The domains of MyoD and Runx1 involved in their interaction.
2010-02-25T00:35:14Z (GMT) by
<p><b>A.</b> The bHLH domain and the C-terminal transactivating domain of MyoD are involved in the interaction with Runx1 and CBFβ. <i>Top panel</i>, Schematic diagram of MyoD functional domains, TAD: transactivation domain, bHLH: basic Helix Loop Helix. <i>Lower panel</i>, Runx1 and CBFβ interact with MyoD wild-type and with MyoD deletion mutant containing the bHLH and the C-terminal domain. <i>Lower panel</i>, expression vectors for wild-type MyoD and its deletion mutants were transfected in HEK 293 cells using Calcium phosphate precipitation as described in Material and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0009425#s4" target="_blank">Methods</a> section. 48 h post-transfection, cells were lysed and lysates were subjected to anti-HA immunoprecipitation. Precipitates were then analyzed by western blotting using with the indicated antibodies (WB Ab). *: <i>specific bands</i>. <b>B.</b> The transcription regulation domain located in C-terminal part of Runx1 is required for the interaction with MyoD. <i>Top panel</i>, Schematic diagram of Runx1 protein. The Runt, transactivation (TA), and transcription inhibition (ID) domains are indicated. <i>Lower panel</i>, Runx1 and its deletion mutants, or luciferase (Luc) were <i>in vitro</i> translated in the presence of <sup>35</sup>S-Methionine (inputs shown on the right panel) and incubated with equivalent amounts of GST-MyoD or GST agarose beads (normalization is shown in the lower panel). GST pull-down was then conducted as described in the Material and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0009425#s4" target="_blank">Methods</a> section, and the radiolabeled proteins were detected by autoradiography.</p>