Research Information System
IVEK-3rd International Convention of Pharmaceuticals and Pharmacies, İstanbul, Turkey, 26 - 29 April 2017, pp.121, (Summary Text)
Influenza A viruses are contagious and causative of recurrent pandemics in humans. These viruses are enveloped
viruses classified as Orthomyxoviridae family. The genome of influenza A virus is composed of eight single-
stranded, negative sense RNA molecules. Nuclear export protein (NEP or NS2), which is the subject of this
work, is encoded from the eighth segment of the viral genome with alternative splicing. The function of NEP is
to mediate the nuclear export of viral ribonucleoprotein complexes (vRNPs) by acting as an adaptor between
vRNPs and cellular nuclear pore complex (NPC) related proteins. In this work, it was aimed to reveal the relationship
between NEP and NPC related proteins, NUPL2 and NUP214 at molecular level.
NUPL2 and NUP214 proteins were determined as candidate proteins with yeast two-hybrid (Y2H) assay. For
the Y2H assay, influenza A virus NEP protein was used as a bait and assayed for interaction with the proteins
encoded by cDNAs of HEK293 cells. The genes coding NUPL2 and NUP214 proteins that showed interaction
with the NEP in yeast cells and the NEP gene were cloned in the mammalian expression vectors, and were used
in the research carried out with mammalian cells. The expression and cellular localization of these proteins in
HEK293 or HeLa cells were detected with SDS-PAGE / Western blotting and immunofluorescence assays
(IFA).
Western blotting analysis showed that viral NEP and host proteins NUPL2 and NUP214 were efficiently expressed
in HEK293 cells. HeLa cells were used for detection of cellular localization of these proteins with IFA.
Influenza virus NEP tagged with flag proteins showed dominantly nuclear localization. Carboxyl-terminal half
of NUP214 protein tagged with HA was localized in nucleus of HeLa cells when expressed alone. NUPL2 protein
tagged with HA showed nuclear or cytoplasmic localization about half. The relationship of viral NEP and
host proteins was investigated in co-tranfected HeLa cell with IFA. It was shown that co-expression of NEP/
NUPL2 and NEP/NUP214 interfere with the cellular localization of each other and shows co-localization in
HeLa cells. These results strongly confirmed the functional relationship between viral NEP and host NUPL2
and NUP214 proteins not only in yeast cells, but also in the mammalian cells. Thus, the data obtained in this
study suggested that at least two of the NPC-related proteins, NUPL2 and NUP214 have a relationship with
NEP to transport of vRNPs into the cytoplasm of host cells, which is a vital function for the viral maturation in
the cells. In this regard, NEP and NPC related proteins can be considered as targets for the developing of antiviral
drugs.