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p300 is required for MyoD-dependent cell cycle arrest and muscle-specific gene transcription
P L Puri1, M L Avantaggiati, C Balsano
1Fondazione Andrea Cesalpino, Università degli Studi di Roma La Sapienza, Rome, Italy.
Abstract:
The nuclear phosphoprotein p300 is a new member of a family of 'co-activators' (which also includes the CREB binding protein CBP), that directly modulate transcription by interacting with components of the basal transcriptional machinery. Both p300 and CBP are targeted by the adenovirus E1A protein, and binding to p300 is required for E1A to inhibit terminal differentiation in both keratinocytes and myoblasts. Here we demonstrate that, in differentiating skeletal muscle cells, p300 physically interacts with the myogenic basic helix-loop-helix (bHLH) regulatory protein MyoD at its DNA binding sites. During muscle differentiation, MyoD plays a dual role: besides activating muscle-specific transcription, it induces permanent cell cycle arrest by up-regulating the cyclin-dependent kinase inhibitor p21. We show that p300 is involved in both these activities. Indeed, E1A mutants lacking the ability to bind p300 are greatly impaired in the repression of E-box-driven transcription, and p300 overexpression rescues the wild-type E1A-mediated repression. Moreover, p300 potentiates MyoD- and myogenin-dependent activation of transcription from E-box-containing reporter genes. We also provide evidence, obtained by microinjection of anti-p300 antibodies, that p300 is required for MyoD-dependent cell cycle arrest in either myogenic cells induced to differentiate or in MyoD-converted C3H10T1/2 fibroblasts, but is dispensable for maintenance of the postmitotic state of myotubes.
Insights
The nuclear phosphoprotein p300 interacts with MyoD to regulate muscle cell differentiation and cell cycle arrest. This co-activator is crucial for MyoD
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Nuclear phosphoprotein p300 acts as a co-activator, modulating transcription by interacting with basal transcriptional machinery.
- p300 and CBP are targeted by adenovirus E1A protein, crucial for inhibiting keratinocyte and myoblast differentiation.
Purpose of the Study:
- To investigate the role of p300 in the physical interaction with MyoD during skeletal muscle differentiation.
- To elucidate p300's involvement in both MyoD-mediated transcription activation and cell cycle arrest.
Main Methods:
- Demonstration of physical interaction between p300 and MyoD at DNA binding sites in differentiating skeletal muscle cells.
- Analysis of E1A mutants lacking p300 binding ability to assess transcriptional repression.
- Overexpression of p300 to rescue E1A-mediated repression.
- Microinjection of anti-p300 antibodies to evaluate p300's requirement in MyoD-dependent processes.
Main Results:
- p300 physically interacts with MyoD at its DNA binding sites during muscle differentiation.
- p300 is essential for both MyoD-dependent activation of muscle-specific transcription and induction of cell cycle arrest via p21.
- Adenovirus E1A mutants deficient in p300 binding show impaired repression of E-box-driven transcription, which can be rescued by p300 overexpression.
- p300 is required for MyoD-dependent cell cycle arrest but not for maintaining the postmitotic state of myotubes.
Conclusions:
- p300 plays a critical role in regulating skeletal muscle differentiation by modulating MyoD activity.
- The co-activator p300 is indispensable for MyoD-driven transcriptional activation and cell cycle arrest during myogenesis.
- p300's interaction with MyoD is a key mechanism controlling muscle cell fate and proliferation.