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Targeting a ribonucleoprotein complex containing the caprin-1 protein and the c-Myc mRNA suppresses tumor growth in mice: an identification of a novel oncotarget.

Qiu YQ, Yang CW, Lee YZ, Yang RB, Lee CH, Hsu HY, Chang CC, Lee SJ - Oncotarget (2015)

Bottom Line: Gene expression profiling and gain-of-c-Myc-function experiments also revealed that the downregulated c-Myc contributes to the anti-oncogenic effects of tylophorine compounds.Furthermore, the potent tylophorine derivative dibenzoquinoline-33b elicited a similar effect, as c-Myc protein levels were also decreased in xenograft tumors treated with dibenzoquinoline-33b.Thus, tylophorine compounds exert anti-cancer activity predominantly by targeting and sequestering the caprin-1 protein and c-Myc mRNA associated ribonucleoprotein complex.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli, Taiwan.

ABSTRACT
Tylophorine compounds have been the focus of drug development for decades. Tylophorine derivatives exhibit anti-cancer activities but their cellular targets remain unknown. We used a biotinylated tylophorine derivative to probe for the interacting cellular target(s) of tylophorine. Tylophorine directly binds to caprin-1 and consequently enhances the recruitment of G3BP1, c-Myc mRNA, and cyclin D2 mRNA to form a ribonucleoprotein complex. Subsequently, this tylophorine targeted ribonucleoprotein complex is sequestered to the polysomal fractions and the protein expressions of the associated mRNA-transcripts are repressed. Caprin-1 depleted carcinoma cells become more resistant to tylophorine, associated with decreased formation of the ribonucleoprotein complex targeted by tylophorine. Consequently, tylophorine downregulates c-Myc and cyclins D1/D2, causing hypophosphorylation of Rb and suppression of both processing-body formation and the Warburg effect. Gene expression profiling and gain-of-c-Myc-function experiments also revealed that the downregulated c-Myc contributes to the anti-oncogenic effects of tylophorine compounds. Furthermore, the potent tylophorine derivative dibenzoquinoline-33b elicited a similar effect, as c-Myc protein levels were also decreased in xenograft tumors treated with dibenzoquinoline-33b. Thus, tylophorine compounds exert anti-cancer activity predominantly by targeting and sequestering the caprin-1 protein and c-Myc mRNA associated ribonucleoprotein complex.

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Gain-of-function experiments for tylophorine-induced decreased protein expression of cyclin D1, cyclin D2, and c-MycA. Ectopically overexpressed cyclin D1 or D2 restored the hyperphosphorylation of pRb and the expression of cyclin A2. B. Ectopically overexpressed c-Myc rescued the biological function of its downstream effectors (e.g., cyclin B1 and p-pRb) but not the protein expression of cyclin D1 and D2. C. Ectopically overexpressed cyclin A2 moderately restored the hyperphosphorylation of pRb. HONE-1 cells were transfected with the indicated expression vectors for 24 h prior to treatment with either vehicle (DMSO) or tylophorine at the indicated concentrations for an additional 24 h. The resulting cell lysates were analyzed by western blotting with the indicated antibodies. The relative expression or phosphorylation levels of each protein were normalized with their respective internal loading control α-tubulin or GAPDH. The results shown are representative of 3 independent experiments.
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Figure 4: Gain-of-function experiments for tylophorine-induced decreased protein expression of cyclin D1, cyclin D2, and c-MycA. Ectopically overexpressed cyclin D1 or D2 restored the hyperphosphorylation of pRb and the expression of cyclin A2. B. Ectopically overexpressed c-Myc rescued the biological function of its downstream effectors (e.g., cyclin B1 and p-pRb) but not the protein expression of cyclin D1 and D2. C. Ectopically overexpressed cyclin A2 moderately restored the hyperphosphorylation of pRb. HONE-1 cells were transfected with the indicated expression vectors for 24 h prior to treatment with either vehicle (DMSO) or tylophorine at the indicated concentrations for an additional 24 h. The resulting cell lysates were analyzed by western blotting with the indicated antibodies. The relative expression or phosphorylation levels of each protein were normalized with their respective internal loading control α-tubulin or GAPDH. The results shown are representative of 3 independent experiments.

Mentions: pRb protein, a common downstream effector of cyclins D1 [8, 9] and D2 [10, 11] and c-Myc [1, 7], was further examined for tylophorine effects on the anticancer activity. pRb was hypophosphorylated to the active form in tylophorine-treated carcinoma cells, whereas its protein expression levels remained unchanged (Fig. 3B); this result was expected because the protein expressions of the pRb's upstream regulators, cyclins D1 [8, 9]/D2 [10, 11] and c-Myc 1, 7], were reduced by tylophorine treatment thereby decreasing the phosphorylation of pRb (Fig. 3B). Cyclin A2 was downregulated by tylophorine, which partially accounts for the G1 arrest induced by tylophorine [20, 25]. The cyclin A2 promoter region contains two E2F regulatory sites. When either cyclin D2 (Fig. 4A-upper panel) or cyclin D1 (Fig. 4A-lower panel) was overexpressed, the downstream hyperphosphorylation of pRb resumed, thereby upregulating E2F-mediated cyclin A2 expression [35] in tylophorine-treated cells (Fig. 4A).


Targeting a ribonucleoprotein complex containing the caprin-1 protein and the c-Myc mRNA suppresses tumor growth in mice: an identification of a novel oncotarget.

Qiu YQ, Yang CW, Lee YZ, Yang RB, Lee CH, Hsu HY, Chang CC, Lee SJ - Oncotarget (2015)

Gain-of-function experiments for tylophorine-induced decreased protein expression of cyclin D1, cyclin D2, and c-MycA. Ectopically overexpressed cyclin D1 or D2 restored the hyperphosphorylation of pRb and the expression of cyclin A2. B. Ectopically overexpressed c-Myc rescued the biological function of its downstream effectors (e.g., cyclin B1 and p-pRb) but not the protein expression of cyclin D1 and D2. C. Ectopically overexpressed cyclin A2 moderately restored the hyperphosphorylation of pRb. HONE-1 cells were transfected with the indicated expression vectors for 24 h prior to treatment with either vehicle (DMSO) or tylophorine at the indicated concentrations for an additional 24 h. The resulting cell lysates were analyzed by western blotting with the indicated antibodies. The relative expression or phosphorylation levels of each protein were normalized with their respective internal loading control α-tubulin or GAPDH. The results shown are representative of 3 independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4385842&req=5

Figure 4: Gain-of-function experiments for tylophorine-induced decreased protein expression of cyclin D1, cyclin D2, and c-MycA. Ectopically overexpressed cyclin D1 or D2 restored the hyperphosphorylation of pRb and the expression of cyclin A2. B. Ectopically overexpressed c-Myc rescued the biological function of its downstream effectors (e.g., cyclin B1 and p-pRb) but not the protein expression of cyclin D1 and D2. C. Ectopically overexpressed cyclin A2 moderately restored the hyperphosphorylation of pRb. HONE-1 cells were transfected with the indicated expression vectors for 24 h prior to treatment with either vehicle (DMSO) or tylophorine at the indicated concentrations for an additional 24 h. The resulting cell lysates were analyzed by western blotting with the indicated antibodies. The relative expression or phosphorylation levels of each protein were normalized with their respective internal loading control α-tubulin or GAPDH. The results shown are representative of 3 independent experiments.
Mentions: pRb protein, a common downstream effector of cyclins D1 [8, 9] and D2 [10, 11] and c-Myc [1, 7], was further examined for tylophorine effects on the anticancer activity. pRb was hypophosphorylated to the active form in tylophorine-treated carcinoma cells, whereas its protein expression levels remained unchanged (Fig. 3B); this result was expected because the protein expressions of the pRb's upstream regulators, cyclins D1 [8, 9]/D2 [10, 11] and c-Myc 1, 7], were reduced by tylophorine treatment thereby decreasing the phosphorylation of pRb (Fig. 3B). Cyclin A2 was downregulated by tylophorine, which partially accounts for the G1 arrest induced by tylophorine [20, 25]. The cyclin A2 promoter region contains two E2F regulatory sites. When either cyclin D2 (Fig. 4A-upper panel) or cyclin D1 (Fig. 4A-lower panel) was overexpressed, the downstream hyperphosphorylation of pRb resumed, thereby upregulating E2F-mediated cyclin A2 expression [35] in tylophorine-treated cells (Fig. 4A).

Bottom Line: Gene expression profiling and gain-of-c-Myc-function experiments also revealed that the downregulated c-Myc contributes to the anti-oncogenic effects of tylophorine compounds.Furthermore, the potent tylophorine derivative dibenzoquinoline-33b elicited a similar effect, as c-Myc protein levels were also decreased in xenograft tumors treated with dibenzoquinoline-33b.Thus, tylophorine compounds exert anti-cancer activity predominantly by targeting and sequestering the caprin-1 protein and c-Myc mRNA associated ribonucleoprotein complex.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli, Taiwan.

ABSTRACT
Tylophorine compounds have been the focus of drug development for decades. Tylophorine derivatives exhibit anti-cancer activities but their cellular targets remain unknown. We used a biotinylated tylophorine derivative to probe for the interacting cellular target(s) of tylophorine. Tylophorine directly binds to caprin-1 and consequently enhances the recruitment of G3BP1, c-Myc mRNA, and cyclin D2 mRNA to form a ribonucleoprotein complex. Subsequently, this tylophorine targeted ribonucleoprotein complex is sequestered to the polysomal fractions and the protein expressions of the associated mRNA-transcripts are repressed. Caprin-1 depleted carcinoma cells become more resistant to tylophorine, associated with decreased formation of the ribonucleoprotein complex targeted by tylophorine. Consequently, tylophorine downregulates c-Myc and cyclins D1/D2, causing hypophosphorylation of Rb and suppression of both processing-body formation and the Warburg effect. Gene expression profiling and gain-of-c-Myc-function experiments also revealed that the downregulated c-Myc contributes to the anti-oncogenic effects of tylophorine compounds. Furthermore, the potent tylophorine derivative dibenzoquinoline-33b elicited a similar effect, as c-Myc protein levels were also decreased in xenograft tumors treated with dibenzoquinoline-33b. Thus, tylophorine compounds exert anti-cancer activity predominantly by targeting and sequestering the caprin-1 protein and c-Myc mRNA associated ribonucleoprotein complex.

Show MeSH
Related in: MedlinePlus