Expression and Activation of Signal Regulatory Protein α on Astrocytomas

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Abstract
High-grade astrocytomas and glioblastomas are usually unresectable because they extensively invade surrounding brain tissue. Here, we report the expression and function of a receptor on many astrocytomas that may alter both the proliferative and invasive potential of these tumors. Signal regulatory protein (SIRP) α1 is an immunoglobulin superfamily transmembrane glycoprotein that is normally expressed in subsets of myeloid and neuronal cells. Transfection of many cell types with SIRPα1, including glioblastomas, has been shown to inhibit their proliferation in response to a range of growth factors. Furthermore, the expression of a murine SIRPα1 mutant has been shown to enhance cell adhesion and initial cell spreading but to inhibit cell extension and movement. The extracellular portion of SIRPα1 binds CD47 (integrin-associated protein), although this interaction is not required for integrin-mediated activation of SIRPα1. On phosphorylation, SIRPα1 recruits the tyrosine phosphatases SHP-1 and SHP-2, which are important in its functions. Although SHP-1 is uniquely expressed on hematopoietic cells, SHP-2 is ubiquitously expressed, so that SIRPα1 has the potential to function in many cell types, including astrocytomas. Because SIRPα1 regulates cell functions that may contribute to the malignancy of these tumors, we examined the expression of SIRPs in astrocytoma cell lines by flow cytometry using a monoclonal antibody against all SIRPs. Screening of nine cell lines revealed clear cell surface expression of SIRPs on five cell lines, whereas Northern blotting for SIRPα transcripts showed mRNA present in eight of nine cell lines. All nine cell lines expressed the ligand for SIRPα1, CD47. To further examine the expression and function of SIRPs, we studied the SF126 and U373MG astrocytoma cell lines, both of which express SIRPs, in greater detail. SIRP transcripts in these cells are identical in sequence to SIRPα1. The expressed deglycosylated protein is the same size as SIRPα1, but in the astrocytoma cells, it is underglycosylated compared with SIRPα1 produced in transfected Chinese hamster ovary cells. It is nonetheless still capable of binding soluble CD47. Moreover, SIRPα1 in each of the two cell lines recruited SHP-2 on phosphorylation, and SIRPα1 phosphorylation in cultured cells is CD47 dependent. Finally, examination of frozen sections from 10 primary brain tumor biopsies by immunohistochemistry revealed expression of SIRPs on seven of the specimens, some of which expressed high levels of SIRPs. Most of the tumors also expressed CD47. This is the first demonstration that astrocytomas can express SIRPα. Given the known role of SIRPα in regulating cell adhesion and responses to mitogenic growth factors, the expression of SIRPα1 on astrocytomas may be of considerable importance in brain tumor biology, and it offers the potential of a new avenue for therapeutic intervention.