Figure 6: c-Src stimulated tumorigenesis depends on both HK1 and HK2.
From: c-Src phosphorylation and activation of hexokinase promotes tumorigenesis and metastasis
(a–c) Individual knockdown of HK1, HK2 and c-Src in HeLa cells resulted in proliferation retardation as determined by growth curves (a) MTT assays (b) and BrdU incorporation assays (c). (d) Re-expression of HK1 rescued the decreased proliferation rate in shHK1 HeLa cells. (e,f) The rescued cell proliferation resulted from re-expression of HK1 in shHK1 HeLa cells is antagonized by PP2 treatment (e) and knockdown c-Src (f). (g) shHK1 significantly attenuated xenograft tumour growth in nude mice. HeLa cells stably expressing shHK1 and shGFP were separately injected on the left and right flanks of mouse. Three weeks post-injection, pictures of isolated tumours (scale bars, 6.25 mm) were taken and tumour lysates were detected for HK1 expression (left panel). Tumour weights were determined (n=5). (h) Replacement of HK1 with HK1-Y732F retarded xenograft tumour growth. shHK1 HeLa cells rescued for WT HK1 and HK1-Y732F expressions were injected on the right and left flanks of mice, respectively. Pictures of isolated tumours (scale bars, 6 mm) were taken. Tumours were detected for HK1 expression and HK1-Y732 phosphorylation and tumour weights were determined (n=6). (i) Tumours shown in h were measured for HK activity and lactate production. (j) Overexpression of HK2 rescued cell proliferation rate in shHK1 HeLa cells. (k) Overexpression HK1 rescued the cell proliferation rate in shHK2 HeLa cells. (l) HK1 was knocked down in A549 cells that are deficient for HK2 expression, followed by rescue expression of HK1 and HK2. Cells were then incubated in medium containing 0.2 mM of glucose for 4 h, followed by determination of survival rates. Western blot (WB) was performed to detect proteins’ expressions. The data shown in panels a,c,e,f,i,l are means±s.d. of three independent experiments, in panel b,d,j,k are means±s.e.m. of three independent experiments in triplicates. The data in panels g,h were statistically analysed with two-way ANOVA. Other data were analysed by using unpaired Student’s t-test. *P<0.05, **P<0.01, ***P<0.001.
