A cell adhesive peptide from tropoelastin promotes sequential cell attachment and spreading via distinct receptors

Abstract

Tropoelastin is the dominant monomer that assembles to form elastin, which confers elasticity to vertebrate elastic tissues including skin, arteries, and lungs. Tropoelastin interacts with cells through cell surface receptors including integrins and glycosaminoglycans (GAGs). As the region 17-18 is recognized as a key region in cell attachment and spreading, we utilized C-terminal truncated tropoelastin constructs containing dissected sections of domain 18. We mapped a cell-interactive sequence of tropoelastin to domain 17 and the first six amino acids (aa) of domain 18. Further delineation identified a 21-residue sequence (Peptide 302-322) which promoted cell attachment and spreading indistinguishable from that to N18, a construct encompassing the full domains 17-18. Alanine substitution of the lysines at positions 11 and 14 in Peptide 302-322 effectively abolished cell binding. This reliance on lysines pointed to a role for GAGs, which was assessed by heparan sulfate inhibition, leading to 85.9 ± 4.2% decreased cell binding, while inhibition of integrins using ethylenediaminetetraacetic acid did not affect attachment. In contrast, selective antibody blocking of the integrin α family prevented cell spreading by 92.5 ± 8.9%. We propose a two-step mechanism by which cell interactions occur at this central region of tropoelastin: initially, cell adhesion is mediated by GAGs, which contact the lysine residues within the target sequence, and subsequently facilitate cell spreading modulated by integrins, specifically α β and α β . We conclude that this region comprises a tropoelastin-derived, cell-interactive sequence that independently mediates potent cell binding and spreading via sequential recognition of GAG and integrin cell surface receptors.

Soluble matrix protein is a potent modulator of mesenchymal stem cell performance

“…In contrast, we show that fibronectin in solution does not promote MSC proliferation, possibly due to poor cell recognition as its cell receptor binding sites become exposed only upon adsorption to a surface such as a collagen matrix (37). The effects of tropoelastin in solution are likely enabled by the inherent accessibility (35,38) of its cell binding regions (32,39,40). Before this work, soluble signaling factors derived from ECM proteins, including fibronectin, laminin, collagen, and elastin, were thought to be limited to peptides released by partial proteolysis, termed matrikines (41,42).…”

Tropoelastin is a Flexible Molecule that Retains its Canonical Shape

“…This sequence works synergistically with integrins, where GAG‐binding allows the engagement of the α v integrin family to induce cell spreading . Domains that contribute to integrin binding reside in regions adjacent to the GAG‐binding sequence spanning domains 17 and 18 . We propose that the flexibility of the central region may therefore facilitate cell–receptor interactions by exposing relevant domains.…”

“…Second, the variability in domain positioning may promote cellular interaction. Earlier work suggests that the central region of the molecule interacts with cell surface receptors including glycosaminoglycans (GAGs) and integrins, promoting cell binding and cell spreading . In particular, domains 17 and 18 contain a glycosaminoglycan‐binding site responsible for cell adhesion.…”

“…In particular, domains 17 and 18 contain a glycosaminoglycan‐binding site responsible for cell adhesion. This sequence works synergistically with integrins, where GAG‐binding allows the engagement of the α v integrin family to induce cell spreading . Domains that contribute to integrin binding reside in regions adjacent to the GAG‐binding sequence spanning domains 17 and 18 .…”

“…Spherule formation through coacervation begins at the cell surface by surface‐tethered tropoelastin molecules, until released for integration into the growing elastic fiber. Tropoelastin aggregates adhere to the cell surface through GAG‐ and integrin–mediated interactions at specific interaction sites . Spherules are deposited on a microfibrillar scaffold where they are stabilized by cross‐links facilitated by lysyl oxidase and eventually integrated into a mature elastic fiber.…”