Fig 2: Shp-2 is largely dispensable for NK cell effector functions. a Graphs depict percentages of CD94+, Ly49A+, Ly49G2+, Ly49I+, Ly49D+, Ly49H+, and KLRG1+ splenic NK cells (gated as NK1.1+ CD3/CD19-) from Ncr1Ki Ptpn11wt/wt (white), Ncr1Ki Ptpn11fl/fl (dark gray), and B2m-/- (green) mice. b Graph and a representative cytometric plot illustrate the production of granzyme A and B by splenic NK cells (gated as NK1.1+NKp46+CD3/CD19-) after phorbol 12-myristate 13-acetate and ionomycin stimulation (PMA/Iono). c NK cells isolated from polyinosinic:polycytidylic acid (polyI:C)-treated Ncr1Ki Ptpn11wt/wt or Ncr1Ki Ptpn11fl/fl mice were plated with RMA, RMA-S, or RMA-H60 cells at the indicated ratios. The graph depicts percentage killing of target cells, as measured by quantifying PI- living target cells. d Naive splenocytes from Ncr1Tg Ptpn11fl/fl (light gray) mice or heterozygote littermate controls (white) were incubated with RMA cells or RMA-m157 cells (expression of m157 is illustrated in the graph on the left). Percentage of YFP+Ly49H+CD107a+ NK cells in each group was determined by flow cytometry (illustrated in the graph on the right). a, b Results represent the mean ± SEM of n = 4 (Ncr1Ki Ptpn11fl/fl or B2m-/-) and n = 6 (Ncr1Ki Ptpn11wt/wt) mice per genotype and are representative of at least three (a) and two (b) independent experiments. c, d Results represent the mean ± SD of n = 3 (c), n = 4 (for non-stimulated conditions) and n = 5 (for RMA conditions) (d) technical replicates and are representative of at least two (c) and three (d) independent experiments. Statistical comparisons are shown; *p = 0.05, **p = 0.01, ***p = 0.001, ****p = 0.0001, NS, non-significant; Student’s t-test. Source data are provided as a Source Data file

Fig 3: Analysis of cytotoxic activity of YT cells mutant for shp-2 (C1-CAR, C1) or harboring wild-type shp-2 gene (CAR-YT, YT), against peripheral blood mononuclear cells of a healthy donor.Incubation time was 4 hrs at a 1:1 E:T ratio. K – no effector cells were added. Average and standard deviation values for at least three independent measurements are shown.

Fig 4: Loss of LIFR activates NF-?B signaling through SHP1, leading to upregulation of LCN2.a HEK293T cells were transfected with HA-FLAG-SHP1 and SFB-tagged GFP or LIFR. LIFR-SFB protein was pulled down with S-protein beads, followed by immunoblotting with antibodies against FLAG and HA. b HEK293T cells were transfected with MYC-SHP2 and SFB-tagged GFP or LIFR. LIFR-SFB protein was pulled down with S-protein beads, followed by immunoblotting with antibodies against FLAG and MYC. c HEK293T SFB-GFP and SFB-LIFR stable cell lines were infected with the scrambled (Scr) or sh-SHP1 lentivirus, followed by transfection with a K63-specific mutant of His-Xpress-ubiquitin (Ub). 48 h later, cells were subjected to pulldown with nickel beads and immunoblotting with antibodies against TRAF6 and Xpress. d Control and LIFR-overexpressing PLC/PRF/5 cells were transduced with SHP1 shRNA and immunoblotted with the indicated antibodies. e Control (Scr) and LIFR-knockdown HEK293T cells were transfected with FLAG-TRAF6. 48 h later, cells were immunoprecipitated with a FLAG-specific antibody and immunoblotted with antibodies against LIFR, SHP1, and FLAG. f, g qPCR of LCN2, LIFR, and RELA in HEK293T (f) and PLC/PRF/5 (g) cells transduced with LIFR shRNA alone or in combination with p65 shRNA. n = 3 technical replicates. h qPCR of Lcn2, Lifr, and RelA in control and Lifr-knockout PHM cells transduced with the scrambled shRNA (Scr) or p65 shRNA. n = 3 technical replicates. i, j Immunohistochemical staining (i) and quantification (j) of Lcn2 in livers from Lifrfl/fl (F/F) and Lifrfl/fl;Alb-Cre (LKO) mice, 57 days after hydrodynamic injection of plasmids expressing the Sleeping Beauty transposase, myrAKT, RasV12, and shRNA (sh-p65, sh-Lcn2, or scrambled). n = 10, 8, 10, and 12 mice from left to right. Scale bars, 200 µm. k, l Liver weight (k) and liver-to-body weight ratio (l) of the mice described in i and j. n = 10, 8, 10, and 12 mice from left to right. m H&E staining of livers described in i and j. Scale bars, 300 µm. Statistical significance in f-h and j-l was determined by a two-tailed unpaired t-test. Error bars are s.e.m. Source data are provided as a Source Data file.

Fig 5: SHP2 inhibitors trigger apoptosis in MM cells. (A) RPMI-8226 and NCI-H929 cells were treated with 1, 10, 20, and 30 µM of SHP099 for 48 h; then, apoptotic rates were analyzed by flow cytometry. (B) RPMI-8226 and NCI-H929 cells were incubated with RMC-4550 (0~20 µM) for 48 h, and apoptotic cells were detected via flow cytometric analyses. The results represent the mean ± SD of three independent experiments. *p < 0.05, **p < 0.01, and ***p < 0.001.