Taurodeoxycholate Increases the Number of Myeloid-Derived Suppressor Cells That Ameliorate Sepsis in Mice.

Bile acids (BAs) control metabolism and inflammation by interacting with several receptors. Here, we report that intravenous infusion of taurodeoxycholate (TDCA) decreases serum pro-inflammatory cytokines, normalizes hypotension, protects against renal injury, and prolongs mouse survival during sepsis. TDCA increases the number of granulocytic myeloid-derived suppressor cells (MDSCLT) distinctive from MDSCs obtained without TDCA treatment (MDSCL) in the spleen of septic mice. FACS-sorted MDSCLT cells suppress T-cell proliferation and confer protection against sepsis when adoptively transferred better than MDSCL. Proteogenomic analysis indicated that TDCA controls chromatin silencing, alternative splicing, and translation of the immune proteome of MDSCLT, which increases the expression of anti-inflammatory molecules such as oncostatin, lactoferrin and CD244. TDCA also decreases the expression of pro-inflammatory molecules such as neutrophil elastase. These findings suggest that TDCA globally edits the proteome to increase the number of MDSCLT cells and affect their immune-regulatory functions to resolve systemic inflammation during sepsis.

Twist1 and AP-1 cooperatively upregulate integrin α5 expression to induce invasion and the epithelial-mesenchymal transition.

Epithelial-mesenchymal transition (EMT) is an important process implicated in tumor invasion and metastasis. Twist1 is a transcription factor that induces EMT, including E-cadherin suppression and cancer cell migration and invasion; hence it promotes cancer metastasis. Twist1 directly or indirectly regulates the expression of various genes and cellular functions involved in cancer progression. However, the underlying mechanisms remain largely unknown. In this study, we investigated the molecular basis for Twist1-mediated invasion and EMT. In human cancer cells, Twist1 was found to directly upregulate transcription of the mesenchymal gene integrin α5 in an E-box-independent, but activating protein-1 (AP-1) element-dependent, manner. Twist1 activated the integrin α5 promoter by interacting with and activating the transcription factor AP-1, composed of c-Jun and activating transcription factor-2 (ATF-2); it also enhanced the nuclear presence of ATF-2. AP-1 was critical for Twist1-induced cancer cell invasion, primarily through the induction of integrin α5, which activated c-Jun N-terminal kinase and focal adhesion kinase-signaling activities. Using data from The Cancer Genome Atlas, we found that Twist1 expression positively correlates with integrin α5 expression in human colorectal cancers. These findings suggest that cooperation between Twist1 and AP-1 represents a novel mechanism for EMT and tumor invasiveness. This study supports further investigation into the molecular basis underlying the diverse Twist1-mediated functions that occur during tumor progression.

ZEB2-Sp1 cooperation induces invasion by upregulating cadherin-11 and integrin α5 expression.

Epithelial-mesenchymal transition (EMT) is a process implicated in invasion and metastasis. EMT is characterized by repression of epithelial markers and induction of mesenchymal markers. ZEB2 is a transcriptional repressor of E-cadherin, leading to EMT. Previously, we have shown that ZEB2 directly upregulates integrin α5 transcription by cooperating with the transcription factor Sp1. In this study, we investigated the precise mechanism by which ZEB2 modulates invasion and EMT events and the role of Sp1 in ZEB2-induced invasion. We found that ZEB2 directly induced cadherin-11 transcription in an Sp1-dependent, but Smad- and E-box-independent, manner and repressed E-cadherin expression in an Sp1- and Smad-independent manner, leading to cadherin switch. Furthermore, ZEB2 upregulated Sp1 by enhancing Sp1 protein stability, and Sp1 was found to be critical for ZEB2-induced cancer cell invasion, mainly through induction of cadherin-11 and integrin α5. Expression levels of cadherin-11 and integrin α5 were interdependent and both modulated c-Jun N-terminal kinase-signaling activity and invasion. Immunofluorescence analysis showed that nuclear expression of ZEB2 was positively correlated with Sp1 expression in human colorectal cancers. Together, these findings demonstrate a previously unrecognized interplay between ZEB2, Sp1, cadherin-11 and integrin α5 that is, probably, significant in tumor progression and metastasis.

ZEB2 upregulates integrin α5 expression through cooperation with Sp1 to induce invasion during epithelial-mesenchymal transition of human cancer cells.

Epithelial-mesenchymal transition (EMT) is a process implicated in tumor invasion, metastasis, embryonic development and wound healing. ZEB2 is a transcription factor involved in EMT that represses E-cadherin transcription. Although E-cadherin downregulation is a major event during EMT and tumor progression, E-cadherin reduction is probably not sufficient for full invasiveness. The mechanisms by which E-cadherin transcriptional repressors induce mesenchymal genes during EMT remain largely unknown. Here, we investigated the role of ZEB2 in the induction of integrin α5 during cancer EMT and its underlying mechanism. In human cancer cells, ZEB2 was found to directly upregulate integrin α5 transcription in a manner that is independent of the regulation of E-cadherin expression. Conversely, depletion of ZEB2 by small interfering RNA suppressed integrin α5 expression, leading to reduced invasion. Suppression of integrin α5 inhibited cancer cell invasion, suggesting an important role for integrin α5 in cancer progression. Furthermore, ZEB2 was found to activate the integrin α5 and vimentin promoters by interacting with and activating the transcription factor Sp1, suggesting that cooperation between ZEB2 and Sp1 represents a novel mechanism of mesenchymal gene activation during EMT. These findings increase our understanding of the pathways beyond E-cadherin reduction that regulate mesenchymal gene expression during EMT and cancer progression.

TGF-beta1 induces mouse dendritic cells to express VEGF and its receptor (Flt-1) under hypoxic conditions.

Angiogenesis is a multi-step process that involves the activation, proliferation, and migration of endothelial cells. We have recently shown that TGF-beta1 can induce mouse macrophages to produce VEGF, a potent angiogenic factor. In the present study, we explored whether TGF-beta1 has a similar effect on mouse dendritic cells. First, we show that under hypoxic conditions, TGF-beta1 induced the expression of VEGF transcripts in bone marrow-derived dendritic cells. Overexpression of Smad3/4 further augmented TGF-beta1-induced VEGF transcription, while overexpression of DN-Smad3 decreased VEGF transcription in DC2.4 cells, a mouse dendritic cell line. We also show that TGF-beta1 and Smads are involved in the induction of VEGF protein secretion. Interestingly, under the same conditions, the expression of VEGF receptor 1 (Flt-1) was also elevated at both the transcriptional and protein levels. Additionally, we found that the TGF-beta1-induced VEGF secretion in activated DC2.4 cells has wound-healing properties. Finally, Smad7 and Smurf1 negatively regulated the TGF-beta1-induced and Smad3/4-mediated VEGF expression. Taken together, these results indicate that TGF-beta1 can enhance the expression of VEGF and Flt-1 through the typical Smad pathway in mouse dendritic cells.

TMPRSS4 induces invasion and epithelial-mesenchymal transition through upregulation of integrin alpha5 and its signaling pathways.

TMPRSS4 is a novel type II transmembrane serine protease that is highly expressed on the cell surface in pancreatic, thyroid and other cancer tissues, although its oncogenic significance and molecular mechanisms are unknown. Previously, we have shown that TMPRSS4 promotes invasion, migration and metastasis of human tumor cells by facilitating an epithelial-mesenchymal transition (EMT). In this study, we explored the molecular basis underlying TMPRSS4-mediated effects. We show that multiple downstream signaling pathways, including focal adhesion kinase (FAK), extracellular signal-regulated kinase (ERK), Akt, Src and Rac1, are activated by TMPRSS4 expression and that FAK signaling and ERK activation are required for TMPRSS4-induced invasiveness and EMT, including cadherin switch. Inhibition of PI3K or Src reduced invasiveness and actin rearrangement mediated by TMPRSS4 without restoring E-cadherin expression. Downregulation of E-cadherin was required for TMPRSS4-mediated effects but was not sufficient to induce EMT and invasion. TMPRSS4 induced integrin alpha5 expression and its signal transduction, leading to invasiveness and EMT accompanied by downregulation of E-cadherin. Functional blocking confirmed that integrin alpha5beta1 is a critical signaling molecule that is sufficient to induce TMPRSS4-mediated effects. Immunohistochemical analysis showed that TMPRSS4 expression was significantly higher in human colorectal cancer tissues from advanced stages than in that of early stage. Furthermore, upregulation of TMPRSS4 was correlated with enhanced integrin alpha5 expression. These observations implicate integrin alpha5 upregulation as a molecular mechanism by which TMPRSS4 induces invasion and contributes to cancer progression.

Tiul1 and TGIF are Involved in Downregulation of TGFbeta1-induced IgA Isotype Expression.

TGF-beta1 is well known to induce Ig germ-line alpha (GLalpha) transcription and subsequent IgA isotype class switching recombination (CSR). Homeodomain protein TG-interacting factor (TGIF) and E3-ubiquitin ligases TGIF interacting ubiquitin ligase 1 (Tiul1) are implicated in the negative regulation of TGF-beta signaling. In the present study, we investigated the roles of Tiul1 and TGIF in TGFbeta1-induced IgA CSR. We found that over-expression of Tiul1 decreased TGFbeta1-induced GLalpha promoter activity and strengthened the inhibitory effect of Smad7 on the promoter activity. Likewise, overexpression of TGIF also diminished GLalpha promoter activity and further strengthened the inhibitory effect of Tiul1, suggesting that Tiul1 and TGIF can down-regulate TGFbeta1-induced GLalpha expression. In parallel, overexpression of Tiul1 decreased the expression of endogenous IgA CSR-predicitive transcripts (GLT(alpha), PST(alpha), and CT(alpha)) and TGFbeta1-induced IgA secretion, but not GLT(gamma3) and IgG3 secretion. Here, over-expressed TGIF further strengthened the inhibitory effect of Tiul1. These results suggest that Tiul1 and TGIF act as negatively regulators in TGFbeta1-induced IgA isotype expression.

Opposing effects of Arkadia and Smurf on TGFbeta1-induced IgA isotype expression.

TGF-beta1 induces Ig germ-line alpha (GLalpha) transcription and subsequent class switching recombination (CSR) to IgA. In the present study, we investigated the roles of two E3-ubiquitin ligases, Smurfs (HECT type) and Arkadia (RING finger type) on TGFbeta1-induced IgA CSR. We found that over-expression of Smurf1 and Smurf2 decreased TGFbeta1-induced GLalpha promoter activity and strengthened the inhibitory effect of Smad7 on the promoter activity. Further, over-expression of Smurf1 and Smurf2 decreased both Smad3/4-mediated and Runx3-mediated GLalpha promoter activities, suggesting that the Smurfs can down-regulate the major TGF-beta1 signaling pathway and decrease GLalpha gene expression. In parallel, the over-expressed Smurf1 decreased the expression of endogenous IgA CSR-predictive transcripts (GLT(alpha), PST(alpha), and CT(alpha)) and also TGFbeta1-induced IgA secretion. Conversely over-expression of Arkadia abolished the inhibitory effect of Smad7 on TGFbeta1-induced GLT(alpha) expression and IgA secretion. Similar results were obtained in the presence of over-expressed Smad7 and Smurf1. These results indicate that Arkadia can amplify TGFbeta1-induced IgA CSR by degrading Smad7, which interacts with Smurf1. We conclude that Smurf and Arkadia have opposite roles in the regulation of TGFbeta1-induced IgA isotype expression.