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RPPA and Western blot confirmation of Smad protein phosphorylation.

figure

posted on 2018-02-06, 18:34 authored by Cynthia de la Fuente, Chelsea Pinkham, Deemah Dabbagh, Brett Beitzel, Aura Garrison, Gustavo Palacios, Kimberley Alex Hodge, Emanuel F. Petricoin, Connie Schmaljohn, Catherine E. Campbell, Aarthi Narayanan, Kylene Kehn-Hall

A) Phosphorylation status of nine RPPA targets after RVFV infection of HSAECs. Bar graph represents the mean fold change with standard deviation of three independent replicates. Phospho-specific Smad antibodies are noted as: pSmad1/5/9 MH2 [phosphorylated Smad1/5 (S463/465)/9 (S465/467)], pSmad1/5 MH2 [phosphorylated Smad1/5 (S463/465)], pSmad2 MH2 [phosphorylated Smad2 (S465/467)], and pSmad2 L [phosphorylated Smad2 (S245/250/255)]. Statistical significance of the phosphorylation changes are highlighted in Table 1. HSAEC (B) and Huh-7 cells (C) were infected at an MOI 5 with MP12 or ZH548, or mock infected (media alone). Protein lysates harvested at 3, 9, and 18hpi were analyzed for pSmad1/5/9 MH2, pSmad1/5 MH2, pSmad2 MH2, pSmad2 L, RVFV NP, Actin as well as total protein levels for Smad1, Smad5, and Smad2 by western blotting. Blots represent one replicate of three independent experiments. M:Mock; MP:MP12; ZH: ZH548.