Integrin α3β1 Represses Reelin Expression in Breast Cancer Cells to Promote Invasion

Abstract

Integrin α3β1, a cell adhesion receptor for certain laminins, is known to promote breast tumor growth and invasion. Our previous gene microarray study showed that the RELN gene, which encodes the extracellular glycoprotein Reelin, was upregulated in α3β1-deficient (i.e., α3 knockdown) MDA-MB-231 cells. In breast cancer, reduced RELN expression is associated with increased invasion and poor prognosis. In this study we demonstrate that α3β1 represses RELN expression to enhance breast cancer cell invasion. RELN mRNA was significantly increased upon RNAi-mediated α3 knockdown in two triple-negative breast cancer cell lines, MDA-MB-231 and SUM159. Modulation of baseline Reelin levels altered invasive potential, where enhanced Reelin expression in MDA-MB-231 cells reduced invasion, while RNAi-mediated suppression of Reelin in SUM159 cells increased invasion. Moreover, treatment of α3β1-expressing MDA-MB-231 cells with culture medium that was conditioned by α3 knockdown MDA-MB-231 cells led to decreased invasion. RNAi-mediated suppression of Reelin in α3 knockdown MDA-MB-231 cells mitigated this effect of conditioned-medium, identifying secreted Reelin as an inhibitor of cell invasion. These results demonstrate a novel role for α3β1 in repressing Reelin in breast cancer cells to promote invasion, supporting this integrin as a potential therapeutic target.

Comparative use of CRISPR and RNAi to modulate integrin α3β1 in triple negative breast cancer cells reveals that some pro-invasive/pro-metastatic α3β1 functions are independent of global regulation of the transcriptome

“…Previously, we showed that RNAi-mediated suppression of ITGA3 in MDA-MB-231 cells substantially reduced cell invasion in vitro and metastatic lung colonization in vivo, and we have shown that these phenotypes are due at least partly to the loss of α3β1-dependent gene regulation in these cells [11][12][13]. However, α3β1 also has important roles in cell adhesion and migration, raising the possibility that this integrin also promotes invasion and/or metastasis independently of its ability to regulate gene expression.…”

“…To build from our previous investigations of integrin α3β1 in promoting pro-tumorigenic functions of TNBC cells [11][12][13], we used CRISPR to target the ITGA3 gene (which encodes the α3 integrin subunit) within the MDA-MB-231 cell line. As α3 is known to partner exclusively with the β1 integrin subunit [1], this strategy was employed to generate cells in which expression of α3β1 is completely ablated (i.e., α3-Cr cells).…”

“…One of the most common genetic approaches to identify functions of a specific integrin is use of RNA interference (RNAi) to suppress the expression of either the α or β subunit, resulting in reduced cell surface expression of the αβ heterodimer [8][9][10][11][12][13][14]. RNAi is widely used to suppress or "knock down" the expression of a target gene through transient expression of a small interfering RNA (siRNA), or through stable expression of a short hairpin RNA (shRNA), that is designed to neutralize the target mRNA transcript and inhibit gene expression or translation [15].…”

“…The laminin-binding integrin α3β1 promotes tumor growth, invasion, and metastasis of breast cancer cells, although this role may be context dependent [27][28][29]. Studies using RNAi to knockdown the α3 integrin subunit in breast cancer cells have identified α3β1-dependent gene regulation that promotes tumor cell growth and invasion [11][12][13]. In the current study, we used CRISPR to target the ITGA3 gene that encodes the α3 integrin subunit in MDA-MB-231 cells, a widely used model of triple-negative breast cancer (TNBC), thereby generating a variant line in which expression of α3β1 is entirely absent (hereafter referred to as α3-Cr cells).…”

Evidence of Reelin Signaling in GBM and Its Derived Cancer Stem Cells

“…Interestingly, a number of studies has reported changes in the expression of reelin in different cancer types even outside the nervous system [37]. Reelin expression has been found reduced in breast [38][39][40], colorectal [41,42], and pancreatic cancers [43], while it has been found to be increased in retinoblastoma [44], myelomas [45], and prostate cancers [39,46,47]. Here, we investigated the expression of reelin in human tissue samples obtained from tumoral lesion and the peritumoral area of GBM.…”