Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2015 Nov 17;44(6):2538–2553. doi: 10.1093/nar/gkv1244

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

PMC Copyright notice

Figure 4. — HEY protein links DPF3a to DNA. (A) DPF3a specifically interacts with HEY1. HEK293 cells were transiently transfected with Flag-tagged DPF3a and with HA-tagged MYOD1, HAND1, HES1 or HEY1. Cell lysates were incubated with an anti-HA matrix or anti-Flag matrix to pull down HA-tagged or Flag-tagged protein together with interacting proteins, which were further probed with anti-Flag or anti-HA antibody in immunoblots. (B) C2C12 cells were transfected with Hey1 siRNA for 48 h and knockdown efficiency of Hey1 in C2C12 cells was analyzed by qPCR and immunoblotting. (C and D) Dpf3a ChIP experiments were performed and Dpf3a enrichment at selected promoters in siHey1 knockdown was compared to siNon control. Binding of Dpf3a was quantified by qPCR (triplicate experiments, **P < 0.01, t-test). (E) The interaction of DPF3a and HEY1 is mediated via the half PHD finger. Co-immunoprecipitation (CoIP) of HA-tagged HEY1 with different Flag-tagged DPF3a constructs (WT, aa1–246, aa1–292 and aa292–357) from lysates of transiently transfected HEK293 cells. (F) CoIP of Flag-tagged DPF3a and HA-tagged HEY1 followed by de-phosphorylation assay using FastAP Alkaline phosphatase. S: supernatant. (G) Phosphorylation of serine 348 of DPF3a is critical for the interaction with HEY1. CoIP of HA-tagged HEY1 with Flag-tagged DPF3a mutants (WT, S348A, 5xAla). (H) Quantification of the DPF3a and HEY1 interaction using Image Lab 5.0 (triplicate experiments, **P < 0.01, t-test).