Year: 2011
Type: dissertation
Abstract: The human ether-à-go-go potassium channel (hEag1) can be detected in adult brain and in both, tumor cell lines and primary tumors. The physiological function of this channel in the brain still remains to be elucidated. The presence of hEag1 in tumors has been linked to an increase in proliferation. Published reports claimed that the IGR39 melanoma cell line is negative for hEag1, what allowed this cell to be used as negative control. We observed that IGR39 cells shows positive amplification for hEag1, at mRNA level, and a protein with compatible molecular weight and reactivity for hEag1 antibody, was detected in western-blot experiments. The biophysical properties of the current registered in IGR39 presents a different kinetics, but it is still sensitive to blocking by drugs and siRNA specific for hEag1. Using molecular biology techniques, we found the expression of two new splice variants, E65 and E70, in two melanoma cell lines (IPC298 and IGR39) and in neural stem cells. The two splice variants have no transmembrane domains and can not form a functional membrane-bound channel. A characterization of the new splice variants was done in Xenopus leavis oocytes, to verify the interaction of the splice variants with the full-length hEag1 (fl-hEag1). Electrophysiological measurements of the oocytes, co-injected with the splice variant E65 and the fl-hEag1, showed a rectified current with diminished amplitude. Moreover, the co-injection of E65 and fl-hEag1 enables these oocytes to mature, which was the same outcome of the treatment of oocytes with progesterone. Based in these results, we conclude that IGR39 cell line express Eag1 channels and the splice variants found in these melanoma cells, could be either interacting with the full-length Eag1 or playing a role in the intracellular signaling.
DOI: 10.53846/goediss-369
Author: Gomes, Fernanda Ramos
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
OpenAlex ID: W4212847511