Neuritogenesis and the nerve growth factor-induced differentiation of PC-12 cells requires annexin II-mediated plasmin generation.
One of the key morphological changes associated with the nerve growth factor (NGF)-induced differentiation of rat adrenal pheochromocytoma (PC-12) cells is the growth of axon-like processes called neurites. A growing body of evidence suggests that this process may be dependent upon plasmin, a serine protease generated from plasminogen (Plg) by either urokinase Plg activator (u-PA) or tissue Plg activator (t-PA). Prior work in our laboratory has identified annexin II (Ann-II) as a co-receptor for Plg and t-PA that promotes and localizes plasmin generation near the cell surface. In the present study, we report a 3-9-fold increase in Ann-II protein and message levels in NGF-treated PC-12 cells. Message stability and nuclear run-on assays suggest that this induction occurs at the level of gene transcription. Neurite outgrowth assays on and within a three-dimensional matrix demonstrate the inhibition of NGF-induced PC-12 cell differentiation by polyclonal and monoclonal antibodies directed against Ann-II as well as by the overexpression of antisense Ann-II mRNA. Neuritogenesis is also impaired by alpha(2)-plasmin inhibitor, antibodies directed against t-PA and u-PA, and epsilon-aminocaproic acid, a lysine analog that inhibits Plg activation and the binding of Plg to Ann-II. Plasmin generation assays reveal a 2-fold increase in plasmin production on NGF-treated PC-12 cells, which can be blocked by a polyclonal antibody directed against the tail region of Ann-II. From these data, we conclude that Ann-II is transcriptionally up-regulated by NGF and that Ann-II-mediated plasmin generation may play an important role during neurite development in the differentiating PC-12 cell.
Jacovina
A T
AT
Division of Hematology-Oncology, Department of Pediatrics, Weill Medical College of Cornell University, New York, New York 10021, USA.
Zhong
F
F
Khazanova
E
E
Lev
E
E
Deora
A B
AB
Hajjar
K A
KA
D015119:Aminocaproic Acid / Q000494:pharmacology; D000818:Animals; D017306:Annexin A2 / Q000235:genetics / Q000378:metabolism*; D000906:Antibodies; D002454:Cell Differentiation; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D007150:Immunohistochemistry; D020932:Nerve Growth Factor / Q000494:pharmacology*; D016501:Neurites / Q000187:drug effects / Q000502:physiology*; D009474:Neurons / Q000166:cytology*; D016716:PC12 Cells; D010455:Peptides / Q000276:immunology / Q000378:metabolism; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000378:metabolism; D051381:Rats; D010959:Tissue Plasminogen Activator / Q000276:immunology; D015854:Up-Regulation / Q000502:physiology; D014568:Urokinase-Type Plasminogen Activator / Q000276:immunology
D017306:Annexin A2; D000906:Antibodies; D010455:Peptides; D020932:Nerve Growth Factor; D010959:Tissue Plasminogen Activator; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras); D015119:Aminocaproic Acid