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http://dx.doi.org/10.5762/KAIS.2017.18.12.575

Analysis of the Role of RGG box of human hnRNP A1 protein  

Choi, Mieyoung (Department of BT-Convergent Pharmaceutical Engineering, Sun Moon University)
Publication Information
Journal of the Korea Academia-Industrial cooperation Society / v.18, no.12, 2017 , pp. 575-580 More about this Journal
Abstract
This study analyzed the effects of RGG box of hnRNP A1 on its subcellular localization and stabilization of hnRNP A1 over a three year period from October 2014. First, a 6R/K mutation in RGG box was generated, and pcDNA1-HA-hnRNP A1(6R/K) was constructed. The subcellular localization of hnRNP A1(6R/K) from the HeLa cells transfected with this plasmid DNA was analyzed by immunofluorescence microscopy. HA-hnRNP A1(6R/K) was found to exhibit nuclear and cytoplasmic fluorescence. The stability of hnRNP A1(6R/K) was checked by Western blot analysis using the expressed protein from the HeLa cells transfected with the pcDNA1-HA-hnRNP A1(6R/K). The results show that HA-hnRNP A1(6R/K) has a smaller size. These confirm that HA-hnRNP A1(6R/K) is localized both in the nuclear and cytoplasm, not because 6R/K mutation affects the nuclear localization of hnRNP A1, but because 6R/K mutation causes hnRNP A1(6R/K) to cleave at the mutation or near the mutation site. The cleaved protein fragment, which lacks the M9 domain (i.e. nuclear localization signal of hnRNP A1), did not exhibit nuclear fluorescence. This suggests that the arginines of RGG box in hnRNP A1 play an important role in stabilizing hnRNP A1. An analysis of the RNA-binding ability of hnRNP A1(6R/K) expressed and purified from bacteria will be a subsequent research project.
Keywords
HA(hemagglutin) epitope; hnRNP A1; immunofluorescence microscopy; M9 domain; nuclear localization signal; RGG box;
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