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RNA G-quadruplexes cause eIF4A-dependent oncogene translation in cancer.

Wolfe AL, Singh K, Zhong Y, Drewe P, Rajasekhar VK, Sanghvi VR, Mavrakis KJ, Jiang M, Roderick JE, Van der Meulen J, Schatz JH, Rodrigo CM, Zhao C, Rondou P, de Stanchina E, Teruya-Feldstein J, Kelliher MA, Speleman F, Porco JA, Pelletier J, Rätsch G, Wendel HG - Nature (2014)

Bottom Line: Accordingly, inhibition of eIF4A with silvestrol has powerful therapeutic effects against murine and human leukaemic cells in vitro and in vivo.Notably, among the most eIF4A-dependent and silvestrol-sensitive transcripts are a number of oncogenes, superenhancer-associated transcription factors, and epigenetic regulators.Hence, the 5' UTRs of select cancer genes harbour a targetable requirement for the eIF4A RNA helicase.

View Article: PubMed Central - PubMed

Affiliation: 1] Cancer Biology and Genetics, Memorial Sloan-Kettering Cancer Center, New York, New York 10065, USA [2] Weill Cornell Graduate School of Medical Sciences, New York, New York 10065, USA [3].

ABSTRACT
The translational control of oncoprotein expression is implicated in many cancers. Here we report an eIF4A RNA helicase-dependent mechanism of translational control that contributes to oncogenesis and underlies the anticancer effects of silvestrol and related compounds. For example, eIF4A promotes T-cell acute lymphoblastic leukaemia development in vivo and is required for leukaemia maintenance. Accordingly, inhibition of eIF4A with silvestrol has powerful therapeutic effects against murine and human leukaemic cells in vitro and in vivo. We use transcriptome-scale ribosome footprinting to identify the hallmarks of eIF4A-dependent transcripts. These include 5' untranslated region (UTR) sequences such as the 12-nucleotide guanine quartet (CGG)4 motif that can form RNA G-quadruplex structures. Notably, among the most eIF4A-dependent and silvestrol-sensitive transcripts are a number of oncogenes, superenhancer-associated transcription factors, and epigenetic regulators. Hence, the 5' UTRs of select cancer genes harbour a targetable requirement for the eIF4A RNA helicase.

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Immunoblots and mRNA expressiona) Lysates from human T-ALL lines treated with CR (25 nM, 24H) and probed as indicated; b) Lysates from JURKAT cells treated with escalating doses of Silvestrol and probed as indicated; c) mRNA levels for the indicated genes treated with vehicle (DMSO, black) or Silvestrol (red, 25 nM) for 45 minutes. Mean and standard deviation are shown, n = 2 biological replicates; d–g) Immunoblots of lysates from murine T-ALL cells expressing either vector control or IRES-MYC (d), IRES-CCND3 T283A (e), IRES-ICN (f), or IRES-BCL2 (g) and probed as indicated.
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Figure 13: Immunoblots and mRNA expressiona) Lysates from human T-ALL lines treated with CR (25 nM, 24H) and probed as indicated; b) Lysates from JURKAT cells treated with escalating doses of Silvestrol and probed as indicated; c) mRNA levels for the indicated genes treated with vehicle (DMSO, black) or Silvestrol (red, 25 nM) for 45 minutes. Mean and standard deviation are shown, n = 2 biological replicates; d–g) Immunoblots of lysates from murine T-ALL cells expressing either vector control or IRES-MYC (d), IRES-CCND3 T283A (e), IRES-ICN (f), or IRES-BCL2 (g) and probed as indicated.

Mentions: We confirmed the Silvestrol effects (25 nM for 24h) on key target proteins (Figure 5d, Extended Data Fig. 7a). The effect on the MYC protein was especially striking, it lasted for up 48h, and was readily detected in xenografts in vivo (Figure 5d–f, Extended Data Fig. 8a/b). The corresponding mRNAs were unchanged, and the increase in MYC mRNA may reflect a known auto-regulatory mechanism35 (Extended Data Fig. 8c). To test which proteins account for Silvestrol’s effects, we expressed several candidate genes and GFP in an eIF4A-independent manner from the HCV IRES. In mixed populations of transduced and parental murine leukemic cells (see Fig. 1c), we found that individually genes were not protective, while co-expression of IRES-MYC and IRES-BCL2 afforded significant protection (Figure 5g, Extended Data Fig. 8d–g). This result indicates that Silvestrol kills cancer cells by disrupting the translation of several critical factors.


RNA G-quadruplexes cause eIF4A-dependent oncogene translation in cancer.

Wolfe AL, Singh K, Zhong Y, Drewe P, Rajasekhar VK, Sanghvi VR, Mavrakis KJ, Jiang M, Roderick JE, Van der Meulen J, Schatz JH, Rodrigo CM, Zhao C, Rondou P, de Stanchina E, Teruya-Feldstein J, Kelliher MA, Speleman F, Porco JA, Pelletier J, Rätsch G, Wendel HG - Nature (2014)

Immunoblots and mRNA expressiona) Lysates from human T-ALL lines treated with CR (25 nM, 24H) and probed as indicated; b) Lysates from JURKAT cells treated with escalating doses of Silvestrol and probed as indicated; c) mRNA levels for the indicated genes treated with vehicle (DMSO, black) or Silvestrol (red, 25 nM) for 45 minutes. Mean and standard deviation are shown, n = 2 biological replicates; d–g) Immunoblots of lysates from murine T-ALL cells expressing either vector control or IRES-MYC (d), IRES-CCND3 T283A (e), IRES-ICN (f), or IRES-BCL2 (g) and probed as indicated.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492470&req=5

Figure 13: Immunoblots and mRNA expressiona) Lysates from human T-ALL lines treated with CR (25 nM, 24H) and probed as indicated; b) Lysates from JURKAT cells treated with escalating doses of Silvestrol and probed as indicated; c) mRNA levels for the indicated genes treated with vehicle (DMSO, black) or Silvestrol (red, 25 nM) for 45 minutes. Mean and standard deviation are shown, n = 2 biological replicates; d–g) Immunoblots of lysates from murine T-ALL cells expressing either vector control or IRES-MYC (d), IRES-CCND3 T283A (e), IRES-ICN (f), or IRES-BCL2 (g) and probed as indicated.
Mentions: We confirmed the Silvestrol effects (25 nM for 24h) on key target proteins (Figure 5d, Extended Data Fig. 7a). The effect on the MYC protein was especially striking, it lasted for up 48h, and was readily detected in xenografts in vivo (Figure 5d–f, Extended Data Fig. 8a/b). The corresponding mRNAs were unchanged, and the increase in MYC mRNA may reflect a known auto-regulatory mechanism35 (Extended Data Fig. 8c). To test which proteins account for Silvestrol’s effects, we expressed several candidate genes and GFP in an eIF4A-independent manner from the HCV IRES. In mixed populations of transduced and parental murine leukemic cells (see Fig. 1c), we found that individually genes were not protective, while co-expression of IRES-MYC and IRES-BCL2 afforded significant protection (Figure 5g, Extended Data Fig. 8d–g). This result indicates that Silvestrol kills cancer cells by disrupting the translation of several critical factors.

Bottom Line: Accordingly, inhibition of eIF4A with silvestrol has powerful therapeutic effects against murine and human leukaemic cells in vitro and in vivo.Notably, among the most eIF4A-dependent and silvestrol-sensitive transcripts are a number of oncogenes, superenhancer-associated transcription factors, and epigenetic regulators.Hence, the 5' UTRs of select cancer genes harbour a targetable requirement for the eIF4A RNA helicase.

View Article: PubMed Central - PubMed

Affiliation: 1] Cancer Biology and Genetics, Memorial Sloan-Kettering Cancer Center, New York, New York 10065, USA [2] Weill Cornell Graduate School of Medical Sciences, New York, New York 10065, USA [3].

ABSTRACT
The translational control of oncoprotein expression is implicated in many cancers. Here we report an eIF4A RNA helicase-dependent mechanism of translational control that contributes to oncogenesis and underlies the anticancer effects of silvestrol and related compounds. For example, eIF4A promotes T-cell acute lymphoblastic leukaemia development in vivo and is required for leukaemia maintenance. Accordingly, inhibition of eIF4A with silvestrol has powerful therapeutic effects against murine and human leukaemic cells in vitro and in vivo. We use transcriptome-scale ribosome footprinting to identify the hallmarks of eIF4A-dependent transcripts. These include 5' untranslated region (UTR) sequences such as the 12-nucleotide guanine quartet (CGG)4 motif that can form RNA G-quadruplex structures. Notably, among the most eIF4A-dependent and silvestrol-sensitive transcripts are a number of oncogenes, superenhancer-associated transcription factors, and epigenetic regulators. Hence, the 5' UTRs of select cancer genes harbour a targetable requirement for the eIF4A RNA helicase.

Show MeSH
Related in: MedlinePlus