To further validate the IL6-induced phosphorylation of these Tyr residues in ACAP4 proteins, we performed in vitro kinase assays again using purified His-Trx-ACAP4415-903WT (ACAP4415-903WT), the nonphosphorylatable mutants His-Trx-ACAP4415-903-Y628F (ACAP4Y628F), His-Trx-ACAP4415-903-Y632F (ACAP4Y632F) and His-Trx-ACAP4415-903-Y843F (ACAP4Y843F) and identified the Tyr843 residue as the major phosphorylation site of ACAP4 induced by IL6 in HCC cells (Figure 4E). This evidence concerns the gene ASAP3 and hepatocellular carcinoma.