Butterfly wing color patterns result from the arrangement of monochromatic scales containing chemical pigments and a delicate architecture that can cause interference or diffraction of light, generating iridescent colors. The latter mechanism is known as structural coloration and, despite its ecological importance, little is known about the molecular mechanisms underlying the development of this trait. The Southern Dogface butterfly, Zerene cesonia, exhibits sexually dimorphic development of ultraviolet wing reflectance. Males possess a UV-reflective patch on the forewing that results from nano-structures on the wing scales, which are absent in females. This dimorphism offers an excellent opportunity to explore the genetic mechanisms involved in pattern formation and cyto-structural variation. We used RNA-seq data from imaginal wing discs through late larval and pupal development to identify genes involved in the regulation of color pattern and scale structure formation. We identified candidate genes for the regulation of wing color pre-patterning and sexually-dimorphic development of wing scales. Our results provide a genomic resource for the identification and characterization of genes that participate in the regulation of wing development in pierid butterflies.

The Srivastava Lab is focused on the identification and characterization of genes that play a role in basement membrane remodeling. Previously, we identified putative basement membrane degraders through a genetic screen. One such gene has been suggested to play a role in the maintenance of the stem cell niche in Drosophila melanogaster, but no other information about the role this gene plays in development or disease has been published. Here, data are presented from experiments utilizing Drosophila genetics and immunohistochemistry that provide important insights on the biological role of this gene. Collagenase activity was up-regulated upon overexpression of this gene, confirming it as a basement membrane degrader. Additionally, RNA in-situ hybridization experiment results showed expression in the developing imaginal discs of the 3rd instar larva tissues. Overexpression and knockdown studies further demonstrated morphological defects in a number of tissues, including the wing and the eye, and are suggestive of apoptosis. Acridine orange staining confirmed that cell death occurred when the gene was overexpressed and a cleaved caspase antibody staining indicated that process to be caspase-mediated apoptosis.

Η μελέτη της φυλοσύνδετης μετάλλαξης wisertsl (1-21.7, 7E) της Drosophila melanogaster αποκάλυψε ότι υπεύθυνο για τους φαινότυπους φαγωμένα/τσιμπημένα φτερά, ελαφρώς ανώμαλα μάτια και τη θερμοευαισθησία είναι το γονίδιο CG32711, που ονομάσαμε wiser (wings scalloped-eyes rough). Το γονίδιο wiser είναι απαραίτητο για τη σωστή ανάπτυξη της Drosophila melanogaster. Η μετάλλαξη wisertsl χαρτογραφείται στη 5΄ ρυθμιστική περιοχή του γονιδίου wiser. Στην ίδια περιοχή χαρτογραφείται και η θανατογόνος μετάλλαξη wiserPL26. Παραπέρα μελέτη του γονιδίου wiser με τη χρήση αυτών των δυο μεταλλάξεων και του διαγονιδίου UAS-wiser αποκάλυψε ότι: α) Οι μεταλλάξεις wisertsl και wiserPL26 ενισχύουν το φαινότυπο των μεταλλάξεων Beadex1 και Serrate1. Το γονίδιο wiser αλληλεπιδρά με τα γονίδια Beadex και Serrate, τα οποία εμπλέκονται στην ενεργοποίηση του Notch μονοπατιού σηματοδότησης κατά μήκος του ραχιοκοιλιακού άξονα του φτερού. Η παρατήρηση αυτή δείχνει, ότι το wiser εμπλέκεται στον προσδιορισμό του ραχιοκοιλιακού άξονα. β) Η μετάλλαξη wisertsl σε ομοζυγωτία μειώνει σημαντικά την έκφραση των διαγονιδίων fringe-lacZ, m8-lacZ, wingless-lacZ, vestigial-lacZ και Distalless-lacZ, αλλάζει το πρότυπο έκφρασης του mβ-lacZ και δεν επηρεάζει την έκφραση του apterous-lacZ στους εμβρυικούς δίσκους του φτερού προνυμφών 3ου σταδίου. Τα αποτελέσματα αυτά έδειξαν, ότι το γονίδιο wiser δρα μετά το γονίδιο apterous και πριν το γονίδιο fringe, που είναι τροποποιητής του υποδοχέα Notch. Επομένως, η δράση του Notch εξαρτάται και από το wiser. γ) Εκτοπική έκφραση του διαγονιδίου UAS-wiser με οδηγό το ap-Gal4, έδειξε ότι διασώζει μερικώς το φαινότυπο apterous- αλλά όχι το φαινότυπο Serrate1. δ) Εκτοπική έκφραση του UAS-wiser με οδηγό το dpp-Gal4, επηρεάζει την έκφραση του wingless-lacZ αλλά όχι των apterous, fringe, mβ, m8, vestigial και Distalless στους εμβρυικούς δίσκους φτερού. ε) Η δημιουργία μιτωτικών κλώνων με το σύστημα FRT/FLP, σε θηλυκά άτομα wiserPL26/+ οδήγησε στη δημιουργία κλώνων +/+ και wiserPL26/wiserPL26 διαφορετικού μέγεθος στους εμβρυικούς δίσκους του φτερού. Οι πρώτοι (+/+), έχουν σημαντικά μεγαλύτερο μέγεθος από τους δεύτερους όταν συμβαίνουν στη περιοχή του εμβρυικού δίσκου που θα δώσει το φτερό του ακμαίου ατόμου. Στα ακμαία θηλυκά οι σωματικοί κλώνοι εκδηλώνονται με το φαινότυπο φαγωμένα φτερά. Σωματικοί κλώνοι παρατηρήθηκαν και στα μάτια των ακμαίων. Τα αποτελέσματα των μιτωτικών κλώνων δείχνουν ότι το γονίδιο wiser εμπλέκεται στον πολλαπλασιασμό των κυττάρων. Όλα τα παραπάνω αποτελέσματα, δείχνουν ότι το γονίδιο wiser είναι απαραίτητο για την ανάπτυξη του φτερού, καθώς εμπλέκεται στο σχηματισμό του ραχιοκοιλιακού άξονα και επηρεάζει τον πολλαπλασιασμό των κυττάρων. / The analysis of the X-linked wisertsl (1-21.7, 7E) mutation in Drosophila melanogaster has shown that responsible for the scalloped phenotype and the temperature sensitivity is the CG32711 gene, which we name wiser (wings scalloped-eyes rough). The gene wiser is essential for Drosophila development. The wisertsl mutation is mapped at the 5′ regulatory region of the gene CG32711. The wiserPL26 lethal mutation is mapped in the same region. Using these two mutations and a UAS-wiser transgene we have shown that: a) The wisertsl and wiserPL26 mutations increase the wing scalloping (phenotype) of the mutations Beadex1 and Serrate1. The genes Beadex and Serrate are implicated in the activation of Notch signaling pathway along the dorsal-ventral axis of the wing. This observation indicates that the wiser gene is involved in determination of dorsal-ventral axis. b) The wisertsl mutation in homozygous condition reduces substantially the expression of fringe-lacZ, m8-lacZ, wingless-lacZ, vestigial-lacZ and Distalless-lacZ transgenes, alters the expression pattern of mβ-lacZ and does not affect the expression of apterous-lacZ transgene in the wing imaginal disc. This indicates that the expression of fringe (a modifier of Notch receptor) is regulated by wiser too. c) Ectopic expression of UAS-wiser by the ap-Gal4 driver partially rescues apterous- but not Serrate1 phenotype. d) Ectopic expression of UAS-wiser by the dpp-Gal4 driver affects the expression of wingless and does not affects the expression of apterous, fringe, mβ, m8, vestigial and Distalless in the wing imaginal disc (revealed by the corresponding -lacZ strains). e) Induction of somatic clones with the FRT/FLP system in wiserPL26/+ mutants led to mitotic +/+ and wiserPL26/wiserPL26 clones of different sizes. The first clones were much larger than the second ones in the territory of wing pouch. Adult females with scalloped wings were also produced. These results indicate that the wiser gene is involved in cell proliferation. All the above findings suggest that the wiser gene is essential for wing development and cell proliferation.

Identifying homologous genes, that is genes from a common ancestor, is important in comparative genomic studies for understanding gene annotation and the predicted function of a gene. Several pieces of software, of which the most well-known is BLAST, have been developed for identifying homologues, but this can be challenging in non-model organisms where sometimes poor quality of genome assemblies and lack of annotation make it difficult to robustly identify homologues. The aim of this project was to build a bioinformatic framework for homology detection using genomes from non-model organisms. The approach developed used genome annotations, annotated polypeptide sequences and genome assembly sequences to detect homologous genes.The framework was applied to identify Drosophila melanogaster homologous wing development genes in the genomes of nine other insect species with the aim to understand the evolution of loss of wings. To identify changes related to wing loss, the homologous protein sequences obtained were aligned and phylogenetic trees were built from them. The aim of creating the multiple protein alignments and phylogenetic trees was to shed light on whether changes in gene sequences can be related to presence or absence of wings. From the set of 21 candidate wing development genes identified with literature and subsequent database searches, I tested eight and was successful in identifying homologues for all of them in eight of the 10 in sectgenomes. This was done using a combination of text searches in genome annotations, searches with Exonerate v. 2.4.0 alignment program in annotated polypeptide sequences and in genome assemblies. The eight genes chosen for testing the framework were based on initial finding of putative homologues in the eight insect genomes when using the first two steps of the framework. For the set of homologous wing development genes examined I was not able to identify any conclusive pattern of potential protein coding changes that correlated with loss of wings in these species. Improvement to the current pipeline could include using query sequences from closer relatives of the 8 test species than D. melanogaster and, of course, testing of the remaining wing development genes as well as further literature study of wing development genes. Together these could improve future studies on the evolution of wing loss in insects.

Insect wings are very specific and unique structures in animal kingdom. Wing morphology is a result of long-standing complicated evolutionary process and until recently the way how the wings have evolved is not completely clarified. The flight ability is one of the most important event in insect history because it allows them to exploit new habitats, escape from predators or find the sexual partner. Here we present the newly discovered material consisting of Palaeodictyoptera immature wings from the Upper Carboniferous (Westphalian A) of Poland. This order became extinct in the end of Permian, however during the Late Paleozoic was remarkably diversified. Until recently, number of adult palaeodictyopterans have been described, however the immature stages are relatively unknown due to lack of suitable fossils. Immature wings present in this thesis have undoubtedly palaeodictyopterous affinities with atribution within superfamilies Breyeroidea and Homoiopteroidea. However, their familial assignment into Breyeriidae (morphotype A) and Homiopteridae (morphotype B) based on fore wing venation characters is not definite because of wing venation limits in early ontogenetic stages. The aim of the present work is a complex description of 14 new palaeodictyopteriids immature wings, and to point out certain important...