Transforming Growth Factor-β: Activation by Neuraminidase and Role in Highly Pathogenic H5N1 Influenza Pathogenesis

Abstract

Transforming growth factor-beta (TGF-β), a multifunctional cytokine regulating several immunologic processes, is expressed by virtually all cells as a biologically inactive molecule termed latent TGF-β (LTGF-β). We have previously shown that TGF-β activity increases during influenza virus infection in mice and suggested that the neuraminidase (NA) protein mediates this activation. In the current study, we determined the mechanism of activation of LTGF-β by NA from the influenza virus A/Gray Teal/Australia/2/1979 by mobility shift and enzyme inhibition assays. We also investigated whether exogenous TGF-β administered via a replication-deficient adenovirus vector provides protection from H5N1 influenza pathogenesis and whether depletion of TGF-β during virus infection increases morbidity in mice. We found that both the influenza and bacterial NA activate LTGF-β by removing sialic acid motifs from LTGF-β, each NA being specific for the sialic acid linkages cleaved. Further, NA likely activates LTGF-β primarily via its enzymatic activity, but proteases might also play a role in this process. Several influenza A virus subtypes (H1N1, H1N2, H3N2, H5N9, H6N1, and H7N3) except the highly pathogenic H5N1 strains activated LTGF-β in vitro and in vivo. Addition of exogenous TGF-β to H5N1 influenza virus–infected mice delayed mortality and reduced viral titers whereas neutralization of TGF-β during H5N1 and pandemic 2009 H1N1 infection increased morbidity. Together, these data show that microbe-associated NAs can directly activate LTGF-β and that TGF-β plays a pivotal role protecting the host from influenza pathogenesis.

IL-10 inhibits neuraminidase-activated TGF-β and facilitates Th1 phenotype during early phase of infection

“…The release of active TGF-b in 6.5 HA-specific CD4 þ T cells increased with augmented viral NA activity (top panels, Fig. 1c), similar to previous reports on TGF-b activation by influenza NA 10,27 . Viral NA also desialylated and exposed LAP on the membrane, and activated TGF-b in the HA-specific CD4 þ T cells in the draining lymph nodes of the lung (mediastinal lymph nodes, MLN; Supplementary Fig.…”

“…27). NA removes sialic acids on the LAP and the enzymatic cleavage causes the release of active TGF-b from its inactive precursor form 10 . TGF-b activation suppresses T-cell activation and implements invasion from the defensive immunity of the host.…”

“…TGF-b is a critical factor in our model of influenza virus infection and is related to the intensity of inflammation and the disease severity. It has also been reported that differential TGF-b activation by different NAs of various strains of influenza viruses is also critical for disease manifestations 10 . Increased TGF-b activity is correlated to better survival.…”

“…Viral NA increases with multiplication of virus during the early phase of infection. NA is known to cause activation of latent TGF-b in the lungs 10 . As most of the cell types possess latent TGF-b, we tested whether viral NA activates TGF-b in 6.5 HA antigen-specific CD4 þ T cells.…”

“…This cleavage facilitates viral entry and exit from the cells 9 . It also activates latent transforming growth factor (TGF)-b by removing the sialic acids on the N-terminal latency-associated peptide (LAP) 10 . Active TGF-b mediates immune suppression, including regulation of T cells [11][12][13][14] , and contributes to evasion by the pathogen from the immune response [15][16][17][18] .…”