Elastin Peptides Activate Extracellular Signal-Regulated Kinase 1/2 via a Ras-Independent Mechanism Requiring Both p110γ/Raf-1 and Protein Kinase A/B-Raf Signaling in Human Skin Fibroblasts

Abstract

Elastin peptides (EPs) produced during cancer progression bind to the elastin binding protein (EBP) found at the surface of dermal fibroblasts, leading to the expression of collagenase-1 gene. The production of this enzyme involved in stromal reaction is caused by the sustained activation of the extracellular signal-regulated kinases 1/2 (ERK1/2) pathway via cAMP/protein kinase A (PKA) and phosphatidylinositol 3-kinase (PI3K). However, the mechanism of these signaling events remains unknown. We show that -elastin (E), a commonly used EP, induces maximum phosphorylation of mitogen-activated protein kinase/extracellular signalregulated kinase (MEK)1/2 and ERK1/2 after 30 min. The simultaneous inhibition of PKA and PI3K, by N-(2-(p-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide (H89) and 2-(4-morpholynil)-8-phenyl-4H-1-bemzopyran-4-one (LY294002), respectively, blocked MEK1/2 and ERK1/2 phosphorylation, as did lactose, an EBP antagonist. E induced Raf-1 phosphorylation and activation in a PI3K-dependent manner. In our system, the PI3K p110␥ is expressed and activated by ␤␥-derived subunits from a pertussis toxin-sensitive G protein after fibroblast stimulation. Pertussis toxin also blocks the Raf-1/MEK1/2/ERK1/2 phosphorylation cascade. In addition, we found that B-Raf is expressed in dermal fibroblasts and activated in a PKA-dependent manner after E treatment, thereby integrating PKA signals to MEK1/2. It is noteworthy that Ras involvement was excluded because ERK1/2 activation by E was not blocked in RasN17-transfected fibroblasts. Together, our results identify a novel Rasindependent ERK1/2 activation system in which p110␥/Raf-1/ MEK1/2 and PKA/B-Raf/MEK1/2 cooperate to activate ERK1/2. Thus, p110␥ and B-Raf seem to be important modulators of dermal fibroblasts physiology and should now qualify as therapeutic targets in strategies aiming at limiting elastin degradation contribution to cancer progression.

Elastin Receptor (Spliced Galactosidase) Occupancy by Elastin Peptides Counteracts Proinflammatory Cytokine Expression in Lipopolysaccharide-Stimulated Human Monocytes through NF-κB Down-Regulation

“…Up to now, EPs were only involved in up-regulation of signaling cascade and gene expression (28,55,56). Thus, biological effects exerted by EPs should take into account the cell type used (37,57), the signaling pathway analyzed (31,56), and, as delineated in this study, the presence or not of coeffectors. As a matter of fact, most of the effects of EPs have been previously investigated using resting cells.…”

“…For instance, activation of protein kinase G, cAMP, or cGMP by EPs was reported to induce chemotaxis of monocytes (28 -30). Besides, activation of pertussis toxin-sensitive G proteins and MEK/ERK signaling cascade were involved in EP-mediated effects on cell proliferation as observed in arterial smooth muscle cells and fibroblasts (31,32). In turn, S-gal occupancy by EPs led to the activation of both NF-B and p38 in melanoma cells (33).…”

Elastin-Derived Peptides Induce Inflammatory Responses through the Activation of NF-κB in Human Ligamentum Flavum Cells

“…In human dermal fibroblasts, EDPs upregulate collagenase-1 expression through PKA, PI3K, and ERK1/2 pathway activation [24]. They further demonstrated that κE activates ERK1/2 via a Ras-independent mechanism involving p110γ/Raf-1/MEK1/2 and PKA/B-Raf/ MEK1/2 cascades [10]. Our study demonstrated that EDPs activated ERK and NF-κB signaling pathways in human LF cells ( Figure 5).…”

“…In elastin-rich tissues such as artery, lung, and skin, inflammation is concomitant with elastolysis, leading to EDPs generation [10,13,14]. In those pathologies, a direct relationship between inflammation and EDP levels has been established.…”

“…This conception has evolved and changed since EDPs have been demonstrated as potent and specific modulators of cellular functions in a number of normal and transformed cells [10,11], but complete knowledge of their distinct effects in LF cells has not been reported.…”