Fig. 3

Differentially present proteins in the CM of AGR2-expressing CMT cells. A, B Heatmap of the proteins identified as AGR2-increased (A) and AGR2-decreased (B) in the CM of CMT-U27 or CF41.Mg. Each CM sample was triply analyzed (Rep1, Rep2, Rep3). Protein abundance was determined by normalizing PSMs of specific proteins to total PSMs in replicates. Cumulative normalized PSM ratios of AGR2-expressing CM were divided by vector-expressing CM. This ratio underwent logarithmic transformation (Log2) as Log2 (A/V). Proteins with Log2 (A/V) above mean + 1.5 SD were classified as AGR2-increased; below mean-1.5 SD were designated as AGR2-decreased. AGR2-increased and AGR2-decreased proteins which are in common in both CMT-U27 CM and CF41.Mg CM are highlighted in red (A) and blue (B), respectively, and are shown in the overlap of Venn diagrams (C). D, E Verification of increased 14-3-3ε and α-actinin 4 levels in the CM of AGR2-expressing CMT cells. CMT-U27 (D) or CF41.Mg (E) cells transfected with the AGR2-expressing or the mock vector were grown in serum-free CM for 24 h, and the CM samples were collected and concentrated. The concentrated CM and whole-cell lysate (WCL) were analyzed by immunoblotting with indicated specific antibodies. Levels of 14-3-3ε and α-actinin 4 in the AGR2-expressing group were presented as fold changes to that in the vector group. Calnexin was used as a negative control in the CM