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Loss of H2A.Z Is Not Sufficient to Determine Transcriptional Activity of Snf2-Related CBP Activator Protein or p400 Complexes.

Bowman TA, Wong MM, Cox LK, Baldassare JJ, Chrivia JC - Int J Cell Biol (2011)

Bottom Line: However, knockdown of SRCAP or p400 reduces deposition of H2A.Z∼50% into all p21 and Sp1 promoter nucleosomes.These results indicate that the deposition of H2A.Z by the p400 or SRCAP complexes is not sufficient to determine how each regulates transcription.This conclusion is further supported by studies that demonstrate a SRCAP(ΔATP ) mutant unable to deposit H2A.Z has similar transcriptional activity as wild-type SRCAP.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacological and Physiological Science, Saint Louis University School of Medicine, 1402 South Grand Boulevard, Saint Louis, MO 63104, USA.

ABSTRACT
The p400 and SRCAP (Snf2-related CBP activator protein) complexes remodel chromatin by catalyzing deposition of histone H2A.Z into nucleosomes. This remodeling activity has been proposed as a basis for regulation of transcription by these complexes. Transcript levels of p21 or Sp1 mRNAs after knockdown of p400 or SRCAP reveals that each regulates transcription of these promoters differently. In this study, we asked whether deposition of H2A.Z within specific nucleosomes by p400 or SRCAP dictates transcriptional activity. Our data indicates that nucleosome density at specific p21 or Sp1 promoter positions is not altered by the loss of either remodeling complex. However, knockdown of SRCAP or p400 reduces deposition of H2A.Z∼50% into all p21 and Sp1 promoter nucleosomes. Thus, H2A.Z deposition is not targeted to specific nucleosomes. These results indicate that the deposition of H2A.Z by the p400 or SRCAP complexes is not sufficient to determine how each regulates transcription. This conclusion is further supported by studies that demonstrate a SRCAP(ΔATP ) mutant unable to deposit H2A.Z has similar transcriptional activity as wild-type SRCAP.

No MeSH data available.


Knockdown of p400  and SRCAP expression differently regulates transcription of the Sp1 promoter and p21 promoter.  A549 cells were transfected where indicated with control, SRCAP or p400 siRNA, harvested 72 hours later and total RNA isolated.  In (a), the level of Sp1 mRNA was assessed using RT-qPCR using primers listed in    [3]. In (b),  the level of p21 mRNA was determined by RT-qPCR using primers listed in Table  S1(a).  The graphs show the mean result and standard error from three experiments.
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fig3: Knockdown of p400 and SRCAP expression differently regulates transcription of the Sp1 promoter and p21 promoter. A549 cells were transfected where indicated with control, SRCAP or p400 siRNA, harvested 72 hours later and total RNA isolated. In (a), the level of Sp1 mRNA was assessed using RT-qPCR using primers listed in [3]. In (b), the level of p21 mRNA was determined by RT-qPCR using primers listed in Table  S1(a). The graphs show the mean result and standard error from three experiments.

Mentions: Previous studies in U2OS cells indicated that knockdown of p400 and SRCAP results in equivalent loss of H2A.Z deposition [5] at the p21 promoter. Surprisingly, loss of p400 expression resulted in activation of transcription of the p21 promoter whereas knockdown of SRCAP had no effect [5]. We subsequently confirmed these results in the lung adenocarcinoma A549 cell line (Figures 3(b) and S1 of the supplementary material available online at doi:10.1155/2001/715642). To understand how the p400 and SRCAP complexes differentially regulate transcription of the p21 promoter we carried out a series of experiments to determine if they deposit H2A.Z into distinct nucleosomes at distinct locations within the p21 promoter.


Loss of H2A.Z Is Not Sufficient to Determine Transcriptional Activity of Snf2-Related CBP Activator Protein or p400 Complexes.

Bowman TA, Wong MM, Cox LK, Baldassare JJ, Chrivia JC - Int J Cell Biol (2011)

Knockdown of p400  and SRCAP expression differently regulates transcription of the Sp1 promoter and p21 promoter.  A549 cells were transfected where indicated with control, SRCAP or p400 siRNA, harvested 72 hours later and total RNA isolated.  In (a), the level of Sp1 mRNA was assessed using RT-qPCR using primers listed in    [3]. In (b),  the level of p21 mRNA was determined by RT-qPCR using primers listed in Table  S1(a).  The graphs show the mean result and standard error from three experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3140016&req=5

fig3: Knockdown of p400 and SRCAP expression differently regulates transcription of the Sp1 promoter and p21 promoter. A549 cells were transfected where indicated with control, SRCAP or p400 siRNA, harvested 72 hours later and total RNA isolated. In (a), the level of Sp1 mRNA was assessed using RT-qPCR using primers listed in [3]. In (b), the level of p21 mRNA was determined by RT-qPCR using primers listed in Table  S1(a). The graphs show the mean result and standard error from three experiments.
Mentions: Previous studies in U2OS cells indicated that knockdown of p400 and SRCAP results in equivalent loss of H2A.Z deposition [5] at the p21 promoter. Surprisingly, loss of p400 expression resulted in activation of transcription of the p21 promoter whereas knockdown of SRCAP had no effect [5]. We subsequently confirmed these results in the lung adenocarcinoma A549 cell line (Figures 3(b) and S1 of the supplementary material available online at doi:10.1155/2001/715642). To understand how the p400 and SRCAP complexes differentially regulate transcription of the p21 promoter we carried out a series of experiments to determine if they deposit H2A.Z into distinct nucleosomes at distinct locations within the p21 promoter.

Bottom Line: However, knockdown of SRCAP or p400 reduces deposition of H2A.Z∼50% into all p21 and Sp1 promoter nucleosomes.These results indicate that the deposition of H2A.Z by the p400 or SRCAP complexes is not sufficient to determine how each regulates transcription.This conclusion is further supported by studies that demonstrate a SRCAP(ΔATP ) mutant unable to deposit H2A.Z has similar transcriptional activity as wild-type SRCAP.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacological and Physiological Science, Saint Louis University School of Medicine, 1402 South Grand Boulevard, Saint Louis, MO 63104, USA.

ABSTRACT
The p400 and SRCAP (Snf2-related CBP activator protein) complexes remodel chromatin by catalyzing deposition of histone H2A.Z into nucleosomes. This remodeling activity has been proposed as a basis for regulation of transcription by these complexes. Transcript levels of p21 or Sp1 mRNAs after knockdown of p400 or SRCAP reveals that each regulates transcription of these promoters differently. In this study, we asked whether deposition of H2A.Z within specific nucleosomes by p400 or SRCAP dictates transcriptional activity. Our data indicates that nucleosome density at specific p21 or Sp1 promoter positions is not altered by the loss of either remodeling complex. However, knockdown of SRCAP or p400 reduces deposition of H2A.Z∼50% into all p21 and Sp1 promoter nucleosomes. Thus, H2A.Z deposition is not targeted to specific nucleosomes. These results indicate that the deposition of H2A.Z by the p400 or SRCAP complexes is not sufficient to determine how each regulates transcription. This conclusion is further supported by studies that demonstrate a SRCAP(ΔATP ) mutant unable to deposit H2A.Z has similar transcriptional activity as wild-type SRCAP.

No MeSH data available.