Sodium hydrogen exchanger and phospholipase D are required for alpha1-adrenergic receptor stimulation of metalloproteinase-9 and cellular invasion in CCL39 fibroblasts.
Matrix metalloproteinase 9 (MMP-9) plays a critical role in digesting the extracellular matrix and has a vital function in tumor metastasis and invasion; this protease activity is significantly increased in non-small cell lung cancers. The sodium hydrogen exchanger isoform 1 (NHE1) functions as a focal point for signal coordination and cytoskeletal reorganization. NHE1 is thought to play a central role in establishing signaling components at the leading edge of a migrating cell. Therefore, we studied the relationship between NHE1 and MMP-9 activity in Chinese hamster lung fibroblasts (CCL39) stimulated with phenylephrine (PE). We show that PE increases MMP-9 gelatinolytic activity in CCL39 cells. The inhibition of phospholipase D (PLD) signaling abrogated PE-induced MMP-9 activity. The role of PLD as an essential signaling intermediate was confirmed when the addition of permeable phosphatidic acid increased MMP-9 activity in the same cells. PE-induced invasion was increased 1.9-fold over controls and the PE response was lost when 1-butanol was used to block PLD signaling. Cells pre-treated with the NHE1 inhibitor, 5-(N-ethyl-N-isopropyl) amiloride (EIPA) prior to PE addition resulted in a notable decrease in MMP-9 activation and cell invasion as compared to untreated PE-stimulated cells. CCL39 NHE1 null cells demonstrated no increase in MMP-9 protease activity or cell invasion in response to PE treatment. Reconstitution of NHE1 expression recovered the PE-induced activation of protease activity and cell invasion. MMP-9 processing was altered in cells expressing a proton transport defective NHE1 but retained the ability to respond to PE. Conversely, cells expressing an ezrin, radixin, moesin (ERM)-binding deficient NHE1 had a lower MMP-9 activity and the protease did not respond to PE addition. Parallel studies on NCI-H358 non-small cell lung cancer (NSCL) cells showed that PE stimulated both MMP-9 activity and cell invasion in an NHE1 dependent manner. This work describes for the first time a PE-induced relationship between NHE1 and MMP-9 and a new potential mechanism by which NHE1 could promote tumor formation and metastasis.
Departments of Chemistry and Biosciences, Minnesota State University Moorhead, Hagen Hall, Moorhead, MN 56563, USA.
Jennifer
J
Taves
Danielle
D
Rastedt
Jenny
J
Canine
Dave
D
Mork
Mark A
MA
Wallert
Joseph J
JJ
Provost
D058646:Adrenergic alpha-1 Receptor Agonists; D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002289:Carcinoma, Non-Small-Cell Lung / Q000201:enzymology / Q000473:pathology*; D002460:Cell Line; D045744:Cell Line, Tumor; D006224:Cricetinae; D003412:Cricetulus; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D020404:Glycerophospholipids / Q000096:biosynthesis; D006801:Humans; D008175:Lung Neoplasms / Q000201:enzymology / Q000473:pathology*; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D009361:Neoplasm Invasiveness; D010656:Phenylephrine / Q000494:pharmacology; D010739:Phospholipase D / Q000037:antagonists & inhibitors / Q000378:metabolism*; D018340:Receptors, Adrenergic, alpha-1 / Q000502:physiology*; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
D058646:Adrenergic alpha-1 Receptor Agonists; D000316:Adrenergic alpha-Agonists; D020404:Glycerophospholipids; D018340:Receptors, Adrenergic, alpha-1; D017923:Sodium-Hydrogen Exchangers; C069139:growth factor-activatable Na-H exchanger NHE-1; C068564:phosphatidylbutanol; D010656:Phenylephrine; D010739:Phospholipase D; D020780:Matrix Metalloproteinase 9