טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentKolpakova Alina
SubjectThe Role of Mef2D Transcription Factor in Germ Layer
Formation and Patterning at Early Development
DepartmentDepartment of Medicine
Supervisor Professor Eyal Bengal
Full Thesis textFull thesis text - English Version


Abstract

In Xenopus, the specification of the three germ layers is between the earliest developmental decisions occurring prior to gastrulation. At the first developmental stages zygotic transcription is repressed and maternally-expressed factors provide blueprint for embryonic development.

Myocyte Enhancer Factor 2 (Mef2) A and D are maternally expressed. At present, most of the studies regarding Mef2 were focused on relatively late developmental stages. We focused our research on the roles of Mef2D at early development.

In this work we have demonstrated that Xenopus Mef2 isoforms A and D are animally localized before the Mid Blastula Transition (MBT) and their expression expands marginally and vegetally after the beginning of zygotic transcription.

Mef2 activity was downregulated by means of antisense morpholino and expression of dominant-negative Mef2-engrailed chimera protein. Knockdown analysis revealed that Mef2D is involved in Spemann organizer formation and mesoderm induction. Moreover, Chromatin Immunoprecipitation analysis indicated that Mef2D protein was associated with promoter regions of Xnr1 and Brachyury. Thus, our results indicate that Mef2 proteins directly regulate the expression of pro-mesodermal genes.

Animal Cap (AC) explants from Mef2-depleted embryos failed to respond to organizer signals and to express neuronal markers. The contacts between cells of appeared loosen and the expression of genes encoding for ectodermal proteins was downregulated. Animal cells from Mef2-depleted embryos failed to re-associate after dissociation. Ectopic expression of Mef2D at the vegetal hemisphere induced the expression of pro-ectodermal genes Ectodermin and FoxI1e. Taken together, these results suggest that Mef2D is important for ectodermal formation and specification.

Overall, we show that marginally-expressed Mef2 proteins regulate the expression of mesoderm and organizer genes, while animally-expressed Mef2 proteins induce the expression of pro-ectodermal genes affecting cell adhesion properties and competence of these cells to respond to organizer neurolizing signals.