Abstract
Expression of the small GTPase, Ras-related C3 botulinum toxin substrate 1B (RAC1B), a RAC1-related member of the Rho GTPase family, in tumor tissues of pancreatic ductal adenocarcinoma (PDAC) has been shown previously to correlate positively with patient survival, but the underlying mechanism(s) and the target genes involved have remained elusive. Screening of a panel of established PDAC-derived cell lines by immunoblotting indicated that both RAC1B and Mothers against decapentaplegic homolog 3 (SMAD3) were more abundantly expressed in poorly metastatic and well-differentiated lines as opposed to highly metastatic, poorly differentiated ones. Both siRNA-mediated RAC1B knockdown in the transforming growth factor (TGF)-β-sensitive PDAC-derived cell lines, Panc1 and PaCa3, or CRISPR/Cas-mediated knockout of exon 3b of RAC1 in Panc1 cells resulted in a dramatic decrease in the expression of SMAD3. Unexpectedly, the knockdown of SMAD3 reproduced the promigratory activity of a RAC1B knockdown in Panc1 and PaCa3, but not in TGF-β-resistant BxPC3 and Capan1 cells, while forced expression of SMAD3 alone was able to mimic the antimigratory effect of ectopic RAC1B overexpression in Panc1 cells. Moreover, overexpression of SMAD3 was able to rescue Panc1 cells from the RAC1B knockdown-induced increase in cell migration, while knockdown of SMAD3 prevented the RAC1B overexpression-induced decrease in cell migration. Using pharmacological and dominant-negative inhibition of SMAD3 C-terminal phosphorylation, we further show that the migration-inhibiting effect of SMAD3 is independent of its activation by TGF-β. Finally, we provide evidence that the antimigratory program of RAC1B-SMAD3 in Panc1 cells is executed through upregulation of the migration and TGF-β inhibitor, biglycan (BGN). Together, our data suggest that a RAC1B-SMAD3-BGN axis negatively controls cell migration and that SMAD3 can induce antimigratory genes, i.e., BGN independent of its role as a signal transducer for TGF-β. Therefore, targeting this novel pathway for activation is a potential therapeutic strategy in highly metastatic PDAC to interfere with invasion and metastasis.
Highlights
Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive tumor with an extremely poor prognosis
Using experimental strategies of related C3 botulinum toxin substrate 1B (RAC1B) knockdown (KD) or knockout (KO) in pancreatic ductal adenocarcinoma (PDAC)-derived Panc1 and breast cancer-derived MDA-MB-231 cells, we have recently shown that the inhibition of transforming growth factor (TGF)-β1-dependent chemokinesis by RAC1B was a consequence of downregulation of the TGF-β type I receptor, activin receptor-like kinase (ALK)5 [9]
We focused our attention on the role of RAC1B in the regulation of basal migratory/chemokinetic activity in PDAC, employing pancreatic carcinoma cells of different metastatic potential
Summary
Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive tumor with an extremely poor prognosis. RAC1B differs from RAC1 by an in-frame insertion of an additional exon (exon 3b) of 57 nucleotides. This results in profound alterations in its biochemical and signaling properties as well as in its cellular effects, some of which are antagonistic to those of RAC1 [3]. RAC1B has been reported to promote cellular proliferation and antiapoptosis, but unlike RAC1 its role in other cancer-related processes, such as epithelial–mesenchymal transition (EMT) and cell migration and invasion remains poorly characterized. RAC1 is known to promote EMT, migration and invasion [4,5] and the same role in EMT induced by matrix metalloprotease (MMP) has been postulated for RAC1B (reviewed in [3])
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
