Surface protein CD20 serves as the critical target of immunotherapy in various B-cell malignancies for decades, however its biological function and regulation remain largely elusive. Better understanding of CD20 function may help to design improved rational therapies to prevent development of resistance. Using CRISPR/Cas9 technique, we have abrogated CD20 expression in five different malignant B-cell lines. We show that CD20 deletion has no effect upon B-cell receptor signaling or calcium flux. Also B-cell survival and proliferation is unaffected in the absence of CD20. On the contrary, we found a strong defect in actin cytoskeleton polymerization and, consequently, defective cell adhesion and migration in response to homeostatic chemokines SDF1α, CCL19 and CCL21. Mechanistically, we could identify a reduction in chemokine-triggered PYK2 activation, a calcium-activated signaling protein involved in activation of MAP kinases and cytoskeleton regulation. These cellular defects in consequence result in a severely disturbed homing of B cells in vivo.
- MeSH
- aktiny metabolismus MeSH
- antigeny CD20 genetika metabolismus fyziologie MeSH
- B-buněčný lymfom metabolismus patologie MeSH
- B-lymfocyty patologie fyziologie MeSH
- buněčná adheze fyziologie MeSH
- genový knockdown MeSH
- leukemie B-buněčná metabolismus patologie MeSH
- lidé MeSH
- multimerizace proteinu fyziologie MeSH
- myši inbrední NOD MeSH
- myši SCID MeSH
- myši transgenní MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- pohyb buněk fyziologie MeSH
- polymerizace MeSH
- receptory antigenů B-buněk metabolismus MeSH
- signální transdukce imunologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
OBJECTIVES: TWIST1 is a member of the class B of basic helix-loop-helix transcription factors that regulates cell lineage determination and differentiation and has been implicated in epithelial-to-mesenchymal transition. Here, we aimed to investigate the role of TWIST1 for the activation of resident fibroblasts in systemic sclerosis (SSc). METHODS: The expression of Twist1 in fibroblasts was modulated by forced overexpression or siRNA-mediated knockdown. Interaction of Twist1, E12 and inhibitor Of differentiation (Id) was analysed by co-immunoprecipitation. The role of Twist1 in vivo was evaluated using inducible, conditional knockout mice with either ubiquitous or fibroblast-specific depletion of Twist1. Mice were either challenged with bleomycin or overexpressing a constitutively active transforming growth factor (TGF)β receptor I. RESULT: The expression of TWIST1 was increased in fibroblasts in fibrotic human and murine skin in a TGFβ/SMAD3-dependent manner. TWIST1 in turn enhanced TGFβ-induced fibroblast activation in a p38-dependent manner. The stimulatory effects of TWIST1 on resident fibroblasts were mediated by TWIST1 homodimers. TGFβ promotes the formation of TWIST1 homodimers by upregulation of TWIST1 and by induction of inhibitor of DNA-binding proteins, which have high affinity for E12/E47 and compete against TWIST1 for E12/E47 binding. Mice with selective depletion of Twist1 in fibroblasts are protected from experimental skin fibrosis in different murine models to a comparable degree as mice with ubiquitous depletion of Twist1. CONCLUSIONS: Our data identify TWIST1 as a central pro-fibrotic factor in SSc, which facilitates fibroblast activation by amplifying TGFβ signalling. Targeting of TWIST1 may thus be a novel approach to normalise aberrant TGFβ signalling in SSc.
- MeSH
- fibroblasty účinky léků metabolismus MeSH
- genový knockdown MeSH
- jaderné proteiny biosyntéza nedostatek genetika metabolismus fyziologie MeSH
- kůže patologie MeSH
- lidé MeSH
- malá interferující RNA genetika MeSH
- messenger RNA genetika MeSH
- multimerizace proteinu fyziologie MeSH
- myši knockoutované MeSH
- regulace genové exprese účinky léků fyziologie MeSH
- signální transdukce fyziologie MeSH
- studie případů a kontrol MeSH
- systémová sklerodermie metabolismus patologie MeSH
- transformující růstový faktor beta farmakologie MeSH
- transkripční faktor Twist biosyntéza nedostatek genetika metabolismus fyziologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
