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

. 2013 Oct 14;203(1):57–71. doi: 10.1083/jcb.201305159

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

© 2013 Raghuram et al.

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).

PMC Copyright notice

Figure 2. — Pin1 promotes H1 dephosphorylation. (A) Histones were extracted from Pin1^−/−, Pin1wt, and Ciras-3 cells, and were then probed with either pS173(H1.2/H1.5), pS187(H1.4), a phospho-specific stain that labels all phosphorylated proteins, or with a stain that labels total protein. Levels of pS173, pS187, and net H1 phosphorylation levels were found to be higher in Pin1^−/− cells as compared with Pin1wt cells, similar to those observed in Ciras-3 cells (positive control). (B) Nuclear extracts from Pin1^−/− cells and Pin1wt cells revealed that the levels of Cdk2 and PP2Ac were similar in both cells. (C) The dephosphorylation activity of PP2Ac activity was analyzed using purified H1 as a substrate. PP2Ac was immunoprecipitated from either Pin1wt cells or Pin1^−/− cells and assessed for its ability to dephosphorylate pS187. The kinetics of this dephosphorylation reaction are plotted in E with each dot/square representing the average H1 phosphorylation level obtained from at least three independent experiments. The average intensity from the zero-minute time point is set as the maximum, against which all other time points are compared. (D) PP2Ac was immunoprecipitated from Pin1^−/− cells and was mixed with a constant amount of H1, while levels of purified Pin1 were varied from 0.004 to 8 µg. The former corresponds to a molar stoichiometry of H1/Pin1 = 1:0.0005, whereas the latter corresponds to H1/Pin1 = 1:0.9. The kinetics of dephosphorylation is plotted in F and G, with the average intensity at the zero-minute time point set to 1. These curves were then submitted to a one-phase decay curve analysis and the rate obtained was plotted as a function of the amount of Pin1 added to the reaction (H).