SHP1 tyrosine phosphatase negatively regulates NPM-ALK tyrosine kinase signaling.
Anaplastic large-cell lymphoma (ALCL) is frequently associated with the 2;5 translocation and expresses the NPM-ALK fusion protein, which possesses a constitutive tyrosine kinase activity. We analyzed SHP1 tyrosine phosphatase expression and activity in 3 ALK-positive ALCL cell lines (Karpas 299, Cost, and SU-DHL1) and in lymph node biopsies (n = 40). We found an inverse correlation between the level of NPM-ALK phosphorylation and SHP1 phosphatase activity. Pull-down and coimmunoprecipitation experiments demonstrated a SHP1/NPM-ALK association. Furthermore, confocal microscopy performed on ALCL cell lines and biopsy specimens showed the colocalization of the 2 proteins in cytoplasmic bodies containing Y664-phosphorylated NPM-ALK. Dephosphorylation of NPM-ALK by SHP1 demonstrated that NPM-ALK was a SHP1 substrate. Downregulation of SHP1 expression by RNAi in Karpas cells led to hyperphosphorylation of NPM-ALK, STAT3 activation, and increase in cell proliferation. Furthermore, SHP1 overexpression in 3T3 fibroblasts stably expressing NPM-ALK led to the decrease of NPM-ALK phosphorylation, lower cell proliferation, and tumor progression in nude mice. These findings show that SHP1 is a negative regulator of NPM-ALK signaling. The use of tissue microarrays revealed that 50% of ALK-positive ALCLs were positive for SHP1. Our results suggest that SHP1 could be a critical enzyme in ALCL biology and a potential therapeutic target.
Centre de Physiopathologie de Toulouse-Purpan, CHU-Purpan, Place du Docteur Baylac, 31059 Toulouse Cedex, France.
Jean-François
JF
Honorat
Ashraf
A
Ragab
Laurence
L
Lamant
Georges
G
Delsol
Jeannie
J
Ragab-Thomas
D001483:Base Sequence; D002460:Cell Line; D003593:Cytoplasm / Q000502:physiology; D017931:DNA Primers; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D016403:Lymphoma, Large B-Cell, Diffuse / Q000201:enzymology* / Q000503:physiopathology; D018613:Microscopy, Confocal; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology
D017931:DNA Primers; D047908:Intracellular Signaling Peptides and Proteins; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; C107023:p80(NPM-ALK) protein; D011505:Protein-Tyrosine Kinases; D010744:Phosphoric Monoester Hydrolases; C509249:PTPN6 protein, human; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases