SHP-1 Phosphatase C-Terminus Interacts With Novel Substrates p32/p30 During Erythropoietin and Interleukin-3 Mitogenic Responses

Abstract

SHP-1 protein tyrosine phosphatase is a critical negative regulator of mitogenic signaling, as demonstrated by the heightened growth responses to hematopoietic growth factors in hematopoietic cells of motheaten mice, which lack functional SHP-1 expression due to mutations in the SHP-1 gene. The mitogenic signaling molecules dephosphorylated by SHP-1 have not been fully identified. We detected two proteins (p32/p30) that are hyperphosphorylated in a DA3/erythropoietin receptor (EpoR) cell line that expresses a mutant containing the SHP-1 C-terminus that suppresses the function of the endogenous phosphatase and induces hyperproliferative responses to interleukin-3 (IL-3) and Epo. Hyperphosphorylated p32/p30 are also detected in motheaten hematopoietic cells, demonstrating an association of p32/p30 hyperphosphorylation with SHP-1-deficiency and growth factor-hyperresponsiveness. The hyperphosphorylated p32/30 associate with SHP-1 via its C-terminus, because they coimmunoprecipitate with the phosphatase and the C-terminal mutant and they bind in vitro to a synthetic peptide of the mutant but not the GST fusion proteins of SHP-1 SH2 domains. Induction of p32/p30 phosphorylation by IL-3 or Epo occurs mainly at 2 to 18 hours poststimulation in the DA3/EpoR cell line, indicating p32/p30 as novel signaling molecules during cell cycle progression. These data demonstrate a function for the SHP-1 C-terminus in recruiting potential substrates p32/p30 and suggest that SHP-1 may regulates mitogenic signaling by dephosphorylating p32/p30.

Reduction of Hematopoietic Cell-Specific Tyrosine Phosphatase SHP-1 Gene Expression in Natural Killer Cell Lymphoma and Various Types of Lymphomas/Leukemias

“…The consensus sequence (S/L/I/V)XYXX(L/V), based on sequences originally deduced from several receptors known to bind to the C-terminal SH2 domain of SHP1, defines all immunoreceptor tyrosine-based inhibitory motifs (ITIMs), including NK cell, B cell, and monocyte and dendritic cell inhibitory receptors. [21][22][23][24] SHP1 is known to associate with multiple signaling molecules, including ZAP70, 25 CD3 ⑀, 25,26 CD5 26 and interleukin-2R 27 in T cells; interleukin-3 receptor ␤ chain 28,29 and erythropoietin receptor 30 in hematopoietic cells; CD22, 31,32 B cell receptor, 33 SLP76 34,35 and CD72 36,37 in B cells; and the killer cell inhibitory receptor 38,39 in natural killer cells. These interactions appear to exert primarily inhibitory effects on their signaling cascades.…”