Abstract
The cyclin-dependent kinase inhibitor p21 plays a critical role in regulating cell cycle and cell proliferation. We previously cloned the dog p21 gene and found that unlike human p21, dog p21 is expressed as 2 isoforms due to the proline-directed phosphorylation at serine 123 (S123). Here, we identifi ed that PPM1D, also called Wip1 and a Mg2+-dependent phosphatase, dephosphorylates dog p21 protein at serine 123. Specifically, we showed that the level of S123-phosphorylated dog p21 is increased by a PPM1D inhibitor in a dose-dependent manner. We also showed that over-expression of PPM1D decreases, whereas knockdown of PPM1D increases, the level of S123-phosphorylated dog p21 regardless of p53. Additionally, in vitro phosphatase assay was performed and showed that phosphorylated S123 in dog p21 is dephosphorylated by recombinant rPPM1D, which contains the catalytic domain of human PPM1D (residue 1-420), but not by the phosphatase dead rPPM1D (D314A). Furthermore, dephosphorylation of S123 by rPPM1D can be abrogated by PPM1D inhibitor or by withdrawal of Mg2+. Finally, we showed that upon PPM1D inhibition, the level of S123-phosphorylated dog p21 was increased, concomitantly with decreased expression of cyclin A, cyclin B, Rb, and PCNA. Together, our results indicate that PPM1D functions as a phosphatase of dog p21 at serine 123 and plays a role in cell cycle control via p21.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 641-647 |
| Number of pages | 7 |
| Journal | Cell Cycle |
| Volume | 14 |
| Issue number | 4 |
| DOIs | |
| State | Published - Feb 15 2015 |
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Keywords
- P21
- P53
- Phosphatase
- PPM1D
- Wip1
ASJC Scopus subject areas
- Cell Biology
- Molecular Biology
- Developmental Biology
Cite this
PPM1D regulates p21 expression via dephoshporylation at serine 123. / Cao, Ruibing; Zhang, Jin; Zhang, Min; Chen, Xinbin.
In: Cell Cycle, Vol. 14, No. 4, 15.02.2015, p. 641-647.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - PPM1D regulates p21 expression via dephoshporylation at serine 123
AU - Cao, Ruibing
AU - Zhang, Jin
AU - Zhang, Min
AU - Chen, Xinbin
PY - 2015/2/15
Y1 - 2015/2/15
N2 - The cyclin-dependent kinase inhibitor p21 plays a critical role in regulating cell cycle and cell proliferation. We previously cloned the dog p21 gene and found that unlike human p21, dog p21 is expressed as 2 isoforms due to the proline-directed phosphorylation at serine 123 (S123). Here, we identifi ed that PPM1D, also called Wip1 and a Mg2+-dependent phosphatase, dephosphorylates dog p21 protein at serine 123. Specifically, we showed that the level of S123-phosphorylated dog p21 is increased by a PPM1D inhibitor in a dose-dependent manner. We also showed that over-expression of PPM1D decreases, whereas knockdown of PPM1D increases, the level of S123-phosphorylated dog p21 regardless of p53. Additionally, in vitro phosphatase assay was performed and showed that phosphorylated S123 in dog p21 is dephosphorylated by recombinant rPPM1D, which contains the catalytic domain of human PPM1D (residue 1-420), but not by the phosphatase dead rPPM1D (D314A). Furthermore, dephosphorylation of S123 by rPPM1D can be abrogated by PPM1D inhibitor or by withdrawal of Mg2+. Finally, we showed that upon PPM1D inhibition, the level of S123-phosphorylated dog p21 was increased, concomitantly with decreased expression of cyclin A, cyclin B, Rb, and PCNA. Together, our results indicate that PPM1D functions as a phosphatase of dog p21 at serine 123 and plays a role in cell cycle control via p21.
AB - The cyclin-dependent kinase inhibitor p21 plays a critical role in regulating cell cycle and cell proliferation. We previously cloned the dog p21 gene and found that unlike human p21, dog p21 is expressed as 2 isoforms due to the proline-directed phosphorylation at serine 123 (S123). Here, we identifi ed that PPM1D, also called Wip1 and a Mg2+-dependent phosphatase, dephosphorylates dog p21 protein at serine 123. Specifically, we showed that the level of S123-phosphorylated dog p21 is increased by a PPM1D inhibitor in a dose-dependent manner. We also showed that over-expression of PPM1D decreases, whereas knockdown of PPM1D increases, the level of S123-phosphorylated dog p21 regardless of p53. Additionally, in vitro phosphatase assay was performed and showed that phosphorylated S123 in dog p21 is dephosphorylated by recombinant rPPM1D, which contains the catalytic domain of human PPM1D (residue 1-420), but not by the phosphatase dead rPPM1D (D314A). Furthermore, dephosphorylation of S123 by rPPM1D can be abrogated by PPM1D inhibitor or by withdrawal of Mg2+. Finally, we showed that upon PPM1D inhibition, the level of S123-phosphorylated dog p21 was increased, concomitantly with decreased expression of cyclin A, cyclin B, Rb, and PCNA. Together, our results indicate that PPM1D functions as a phosphatase of dog p21 at serine 123 and plays a role in cell cycle control via p21.
KW - P21
KW - P53
KW - Phosphatase
KW - PPM1D
KW - Wip1
UR - http://www.scopus.com/inward/record.url?scp=84923544381&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84923544381&partnerID=8YFLogxK
U2 - 10.4161/15384101.2014.994922
DO - 10.4161/15384101.2014.994922
M3 - Article
C2 - 25590690
AN - SCOPUS:84923544381
VL - 14
SP - 641
EP - 647
JO - Cell Cycle
JF - Cell Cycle
SN - 1538-4101
IS - 4
ER -