this, a detailed molecular characterisation of the MF1XYRIIIqdel phenotype in developing haploid spermatids was performed. Cellular elutriation and fractionation techniques were employed to separate spermatids at different stages of maturation and isolate different subcellular compartments. Differences in the transcriptional profile between these populations were analysed by microarray and RNA sequencing analysis of total and micro RNA. This work yielded a collection of coding and non-coding transcripts which show distinctive expression and compartmentalisation differences between the deletion model and its wild-type counterpart across several sperm maturation stages. Combining these strategies has led to the identification of several gene products potentially implicated in observed physiological differences and the offspring sex ratio skew, providing candidate genes for further research.

It is known that chromosomal rearrangements play an important role in speciation by limiting gene flow within and between species. Furthermore, this effect may be enhanced by involvement of sex chromosomes that are known to undergo fast evolution compared to autosomes and play a special role in speciation due to their engagement in postzygotic reproductive isolation. The work presented in this study uses various molecular-genetic and cytogenetic techniques to describe karyotype and sex chromosome evolution of two groups of Lepidoptera, namely selected representatives of the family Tortricidae and Leptidea wood white butterflies of the family Pieridae. The acquired knowledge points to unexpected evolutionary dynamics of lepidopteran karyotypes including the presence of derived neo-sex chromosome systems that originated as a result of chromosomal rearrangements. We discuss the significance of these findings for radiation and subsequent speciation of both lepidopteran groups.