Suggesting MFN1 plays a role in the cellular anti-viral response. PRA1 Echinacoside family protein 2 is an integral membrane protein with four-transmembrane domains that is localized to the ER and trans-Golgi network and is believed to play a regulatory role in vesicular trafficking. PRAF2 is also known to interact with C-C chemokine receptor type 5 and Bcl-xL to modulate cell survival. Ubiquitinconjugating enzyme E2 S functions as an auxiliary factor in the multi-subunit anaphase-promoting complex, a cell cycle-regulated E3 ubiquitin ligase that controls progression through mitosis. A number of viruses are now known to modulate the activity of the APC, including through the targeting of APC subunits for degradation, to facilitate their replication. Hypoxia up-regulated protein 1 is an inducible ER chaperone protein that is upregulated in response to cellular stresses including the unfolded protein response and hypoxia. HYOU1 is known to be cytoprotective and is involved in antiapoptotic signaling mechanisms. Initially the amounts of the seven proteins were analyzed in the SILAC labeled cytoplasmic and nuclear fractions used for the MS/MS analysis. In agreement with the MS/MS analysis, there was an observable decrease in the amounts of the proteins CTSL1, ERC1, KPNA2, MFN1, PRAF2 and UBE2S in the cytoplasmic fraction from DENV-2 infected cells whereas there was an increase in HYOU1. The most dramatic decreases were observed for the proteins ERC1 and PRAF2 that correlated with the quantitative MS/MS analysis. MS/MS analysis of the nuclear fractions from DENV-2 and mock infected cells resulted in the quantification of peptides corresponding to KPNA2 and HYOU1, that decreased and increased in infected cells respectively, which was confirmed by Western blot analysis. Although peptides corresponding to CTSL1, PRAF2 and UBE2S were identified in the tryptic digests from the nuclear fraction, they were not used for quantification. However all of these proteins were detected in the nuclear fractions by Western blot analysis at higher levels than suggested by the SILAC-MS analysis. Their abundance in the nuclear faction of DENV-2 infected cells, compared to mock infected cells reflected the changes in protein amounts observed in the cytoplasmic fraction. The levels of GAPDH were examined in the cellular extracts as a loading control as the MS/MS analysis showed that the level of GAPDH was essentially the same in the cytoplasm and nucleus of DENV-2 and mock infected cells. Following the validation of the proteomic analysis for the selected proteins, using the SILAC samples used for MS/MS analysis, it was of interest to examine whether the amounts of the proteins were consistently altered over a time course of DENV-2 infection and in another cell type. The amounts of the selected proteins were therefore examined in human A549 and HEK-293 cells at a number of time points after infection with DENV-2, in comparison to mock infected cells. Cell samples were harvested at 24, 48, 72, and 96 hours p.i. and used to Isoacteoside produce total cell lysates which were examined by Western blot analysis. To ensure that the respective cell lines were infected with DENV-2, the amount of the DENV-2 NS5 protein in the samples was examined, confirming the infections.