Resveratrol suppresses human colon cancer cell proliferation and induces apoptosis via targeting the pentose phosphate and the talin-FAK signaling pathways-A proteomic approach

Abstract

Background: We and others have previously reported that resveratrol (RSV) suppresses colon cancer cell proliferation and elevates apoptosis in vitro and/or in vivo, however molecular mechanisms are not fully elucidated. Particularly, little information is available on RSV's effects on metabolic pathways and the cell-extra cellular matrix (ECM) communication that are critical for cancer cell growth. To identify important targets of RSV, we analyzed whole protein fractions from HT-29 advanced human colon cancer cell line treated with solvent control, IGF-1 (10 nM) and RSV (150 μM) using LC/MS/MS-Mud PIT (Multidimensional Protein Identification Technology).

Results: Pentose phosphate pathway (PPP), a vital metabolic pathway for cell cycle progression, was elevated and suppressed by IGF-1 and RSV, respectively in the HT-29 cell line. Enzymatic assays confirmed RSV suppression of glucose-6 phosphate dehydrogenase (rate limiting) and transketolase, key enzymes of the PPP. RSV (150 μM) suppressed, whereas IGF-1 (10 nM) elevated focal adhesion complex (FAC) proteins, talin and pFAK, critical for the cell-ECM communication. Western blotting analyses confirmed the suppression or elevation of these proteins in HT-29 cancer cells treated with RSV or IGF-1, respectively.

Conclusions: Proteomic analysis enabled us to establish PPP and the talin-pFAK as targets of RSV which suppress cancer cell proliferation and induce apoptosis in the colon cancer cell line HT-29. RSV (150 μM) suppressed these pathways in the presence and absence of IGF-1, suggesting its role as a chemo-preventive agent even in obese condition.

Figure 1

First 10 most differentially represented biological pathways identified in the set of proteins across all experimental conditions. The complete list of top 50 statistically significant pathways is given in Additional file 2, Figure S2.

Figure 2

Map of pentose phosphate pathway (Genego Metacore) overlapped with quantile-normalized estimations of protein abundance. Flags indicate direction of change (red, up-increased, blue; down-decreased) and fold change (length) of protein abundance in relation to solvent control. 1, IGF-1 and 2, Resveratrol. Results were expressed as mean for three replicate experiments for each treatment group.

Figure 3

Effect of RSV (50, 100 and 150 μM denoted as +, ++ and +++ respectively) and/or IGF-1 (10 nM) on G6PDH (A) and TKT (B) enzyme activities in HT-29 colon cancer cells. HT-29 cells were treated with RSV and/or IGF-1 for 24 h and G6PDH and TKT enzyme activities were analyzed as described in materials and methods. Results were expressed as means ± SE for three replicate experiments for each treatment group. Means that differ by a common letter (a, b, c, d, e, f) differ (p < 0.05).

Figure 4

Map of cytoskeleton remodeling pathway (Genego Metacore, fragment) overlapped with quantile-normalized estimations of protein abundance. Flags indicate direction of change (red, up-increased, blue; down-decreased) and fold change (length) of protein abundance in relation to non-treated control. 1, IGF-1 and 2, Resveratrol. Results were expressed as mean for three replicate experiments for each treatment group.

Figure 5

Effect of RSV (50, 100 and 150 μM denoted as +, ++ and +++ respectively) and/or IGF-1 (10 nM) on talin and pFAK proteins in HT-29 colon cancer cells. Western blot analysis was performed as described in materials and methods. β-actin served as a loading control. Results were expressed as mean ± SE for three replicate experiments for each treatment group. Means that differ by a common letter (a, b, c, d, e, f) differ (p < 0.05).