piRNA Function and Biogenesis in the <em>Drosophila</em> Female Germline: A Dissertation
Authors
Klattenhoff, Carla AndreaFaculty Advisor
William Theurkauf, Ph.D.Academic Program
Interdisciplinary Graduate ProgramUMass Chan Affiliations
Program in Molecular MedicineDocument Type
Doctoral DissertationPublication Date
2008-11-20Keywords
RNASmall Interfering
Drosophila Proteins
Biogenesis
Germ Cells
Body Patterning
Cell Cycle Proteins
Mutation
Amino Acids, Peptides, and Proteins
Animal Experimentation and Research
Cells
Embryonic Structures
Genetic Phenomena
Nucleic Acids, Nucleotides, and Nucleosides
Metadata
Show full item recordAbstract
The studies presented in this thesis addressed mainly two aspects of Piwi-interacting RNA (piRNA) biology in the Drosophilagermline. We investigated the role of the piRNA pathway in embryonic axis specification. piRNAs mediate silencing of retrotransposons and the Stellate locus. Mutations in the Drosophila piRNA pathway genes armitage and aubergine disrupt embryonic axis specification, triggering defects in microtubule polarization and asymmetric localization of mRNA and protein determinants in the developing oocyte. Mutations in the ATR/Chk2 DNA damage signal transduction pathway dramatically suppress these axis specification defects, but do not restore retrotransposon or Stellatesilencing. Furthermore, piRNA pathway mutations lead to germline-specific accumulation of γ-H2Av foci characteristic of DNA damage. We conclude that piRNA based gene silencing is not required for axis specification, and that the critical developmental function for this pathway is to suppress DNA damage signaling in the germline. We have also identified a new member of the piRNA pathway. We show that mutations in rhino, which encodes a rapidly evolving Heterochromatin Protein 1 (HP1) chromo box protein, lead to germline specific DNA break accumulation, trigger Chk2 kinase dependent defects in axis specification, and disrupt germline localization of Piwi proteins. Mutations in rhino and the piRNA pathway gene armitage disrupt silencing of all major transposon families, but do not alter expression of euchromatic or heterochromatic protein coding genes. Deep sequencing studies show that rhino mutations significantly reduce or eliminate anti-sense piRNAs derived from the majority of transposable elements in the Drosophila genome, and lead to a dramatic reduction in piRNAs derived from major piRNA production clusters on chromosomes 2R and 4. Rhino protein localizes to distinct nuclear foci, and associates with the chromosome 2R and 4 clusters by chromatin immunoprecipitation. The Rhino HP1 homologue is therefore required for piRNA biogenesis, transposon silencing, and maintenance of germline genome integrity.DOI
10.13028/fkqn-kw46Permanent Link to this Item
http://hdl.handle.net/20.500.14038/31718Rights
Copyright is held by the author, with all rights reserved.ae974a485f413a2113503eed53cd6c53
10.13028/fkqn-kw46