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. 2016 Jun 24;90(14):6598–6610. doi: 10.1128/JVI.00722-16

FIG 4.

FIG 4

Effect of SUMO on MxA localization, expression, and oligomerization. (A) The immunofluorescence of MxA in HeLa-wt, HeLa-SUMO1, and HeLa-SUMO3 cells using rabbit anti-MxA antibodies is shown. (B) One hundred micrograms of protein extracts from HeLa-wt, HeLa-SUMO1 (HeLa-S1), and HeLa-SUMO3 (HeLa-S3) cells was analyzed by Western blotting using a 4 to 12% gradient gel, and the proteins were revealed with rabbit anti-MxA antibodies. (C) Total RNA was extracted from HeLa-wt, HeLa-SUMO1, and HeLa-SUMO3 cells, and mRNAs encoding MxA and RPL13A were quantified by RT-qPCR. Means and standard deviations from three independent experiments are shown. The Student t test was performed to determine the P value. ns, not significant. (D) SUMO enhances MxA oligomerization. Extracts from NIH 3T3-wt, NIH 3T3-MxA, HeLa-wt, HeLa-SUMO1, and HeLa SUMO3 cells were lysed, and proteins were cross-linked with DSS and immunoprecipitated (IP) using MxA antibodies. The eluates were subsequently analyzed by Western blotting using a 4 to 12% gradient gel, and proteins were revealed with anti-MxA, anti-SUMO1, anti-SUMO2/3, or anti-Ubc9 antibodies. The rabbit anti-MxA antibodies used recognize endogenous mouse Mx1 as well as human MxA.