טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentHamdan Rana
SubjectIdentifying Meru, a Regulator of Cell Polarity, as a Gene
Required for Normal Development of Chordotonal
Organs
DepartmentDepartment of Medicine
Supervisor Professor Adi Salzberg
Full Thesis textFull thesis text - English Version


Abstract

            In the fruit fly, Drosophila melanogaster, sensory feedback from within the body is mediated by stretch-receptive chordotonal organs (ChO) and specific subtypes of multiple dendritic neurons. We are using ChO development as a model for studying cell fate determination and diversification of cell properties.

Eight ChO develop in each abdominal hemisegment of the larva, five of them are clustered in the prominent lateral pentascolopidial organ (LCh5). Each ChO originates from a single precursor that goes through a series of asymmetric cell divisions to produce five different cells: a neuron, a scolopale, a cap, a ligament and a cap-attachment (CA) cell. The cap and the ligament cells of the LCh5 organ are anchored to the cuticle by two cap-attachment (CA) cells and one ligament-attachment (LA) that is recruited through an EGFR- dependent mechanism.

An RNAi screen that focused on the ChO accessory and attachment cells and was aimed at identifying new genes required for ChO morphogenesis, has been recently performed in our lab. The screen identified 31 candidate genes required for different aspects of ChO morphogenesis. The genes identified in the screen were grouped into three general phenotypic categories based on the most prominent phenotypic feature they presented: (A) Loss or gain of GFP or RFP expression (fluorescent markers used in the screen), often combined with abnormal morphology of cells. (B) Defective attachment or cell morphology, and (C) Abnormal pattern of cell elongation.

I have joined the RNAi screen project after a primary collection of candidate genes has been established and further characterized some of the phenotypes identified in the screen.

My main focus were three of the 31 genes, CG13653, CG8258 and CG32150, that had no known function at that time. In the course of the work, a paper about the role of the CG32150 gene in external sensory organs was published and the gene was named meru, (Banerjee et al., 2017).  The role of meru in the ChO lineage became the main focus of my research.

A knockdown of meru caused a loss of CA cells in addition to abnormal localization of the cap cells’ nuclei. Based on the findings of Banerjee et al, I hypothesized that meru affects cell fate decision and/or differentiation of the ChO cells. I characterized the distribution of Meru in the ChO using a GFP-meru knock-in strain and found that meru was expressed asymmetrically in the ChO precursors. Later in development meru accumulates at the highest levels in the neurons. To validate the RNAi-induced phenotypes I analyzed the phenotype of meru1 null mutant. I found that the loss of meru led to abnormal number of CA, cap, neurons, scolopale, and ligament cells within the ChO lineage, pointing to defects in ACD. However, no cell fate transformations were evident. Rather, the various cells failed to differentiate properly as suggested by immunohistochemical analyses with a battery of cell-specific markers performed in 3rd instar larvae.