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LARP7 suppresses P-TEFb activity to inhibit breast cancer progression and metastasis.

Ji X, Lu H, Zhou Q, Luo K - Elife (2014)

Bottom Line: The positive transcription elongation factor b (P-TEFb) stimulates transcriptional elongation by phosphorylating Pol II and antagonizing negative elongation factors.A reservoir of P-TEFb is sequestered in the inactive 7SK snRNP where 7SK snRNA and the La-related protein LARP7 are required for the integrity of this complex.Decreased levels of LARP7 and 7SK snRNA redistribute P-TEFb to the transcriptionally active super elongation complex, resulting in P-TEFb activation and increased transcription of EMT transcription factors, including Slug, FOXC2, ZEB2, and Twist1, to promote breast cancer EMT, invasion, and metastasis.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.

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Loss of LARP7 enhances transcription of key EMT genes.(A) LARP7 knockdown significantly increases the expression of key EMT transcriptional factors in T47D breast cancer cells as measured by qRT-PCR. The p values (*p<0.05) were determined by the Student's t test. (B) Cells were treated with flavopiridol for 7 hr at the indicated concentrations. The expression of EMT genes was assessed by qRT-PCR. (C) Cells were transfected with siCtrl, siCDK9-1 or siCDK9-2. After 48 hr, the expression of EMT genes was assessed by qRT-PCR. (D) Control or two shLARP7 T47D cells were subjected to ChIP analysis to determine the levels of CDK9 and phospho-Ser2 (pSer2) Pol II occupancy at various locations of EMT-related transcription factors and non-responsive genes. qRT-PCR was performed using primers specific to the transcription start site (TSS), interior and 3′-UTR of each gene, and the signals were normalized to that of input. Data are presented as the mean ± SD from three independent measurements.DOI:http://dx.doi.org/10.7554/eLife.02907.010
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fig5: Loss of LARP7 enhances transcription of key EMT genes.(A) LARP7 knockdown significantly increases the expression of key EMT transcriptional factors in T47D breast cancer cells as measured by qRT-PCR. The p values (*p<0.05) were determined by the Student's t test. (B) Cells were treated with flavopiridol for 7 hr at the indicated concentrations. The expression of EMT genes was assessed by qRT-PCR. (C) Cells were transfected with siCtrl, siCDK9-1 or siCDK9-2. After 48 hr, the expression of EMT genes was assessed by qRT-PCR. (D) Control or two shLARP7 T47D cells were subjected to ChIP analysis to determine the levels of CDK9 and phospho-Ser2 (pSer2) Pol II occupancy at various locations of EMT-related transcription factors and non-responsive genes. qRT-PCR was performed using primers specific to the transcription start site (TSS), interior and 3′-UTR of each gene, and the signals were normalized to that of input. Data are presented as the mean ± SD from three independent measurements.DOI:http://dx.doi.org/10.7554/eLife.02907.010

Mentions: Because P-TEFb is a transcription elongation factor that most likely affects breast cancer progression at the level of transcription, we decided to examine the expression of a panel of EMT regulators in the two T47D LARP7 KD cell lines (shLARP7-1 and shLARP7-2). Consistent with the enhanced EMT phenotypes, loss of LARP7 markedly increased the expression of many key EMT and metastasis genes (Zeisberg and Neilson, 2009), including Slug, FOXC2, ZEB2, Twist1, ZEB1, Snail, Twist2, and SOX10 (Figure 5A). Notably, treatment of these LARP7 KD cells with flavopiridol or introduction of siCDK9s strongly inhibited the expression of four of those genes: Slug, FOXC2, ZEB2, and Twist1 (Figure 5B,C), suggesting that P-TEFb may directly affect their expression.10.7554/eLife.02907.010Figure 5.Loss of LARP7 enhances transcription of key EMT genes.


LARP7 suppresses P-TEFb activity to inhibit breast cancer progression and metastasis.

Ji X, Lu H, Zhou Q, Luo K - Elife (2014)

Loss of LARP7 enhances transcription of key EMT genes.(A) LARP7 knockdown significantly increases the expression of key EMT transcriptional factors in T47D breast cancer cells as measured by qRT-PCR. The p values (*p<0.05) were determined by the Student's t test. (B) Cells were treated with flavopiridol for 7 hr at the indicated concentrations. The expression of EMT genes was assessed by qRT-PCR. (C) Cells were transfected with siCtrl, siCDK9-1 or siCDK9-2. After 48 hr, the expression of EMT genes was assessed by qRT-PCR. (D) Control or two shLARP7 T47D cells were subjected to ChIP analysis to determine the levels of CDK9 and phospho-Ser2 (pSer2) Pol II occupancy at various locations of EMT-related transcription factors and non-responsive genes. qRT-PCR was performed using primers specific to the transcription start site (TSS), interior and 3′-UTR of each gene, and the signals were normalized to that of input. Data are presented as the mean ± SD from three independent measurements.DOI:http://dx.doi.org/10.7554/eLife.02907.010
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4126343&req=5

fig5: Loss of LARP7 enhances transcription of key EMT genes.(A) LARP7 knockdown significantly increases the expression of key EMT transcriptional factors in T47D breast cancer cells as measured by qRT-PCR. The p values (*p<0.05) were determined by the Student's t test. (B) Cells were treated with flavopiridol for 7 hr at the indicated concentrations. The expression of EMT genes was assessed by qRT-PCR. (C) Cells were transfected with siCtrl, siCDK9-1 or siCDK9-2. After 48 hr, the expression of EMT genes was assessed by qRT-PCR. (D) Control or two shLARP7 T47D cells were subjected to ChIP analysis to determine the levels of CDK9 and phospho-Ser2 (pSer2) Pol II occupancy at various locations of EMT-related transcription factors and non-responsive genes. qRT-PCR was performed using primers specific to the transcription start site (TSS), interior and 3′-UTR of each gene, and the signals were normalized to that of input. Data are presented as the mean ± SD from three independent measurements.DOI:http://dx.doi.org/10.7554/eLife.02907.010
Mentions: Because P-TEFb is a transcription elongation factor that most likely affects breast cancer progression at the level of transcription, we decided to examine the expression of a panel of EMT regulators in the two T47D LARP7 KD cell lines (shLARP7-1 and shLARP7-2). Consistent with the enhanced EMT phenotypes, loss of LARP7 markedly increased the expression of many key EMT and metastasis genes (Zeisberg and Neilson, 2009), including Slug, FOXC2, ZEB2, Twist1, ZEB1, Snail, Twist2, and SOX10 (Figure 5A). Notably, treatment of these LARP7 KD cells with flavopiridol or introduction of siCDK9s strongly inhibited the expression of four of those genes: Slug, FOXC2, ZEB2, and Twist1 (Figure 5B,C), suggesting that P-TEFb may directly affect their expression.10.7554/eLife.02907.010Figure 5.Loss of LARP7 enhances transcription of key EMT genes.

Bottom Line: The positive transcription elongation factor b (P-TEFb) stimulates transcriptional elongation by phosphorylating Pol II and antagonizing negative elongation factors.A reservoir of P-TEFb is sequestered in the inactive 7SK snRNP where 7SK snRNA and the La-related protein LARP7 are required for the integrity of this complex.Decreased levels of LARP7 and 7SK snRNA redistribute P-TEFb to the transcriptionally active super elongation complex, resulting in P-TEFb activation and increased transcription of EMT transcription factors, including Slug, FOXC2, ZEB2, and Twist1, to promote breast cancer EMT, invasion, and metastasis.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.

Show MeSH
Related in: MedlinePlus