Abstract
Adhesion of basal keratinocytes to the underlying extracellular matrix (ECM) plays a key role in the control of skin homeostasis and response to injury. Integrin receptors indirectly link the ECM to the cell cytoskeleton through large protein complexes called focal adhesions (FA). FA also function as intracellular biochemical signaling platforms to enable cells to respond to changing extracellular cues. The α4β1 and α9β1 integrins are both expressed in basal keratinocytes, share some common ECM ligands, and have been shown to promote wound healing in vitro and in vivo. However, their roles in maintaining epidermal homeostasis and relative contributions to pathological processes in the skin remain unclear. We found that α4β1 and α9β1 occupied distinct regions in monolayers of a basal keratinocyte cell line (NEB-1). During collective cell migration (CCM), α4 and α9 integrins co-localized along the leading edge. Pharmacological inhibition of α4β1 and α9β1 integrins increased keratinocyte proliferation and induced a dramatic change in cytoskeletal remodeling and FA rearrangement, detrimentally affecting CCM. Further analysis revealed that α4β1/α9β1 integrins suppress extracellular signal-regulated kinase (ERK1/2) activity to control migration through the regulation of downstream kinases including Mitogen and Stress Activated Kinase 1 (MSK1). This work demonstrates the roles of α4β1 and α9β1 in regulating migration in response to damage cues.
Highlights
The epidermis functions as a barrier to protect against chemical, biological, and physical insults
We further demonstrate that α4 and α9 integrins share common intracellular binding partners that are required to suppress local activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and Mitogen and Stress activated kinase 1 (MSK1) in keratinocyte monolayers
We hypothesized that α4 and α9 integrins may regulate shared signaling pathways to elicit these effects. α4/α9 integrins have previously been implicated in restricting ERK1/2 activity in keratinocytes (Modica et al, 2017; Danussi et al, 2011), and ERK1/2 can modulate cell motility by directly phosphorylating and activating several components of the actin cytoskeleton machinery (Tanimura and Takeda 2017). In agreement with these data, we found that short term inhibition of α4 and α9 integrins increased ERK1/2 phosphorylation compared to MBOP and our vehicle control (DMSO) (Figures 3A,B)
Summary
The epidermis functions as a barrier to protect against chemical, biological, and physical insults. Basal keratinocytes are the only layer of the epidermis in contact with the extracellular matrix (ECM) at the dermal-epidermal junction and must undergo a phenotypic switch to enable rapid collective cell migration (CCM) to reinstate the skin barrier following injury (Haeger et al, 2015). Studies of migrating mesenchymal cells demonstrate that both α4 and α9 integrins regulate lamellipodia actin polymerization and maintain front-rear polarity, both of which are important during CCM (Hight-Warburton and Parsons 2019) These data suggest that α4 and α9 integrins may regulate processes associated with the migratory phase of skin repair following injury. We further demonstrate that α4 and α9 integrins share common intracellular binding partners that are required to suppress local activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and Mitogen and Stress activated kinase 1 (MSK1) in keratinocyte monolayers This signaling leads to coordinated actin protrusion and efficient directed migration. Our data provide new insight into α4 and α9 integrin functions in epithelial homeostasis and repair
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