Study of the functional role of the Set protein in the early developmental stages of the model organism Danio rerio (zebrafish) (Doctoral thesis)

Σερίφη, Ηλιάνα

SET oncoprotein is an evolutionarily conserved protein that is widely expressed in all cell types and tissues. It is a multifunctional protein implicated in diverse cellular processes including chromatin remodelling, gene expression, cell cycle regulation, apoptosis and migration. SET is a potent inhibitor of the protein phosphatase 2A (PP2A). It has also been shown that SET is involved in the development of Alzheimer’s disease as well as of various types of cancer, which has led to new studies towards its use as a target in cancer treatment. The aim of this work was to understand the physiological roles of SET at the organismal level focusing on the zebrafish orthologs, Seta and Setb. Seta/b proteins share high homology with their human counterpart. In situ hybridization revealed similar expression patterns for seta and setb genes. Both transcripts distributed at the midbrain-hindbrain boundary, the retinal pigment epithelium, the ganglion cell layer and the ciliary marginal zone, as well as, the sensory hair cells of the otic vesicle and the lateral line neuromasts. Similar results were obtained after whole mount immunofluorescence experiments. Taken together, these data provide a first link between the expression profiles of these genes and the development of the sensory organs of zebrafish. Loss-of-function studies using antisense morpholino oligonucleotides targeting both seta /b genes (MOab), resulted in increased apoptosis, reduced cell proliferation and morphological defects. The morphant phenotypes were partially rescued when MOab was co-injected with the human SET mRNA. Similar phenotypic defects were observed upon knockdown of setb with a transcription-blocking morpholino (MOb) or by CRISPR/Cas9. Moreover, in vivo labeling of hair cells using the fluorescent dye FM1-43, showed a decreased number of active neuromasts in MOab-, MOb- and gRNA/Cas9-injected embryos. The effects of the pharmacological inhibition of Seta/b with the sphingosine analog FTY720 were in accordance with the previous data, further supporting the functional correlation of Seta/b with these mechanoreceptive organs. In line with this, increased apoptosis was also detected in the hair cells of the neuromasts and otic vesicle upon seta/b knockdown or drug-mediated inhibition. Microarray analysis of the MOab transcriptome revealed differential expression in networks controlling gene transcription in the sensory organs. Collectively, the present work provides new insights on the physiological roles of seta and setb genes and shows for the first time that their functions are essential for embryonic and sensory system development.
Institution and School/Department of submitter: Πανεπιστήμιο Ιωαννίνων. Σχολή Επιστημών Υγείας. Τμήμα Ιατρικής
Subject classification: Πρωτεΐνη SET
Keywords: Πρωτεΐνη SET,Μοντέλο-οργανισμός,SET protein,Μodel-organism,Ζebrafish
Appears in Collections:Διδακτορικές Διατριβές

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