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The role of Na⁺/H⁺ exchanger isoform 5 (NHE5) in glioma

University of British Columbia

Up-regulation of the receptor tyrosine kinases (RTKs) signalling cascades constitutes the basis of tumorigenesis. Signal termination that normally occurs through receptor degradation is disrupted in various cancers. Therefore, persistent receptor-mediated signal occurs through increased recycling and elevated surface expression of these receptors. Previous discoveries of endosomal pH modulators that promote tumour progression suggest the potential roles of acidic endosomal pH in receptor turnover from the endosomes and oncogenic signalling. In this thesis study, I showed that neuron-enriched Na⁺/H⁺ exchanger NHE5 plays an important role in trafficking of c-Met RTK in C6 glioma cells. NHE5 is predominantly expressed in recycling endosomes of C6 cells. By fluorescence ratiometric analysis, I showed that NHE5 depletion with short-hairpin RNA significantly increases pH of Tfn positive recycling endosomes, implicating a prominent role of NHE5 in endosomal acidification. Using cell surface biotinylation, I found that cell surface abundance of hepatocyte growth factor (HGF) receptor c-Met is significantly reduced in NHE5-deficient cells. Further analysis revealed that recycling circuitry is impaired and degradation of c-Met is enhanced by a previously uncharacterized, non-proteasomal degradation when NHE5 is depleted. Consequently, reduced expression of c-Met by NHE5 knockdown (KD) causes severe migratory defect and loss of cell polarity. I further demonstrated that disrupted phenotypes in NHE5-knockdown cells were restored by NHE5 complementation. Stable expression of shRNA-resistant human NHE5 into NHE5-knockdown cells acidifies endosomal pH, increases surface c-Met abundance, restores Rac1 activity, and enhances cell migration. In summary, this thesis study highlights critical role of endosomes pH in regulating receptor-mediated signalling through vesicular trafficking, and a potential role of NHE5 in promoting sustained signalling essential for the tumorigenicity of glioma.

Text

2015-11-27

Attribution-NonCommercial-ShareAlike 2.5 Canada

http://hdl.handle.net/2429/55550

Biochemistry and Molecular Biology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-sa/2.5/ca/