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PMID: 1319241 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Spreading of HeLa cells on a collagen substratum requires a second messenger formed by the lipoxygenase metabolism of arachidonic acid released by collagen receptor clustering.

Molecular biology of the cell ·Vol. 3 ·No. 5 ·1992-05-00 ·Pages 481-92

Chun JS, Jacobson BS

Abstract

HeLa cells attach to a variety of substrata but spread only on collagen or gelatin. Spreading is dependent on collagen-receptor upregulation, clustering, and binding to the cytoskeleton. This study examines whether second messengers are involved in initiating the spreading process on gelatin. The levels of cytosolic free calcium ([Ca++]i), cAMP, and cytoplasmic pH (pHi) do not change during cell attachment and spreading. However, a basal level of [Ca++]i and an alkaline pH(i) are required for spreading. There is an activation of protein kinase C (PKC) and a release of arachidonic acid (AA) on attachment and before cell spreading. Inhibition of PKC does not block cell spreading, indicating that PKC activation is not essential for spreading. Inhibition of phospholipase A2 blocks cell spreading, whereas addition of exogeneous AA overcomes this inhibitory effect. Among AA metabolic pathways, inhibitors of lipoxygenase (LOX) block cell spreading, suggesting that a LOX product(s) formed from AA initiates spreading. Clustering receptors for collagen with polyclonal antibodies, or with anti-collagen-receptor antigen-binding fragments (Fab) in combination with a secondary antibody, induce AA release. Also, AA is released when cells attach to either immobilized gelatin or immobilized Arg-Gly-Asp (RGD) peptide. Thus, AA is released whenever receptor clustering is observed. Receptor occupancy is not sufficient to release AA; when cells are treated with gelatin or RGD peptide in solution or anti-collagen-receptor Fab fragments without secondary antibody, conditions where receptor clustering is not observed, AA is not released. Thus, a LOX metabolite(s) of AA formed by collagen-receptor clustering is a second messenger(s) that initiates HeLa cell spreading. LOX inhibitors also block the spreading of bovine aortic endothelial cells, chicken embryo fibroblasts, and CV-1 fibroblasts on gelatin or fibronectin, indicating that other cells might use the same second messenger system in initiating cell-substratum adhesion.

MeSH Terms
Amino Acid Sequence Arachidonic Acid/metabolism Calcium/physiology Cells, Cultured Collagen/metabolism Cyclic AMP/physiology HeLa Cells/enzymology,metabolism Humans Hydrogen-Ion Concentration Lipoxygenase/metabolism Molecular Sequence Data Phospholipases A/metabolism Phospholipases A2 Protein Kinase C/physiology Receptors, Cell Surface/physiology Receptors, Collagen Second Messenger Systems/physiology
Chemicals
Receptors, Cell Surface Receptors, Collagen Arachidonic Acid Collagen Cyclic AMP Lipoxygenase Protein Kinase C Phospholipases A Phospholipases A2 Calcium
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Chun J S
Department of Biochemistry and Molecular Biology, University of Massachusetts, Amherst 01003.
Jacobson B S
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Article Info
Journal
Molecular biology of the cell
Abbr.
Mol Biol Cell
ISSN
1059-1524
DOI
10.1091/mbc.3.5.481
Published
1992-05-00
Pages
481-92
Language
English
Region
United States
NLM ID
9201390
PMCID
PMC275602
Subset
IM
Grants
NIGMS NIH HHS · GM-29127 · United States
External Links
PubMed source DOI: 10.1091/mbc.3.5.481 PMC Full Text View journal details
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