טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentGlatt Yael
SubjectThe Role of Dpax2 in the Organogenesis of Drosophila
Melanogaster Proprioceptors
DepartmentDepartment of Medicine
Supervisor Professor Adi Salzberg
Full Thesis textFull thesis text - English Version


Abstract

In each abdominal hemisegment of Drosophila melanogaster embryos eight Chordotonal Organs (ChOs) are developed. ChOs are internal stretch receptors that provide proprioceptive feedback on larval locomotion. Five of them are grouped together and form the lateral pentascolopidial organ (LCh5). Each unit comprising the LCh5 arises from a single precursor cell through a series of asymmetric cell divisions, producing five different cells. A Neuron cell ensheathed by a Scolopale (Sc) cell are the core sensory unit of this organ. These two cells are suspended by two support cells - the Cap (C) and the Ligament (L) cells that in turn are anchored to the body wall by the Cap Attachment (CA) cell and an additional attachment cell - the Ligament attachment (LA) that is recruited to the LCh5 at a later stage.  This specific way of ChOs’ development serves as an excellent model for studying diversification of cell properties and cell fate decisions during organogenesis.

In this work we identify Dpax2, the Drosophila homolog of the vertebrate Pax2 gene, as an important determinant of ChOs’ organogenesis. We show that Dpax2 has an essential role in the differentiation of the Cap cells and that it exerts at least some of its effects on their differentiation through the direct regulation of Delilah (Dei) - a bHLH transcription factor, expressed in the attachment and support cells of ChOs and required for their terminal differentiation. We also determine that Dpax2’s expression in the ChOs is regulated via the two separate (early and late) modules of the “shaven” enhancer, that also regulate Dpax2’s expression in the External sensory organs (ESOs), and that, as in the ESOs, the late enhancer that maintains Dpax2 expression in the C and Sc cells is regulated by DPax2 itself through positive feedback loop.

Flies bearing strong loss-of-function mutations in the “shaven” enhancer of Dpax2 exhibited loss of several Cap cell-specific markers, including Dei; while over-expressing Dpax2 in the ChO lineage resulted in the ectopic expression of Dei in the Sc cells and what appears to be additional C cells. Furthermore, we demonstrated that the elimination of a minimal fragment of the deiChO enhancer, containing three putative DPax2 binding sites, by CRISPR/Cas9, completely abolished Dei’s expression in the support cells (C and L) while its expression in the attachment cells remained intact. Therefore we suggest that DPax2 is an essential transcriptional regulator of C cell differentiation at least in part through its direct regulation of dei.