TGF-β-Induced Endothelial to Mesenchymal Transition Is Determined by a Balance Between SNAIL and ID Factors

Abstract

Endothelial-to-mesenchymal transition (EndMT) plays an important role in embryonic development and disease progression. Yet, how different members of the transforming growth factor-β (TGF-β) family regulate EndMT is not well understood. In the current study, we report that TGF-β2, but not bone morphogenetic protein (BMP)9, triggers EndMT in murine endothelial MS-1 and 2H11 cells. TGF-β2 strongly upregulates the transcription factor SNAIL, and the depletion of Snail is sufficient to abrogate TGF-β2-triggered mesenchymal-like cell morphology acquisition and EndMT-related molecular changes. Although SLUG is not regulated by TGF-β2, knocking out Slug also partly inhibits TGF-β2-induced EndMT in 2H11 cells. Interestingly, in addition to SNAIL and SLUG, BMP9 stimulates inhibitor of DNA binding (ID) proteins. The suppression of Id1, Id2, or Id3 expression facilitated BMP9 in inducing EndMT and, in contrast, ectopic expression of ID1, ID2, or ID3 abrogated TGF-β2-mediated EndMT. Altogether, our results show that SNAIL is critical and indispensable for TGF-β2-mediated EndMT. Although SLUG is also involved in the EndMT process, it plays less of a crucial role in it. In contrast, ID proteins are essential for maintaining endothelial traits and repressing the function of SNAIL and SLUG during the EndMT process. These data suggest that the control over endothelial vs. mesenchymal cell states is determined, at least in part, by a balance between the expression of SNAIL/SLUG and ID proteins.

TGF-β-mediated Endothelial to Mesenchymal Transition (EndMT) and the Functional Assessment of EndMT Effectors using CRISPR/Cas9 Gene Editing

“…In addition, the TGF-β2-mediated decline in Pecam-1 and enhancement of Sm22α were completely blocked in Snail-depleted MS-1 cells. In summary, we demonstrated that Snail is critical for TGF-β2-mediated EndMT in MS-1 cells (Figure 3B) 15 .…”

“…To demonstrate the function of Snail in TGF-β2-mediated EndMT, we performed an EndMT assay in Snail-depleted cells and compared it with parental MS-1 cells. As shown in Figure 3A, the knockout of Snail was sufficient to inhibit the fibroblast-like cell morphology driven by TGF-β2 in MS-1 cells 15 . In addition, the TGF-β2-mediated decline in Pecam-1 and enhancement of Sm22α were completely blocked in Snail-depleted MS-1 cells.…”

“…TGF-β is one of the cytokines with greatest potential to induce EndMT. After treating MS-1 cells with TGF-β2 (1 ng/mL) for 3 days, endothelial MS-1 cells lose their cobblestone-like structure and differentiate into spindle-shaped mesenchymallike cells (Figure 1A) 15 Snail protein expression was abrogated (Figure 2D) 15 .…”

“…In this study, we used murine pancreatic microvascular endothelial cells (MS-1) 14 as a model system to examine the TGF-β2-induced EndMT process. Our previous study demonstrated that Snail is the main transcription factor increased by TGF-β2, by which EndMT is induced in MS-1 cells 15 . Upon CRISPR/Cas9 gene editing to abrogate Snail expression in MS-1 cells, TGF-β2 failed to mediate EndMT.…”

Endothelial Heterogeneity in Development and Wound Healing

“…FGF signalling and inhibitor of DNA proteins (Id proteins) are among the most notable pathways countering TGFβ-driven EndMT [ 113 , 114 , 115 ]. FGF2 increases miR20 expression which targets ALK5 and TGFβRII that transduce TGFβ signalling thereby inhibiting EndMT [ 116 ].…”