In the present work I investigated the role of two closely related proteins Nkx5-1 and Nkx5-2 in neuronal apoptosis and differentiation. These two proteins show high sequence conservation in several vertebrate species. It was postulated, that they play overlapping roles in the inner ear and nervous system development. Nkx5-1 and Nkx5-2 are expressed during inner ear development as well as during adult stages in the mouse. In addition to the inner ear structures, they are also expressed in post- mitotic neurons in several central and peripheral locations. Nkx5-1 knockout leads to severe defects of the vestibular apparatus of the inner ear. However, singular Nkx5-1 gene knockout mice did not reveal any obvious neuronal phenotype. Based on the fact that double Nkx5-1/2 (also called Hmx2/3) knockout led to a severe postnatal lethal phenotype, redundant functions for both Nkx5 genes were postulated. In the double knockout mice defects in some hypothalamic functions were documented, however, no neuronal loss was observed in the functionally affected regions and the molecular basis of Nxk5 genes action remains unresolved.To investigate molecular mechanisms of Nkx5-1 and Nkx5-2 genes functions in neuronal cells, PC12 rat pheochromocytoma cells were used as an experimental model. This cell line is a commonly used system for the investigation of the neurogenesis and undergoes neuronal differentiation upon treatment with nerve growth factor NGF. Recently, it was shown that BMP family members, BMP4 and BMP6, support NGF-mediated neuronal differentiation of PC12 cells. In contrast to such coordinated action of BMP and NGF signalling, another BMP- family member, BMP2, is able to stimulate neurite outgrowth in PC12 without NGF.Interestingly, a multifunctional cellular regulator protein p53 was also recently demonstrated to be another player within the NGF-differentiation pathway: p53 knockdown inhibited NGF-induced differentiation. The high affinity NGF receptor TrkA was revealed as a direct target for p53-mediated transcriptional regulation. Depending on the cellular context p53 may regulate TrkA either to induce cell cycle arrest and differentiation or apoptosis.To study the potential role of Nkx5-1 in the adult neuronal structures I used Nkx5-1 knockout mouse strain generated previously in our laboratory.
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