Abstract
E-cadherin plays a powerful tumor suppressor role. Germline E-cadherin mutations justify 30% of Hereditary Diffuse Gastric Cancer (HDGC) and missense mutations are found in 30% of these families. We found possible to restore in vitro mutant E-cadherin associated to HDGC syndrome by using Chemical Chaperones (CCs). Herein, our aim was to disclose the molecular mechanisms underlying the CCs effects in E-cadherin regulation. Using cells stably expressing WT E-cadherin or two HDGC-associated missense mutations, we show that upon DMSO treatment, not only mutant E-cadherin is restored and stabilized at the plasma membrane (PM), but also Arf6 and PIPKIγ expressions are altered. We show that modulation of Arf6 expression partially mimics the effect of CCs, suggesting that the cellular effects observed upon CCs treatment are mediated by Arf6. Further, we show that E-cadherin expression recovery is specifically linked to Arf6 due to its role on endocytosis and recycling pathways. Finally, we demonstrated that, as DMSO, several others CCs are able to modulate the trafficking machinery through an Arf6 dependent mechanism. Interestingly, the more effective compounds in E-cadherin recovery to PM are those that simultaneously inhibit Arf6 and stimulate PIPKIγ expression and binding to E-cadherin. Here, we present the first evidence of a direct influence of CCs in cellular trafficking machinery and we show that this effect is of crucial importance in the context of juxtamembrane E-cadherin missense mutations associated to HDGC. We propose that this influence should be taken into account when exploring the therapeutic potential of this type of chemicals in genetic diseases associated to protein-misfolding.
Highlights
Cadherins are a family of adhesion molecules that play pivotal roles in tissue patterning during development and tissue architecture in adult
In this study we confirmed that E-cadherin expression is strongly increased upon dimethyl sulfoxide (DMSO) treatment – 2.6 fold increase in the wild type (WT) context, from 0.8 to 2.0 (2.5 fold) in the mutant R749W and from 0.4 to 1.5 (3.75 fold) in the mutant E757K, Figure 1A – and we verified that DMSO treatment leads to decreased ADP-ribosylation factor 6 (Arf6) expression (Figure 1A–C), both at the protein and RNA level
The above referred results could suggest that Ecadherin recovery by Chemical Chaperones (CCs) is mediated by an endocytosis decline
Summary
Cadherins are a family of adhesion molecules that play pivotal roles in tissue patterning during development and tissue architecture in adult. The extracellular domain of E-cadherin binds Ca2+ and forms complexes with the extracellular domains of other E-cadherin molecules on neighboring cells [1,2,3]. The cytoplasmic domain of E-cadherin interacts with the actin cytoskeleton through a complex of anchoring proteins including a-, b-, p120- and c-catenins [1,4]. E-cadherin plays a powerful tumor suppressor role and the assembly and maintenance of cadherin-catenins interactions is tightly regulated [2,5]. E-cadherin expression is partially or completely loss in various types of cancer [6,7] and this is associated to increased cell invasion and metastatic potential [8,9]. In sporadic diffuse gastric cancer and lobular breast cancer, E-cadherin loss is associated with somatic mutations, loss of heterozygosity, promoter hypermethylation, aberrant glycosylation and overexpression of transcriptional repressors [10,11,12,13,14,15], but these mechanisms explain only a rather limited percentage of cases with loss of E-cadherin
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