Dengue virus non-structural protein 1 disrupts the TGF-β/Smad signaling

Abstract TGF-β signaling is tightly regulated to ensure cellular functions. Role of DENV on the TGF-β/Smad signaling has not been well established. Therefore, we aimed to study the association between DENV infection and TGF-β/Smad signaling. We observed significant impairment in the expression of Smad2, Smad3, Smad4, Smad6 and Smad7 during DENV replication, which are the key players in TGF-β signaling. Significant reduction in the expression of phosphorylated Smad3 was also documented. Overexpression of Smad2/3/4/6 provided the evidence of slight inhibition on DENV replication indicating these Smads may work against the establishment of DENV replication. DENV non-structural protein 1 (NS1) was noted as crucial viral factor that impaired the expression of Smad2, Smad3 and Smad4 and also physically interacts with these proteins as confirmed by co-immunoprecipitation assay. Additionally, we observed NS1 is also capable of blocking the nuclear translocation of Smad3 and thus further ensuring inhibition of Smad signaling. To figure out degradation mechanisms, we studied the role of two distinct E3 ligases, CHIP and Smurf2, which are essential for the degradation of Smad proteins. Co-expression of Smad2/3/4 and NS1 with Smurf2, Smurf2mut, CHIP or use of CHIP-/- cells suggests that only Smurf2 has significant role in the degradation of Smad proteins during DENV infection. NS1 may acts as a co-factor with Smurf2 to escalate the proteasome and lysosome mediated degradation of Smad3 and Smad4 proteins respectively. Therefore, our results confirm that NS1 interacts with Smad proteins and reduces their expression by utilizing E3 ligase and disrupt the TGF-β/Smad signaling.