A Deletion Mutation in the SH2-N Domain of Shp-2 Severely Suppresses Hematopoietic Cell Development

Abstract

Shp-1 and Shp-2 are cytoplasmic protein tyrosine phosphatases that contain two Src homology 2 (SH2) domains. A negative regulatory role of Shp-1 in hematopoiesis has been strongly implicated by the phenotype of motheaten mice with a mutation in the Shp-1 locus, which is characterized by leukocyte hypersensitivity, deregulated mast cell function, and excessive erythropoiesis. A targeted deletion of 65 amino acids in the N-terminal SH2 (SH2-N) domain of Shp-2 leads to an embryonic lethality at midgestation in homozygous mutant mice. To further dissect the Shp-2 function in hematopoietic development, we have isolated homozygous Shp-2 mutant embryonic stem (ES) cells. Significantly reduced hematopoietic activity was observed when the mutant ES cells were allowed to differentiate into embryoid bodies (EBs), compared to the wild-type and heterozygous ES cells. Further analysis of ES cell differentiation in vitro showed that mutation in the Shp-2 locus severely suppressed the development of primitive and definitive erythroid progenitors and completely blocked the production of progenitor cells for granulocytes-macrophages and mast cells. Reverse transcriptase PCR analysis of the mutant EBs revealed reduced expression of several specific marker genes that are induced during blood cell differentiation. Stem cell factor induction of mitogen-activated protein kinase activity was also blocked in Shp-2 mutant cells. Taken together, these results indicate that Shp-2 is an essential component and primarily plays a positive role in signaling pathways that mediate hematopoiesis in mammals. Furthermore, stimulation of its catalytic activity is not sufficient, while interaction via the SH2 domains with the targets or regulators is necessary for its biological functions in cells. The in vitro ES cell differentiation assay can be used as a biological tool in dissecting cytoplasmic signaling pathways.

A negative role of SHP-2 tyrosine phosphatase in growth factor-dependent hematopoietic cell survival

“…SHP-2 appears to play compound roles in hematopoietic cell processes. Although SHP-2 targeted deletion mutation severely suppresses hematopoietic development (Qu et al, 1997, this might be mainly due to loss of its promoting role in early differentiation process of ES cells Chan et al, 2003). Therefore, it has been unclear how SHP-2 phosphatase is exactly involved in hematopoietic stem/progenitor cell processes.…”

“…Therefore, it has been unclear how SHP-2 phosphatase is exactly involved in hematopoietic stem/progenitor cell processes. Since SHP-2 knockout embryos do not survive past midgestation (Qu et al, 1997;Saxton et al, 1997), the number of SHP-2 mutant hematopoietic cells available is extremely limited. Dominant-negative inhibition and overexpression approaches are very useful for addressing certain aspects of SHP-2 function in the hematopoietic compartment.…”

“…We previously mutated the SHP-2 gene in mouse embryonic stem (ES) cells. Analyses of in vitro (Qu et al, 1997) and in vivo hematopoietic cell differentiation from SHP-2 À/À ES cells demonstrated that SHP-2 was required for primitive hematopoiesis. These results stand in contrast to the negative role of the closely related SHP-1 phosphatase in hematopoietic cell signaling.…”

The tyrosine phosphatase SHP‐2 regulates differentiation and apoptosis of individual primary T lymphocytes

“…However, there is evidence for a role of SHP-2 in distinct signaling cascades in hematopoietic cells from in vitro differentiation of SHP-2-deficient ES cells and the generation of chimeric mice. These experiments showed that SHP-2 is involved in the development and differentiation of all lineages of hematopoietic cells, such as erythroid progenitors, myeloid and lymphoid cells [9][10][11]. SHP-2 has also been implicated in the signaling of cytokine receptors that signal via the common c-chain, the shared b-chain and the receptor subunit gp130 such as IL-2, IL-3 or IL-6 [12][13][14][15][16].…”

Catalytic-dependent and -independent roles of SHP-2 tyrosine phosphatase in interleukin-3 signaling

“…We previously mutated the SHP-2 gene in mouse embryonic stem (ES) cells. Analyses of in vitro (Qu et al, 1997) and in vivo (Qu et al, 1998) hematopoietic cell differentiation from SHP-2 À/À ES cells demonstrated that SHP-2 was required for primitive hematopoiesis. These results stand in sharp contrast to the negative role of the closely related SHP-1 phosphatase in hematopoietic cell signaling.…”