Phosphatase Subfamily PPM1A

Phosphatase Classification: Fold PPM (PP2C): Superfamily PPM (PP2C): Family PPM (PP2C): Subfamily PPM1A

Evolution

The PPM1A subfamily is found across eukaryotes.

Human PPM1A and PPM1B probably arose by gene duplication in jawed vertebrates, but not through whole-genome duplication, as they do not locate in the same double-conserved synteny (see Genomicus).

Human PPM1N probably emerged in placentals and lost by independent evolutionary events in different lineages, as implied by internal orthology database and BLAST NR database. For BLAST, human PPM1N was defined as the hits that have a better (lower) E-values than human PPM1A and PPM1B.

Thus, all jawed vertebrate PPM1As originated from a single ancestral gene, which we call PPM1A-1 at here. Jawless vertebrates and other metazoa have both vtPPM1A-1 and another phosphatase gene, which we call PPM1A-2 at here. Monosiga only has PPM1A-1. Fungi, plants and basal eukaryotes have PPM1As slightly more similar to PPM1A-2 in sequence. Taken together, PPM1A-1 and PPM1A-2 probably arose in holozoa which was lost in jawed vertebrates.

PPM1As duplication happened in many lineages. For instance, budding yeast has three genes, and most plants have two or more genes.

Domains

The PPM1A subfamily has a single structural domain, the phosphatase domain. Human PPM1A and PPM1N have a predicted nuclear localization signal (NLS) at N-terminal. Human PPM1B dephosphorylates at least two transcription factors, but the NLS is weak by prediction. PPM1As from other organisms have weak NLS by prediction, as well.

Functions

The PPM1A subfamily regulates different signaling pathways, such as MAPK, SAPK/JNK, NF-kappaB, TGF-beta pathways.

Common substrates of human PPM1A and PPM1B

• IKKβ at Ser-177 and Ser-181. Both human PPM1A and PPM1B dephosphorylated IKKβ at Ser-177 and Ser-181 and termination of IKKβ-induced NF-κB activation [1, 2].
• CDK at Thr-186 in the T-loop. Both PPM1A and PPM1B dephosphorylated CDK9 at Thr-186 in the T-loop [3]. CDK9 is the catalytic subunit of a general RNA polymerase II elongation factor known as positive transcription elongation factor b (P-TEFb). The kinase function of P-TEFb requires phosphorylation of Thr-186 in the T-loop of Cdk9 to allow substrates to access the catalytic core of the enzyme [3].
• Moesin at Thr-588. Both PPM1A and PPM1B dephosphorylated the membrane-F-actin linking protein moesin at Thr-588, which is phosphorylated during activation of platelets [4].
• CDK2 at Thr-160 [5]. Interetingly, yeast has the ortholog of human CDK2, CDC28, but does not have the PPM1A subfamily. The yeast CDC28 was dephosphorylated by PTC2 and PTC3 [6]. PTC2 and PTC3 belong to another PPM subfamily PPM1G2, which is absent from human. However, another member of PPM1G2 subfamily in yeast, PTC4 has little activity towards CDC28 [6]. In general, the PPM1A subfamily and PPM1G2 subfamily are functional orthologs in the aspect of their phosphatase activity towards human CDK2 at Thr-160 and yeast CDC28 at Thr-169.

Different functions of PPM1A and PPM1B

PPM1A (PP2Cα)

Human PPM1A is widely expressed across different tissues, according to RNA-seq data from GTEx. Below are the substrates reported for PPM1A but not PPM1B, but we cannot rule out PPM1B may dephosphorylate some of them:

• mGluR3 at Ser-845. mGluR3 is a metabotropic glutamate receptor 3. It interacted with PPM1A through a C-terminal region from 836 to 855. The region was specific for mGluR3 and was not found in mGluR2, which was its cloest gene in sequence. The region contained Ser-845 phosphorylated by PKA. As PPM1A dephosphorylated the region in vitro, it was probably dephosphorylated Ser-845 [7].
• RelA at Ser-536 and Ser-276. PPM1A directly dephosphorylated RelA at the two positions and selectively inhibited NF-kappaB pathway [8].
• p85 subunit of PI3K at Ser-608 [9].
• p38. PPM1A directly interacted with and dephosphorylated MAPK p38, classfied as CMGC:MAPK:p38 in Kinbase [10].
• MKK6. PPM1A dephosphorylated MAPKK MKK6, classified as STE:STE7:MEK3 in KinBase [10].
• SEK1. PPM1A dephosphorylated MAPKK SEK1, classified as STE:STE7:MEK4 in KinBase [10].
• Axin. PPM1A formed a complex with Dvl, beta-catenin, and Axin. It dephosphorylated Axin, a negative regulator of WNT signaling, both in vitro and in vivo. The dephosphorylation resulted in decreasing Axin's half-life. PPM1A therefore is a positive regulator of WNT signal [11].
• PKCdelta. In addition to PPM1A, PP1 and PP2A (PP3) can also dephosphorylate PKCdelta, albeit lower specific activity [12].
• SMAD1 and SMAD2/3 at SxS motif [13, 14]. SMADs are critical players in TGF-beta signaling. The dephosphorylation increases their nuclear exporter activity.
• RanBP3, nuclear exporter. PPM1A directly interacted with and dephosphorylated RanBP3 at Ser-58 both in vitro and in vivo. The dephosphorylation promoted RanBP3's ability to export Smad2/3 and terminate TGF-beta signaling [15].

In addition, PPM1A regulated cell cycle [16].

PPM1B (PP2Cβ)

PPM1B is expressed in a broad types of tissues, most abundantly in skeletal muscle as shown by RNA-seq data from GTEx and northern blot [17]. Below are the substrates reported for PPM1A but not PPM1B, though PPM1B may dephosphorylate some of them:

• TAK1 at Ser-192. PPM1B but not PPM1A associated with and dephosphorylated TAK1 in vitro [18]. TAK1 is a player in SAPK/JNK pathway, upstream of MKK4/MKK6 and JNK. It belongs to TKL group, MLK family and TAK1 subfamily according the KinBase classification. It emerged in holozoa. In the same study, it was shown that PPM1B did not dephosphorylate MKK6 and did not associate with MEKK3, MKK4, MKK6, JNK, or p38, which was dephosphorylated and inactivated by PPM1A [18]. TAK1 can be also dephosphorylated by another PPM, PPM1L.
• TBK1 at Ser-172. TBK1 is a kinase activates antiviral response. The dephosphorylation therefore results in negatively regulates antiviral response [19].
• PAX2 [20]. PAX2 is a transcription factor which is activated by the phosphorylation at Ser/Thr sites within its C-terminal activation domain, depending on WNT signals and JNK activity.
• PPARγ (peroxisome-proliferator-activated receptor γ) at Ser-112 [21]. PPARγis a nuclear receptor whose activity is altered by the phosphorylation state of Ser-112 and Ser-273. PPM1B can directly dephosphorylate PPARγ both in intact cells and in vitro. In addition, PPM1B specifically targeted Ser-112 rather than Ser-273, which increase PPARγ activity [21]..

PPM1B interacts with the proteins, but there is no direct evidence that they are the substrates:

• Erythroid Kruppel-like factor, a erythroid-specific transcription factor. Mouse Ppm1b probably has an indirect role in regulating EKLF turnover via its zinc finger domain [22].

PPM1B is phosphorylated by PKA at Ser-195 [23]. The Ser-195 is a signature of PPM1A subfamily, because 1) all PPM1A members have serine at the position, 2) PPMs of other subfamilies only have serine, occasionally.

PPM1B is modified with ISG15 at least through Lys-12 and Lys-142 [24]. ISG15 is an interferon-upregulated ubiquitin-like protein, which is covalently conjugated to various cellular proteins (ISGylation). ISG15 is found mainly in marsupials and placentals. In comparison, Lys-12 is found in all vertebrate PPM1Bs and PPM1As. Lys-142 is only found in primate and eutheria PPM1Bs, and was replaced by Thr in rodent PPM1B, by Asp or Glu even Ser in marsupial, reptile and fish PPM1Bs, by Gln and Ser even His in PPM1A.

Human PPM1N

PPM1N's function is unknown. It is generally expressed at low level across tissues except spleen (see RNA-seq data from GTEx).

Fruit fly alph (alphabet): negatively regulates RAS/MAPK and SAPK/JNK

Alphabet, the PPM1A in fruit fly, negatively regulates RAS/MAPK signaling in Drosophila [25] and stress-activated protein kinase (SAPK) [26]. However, its substrates in these signaling pathways are unclear.

References

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2. Sun W, Yu Y, Dotti G, Shen T, Tan X, Savoldo B, Pass AK, Chu M, Zhang D, Lu X, Fu S, Lin X, and Yang J. PPM1A and PPM1B act as IKKbeta phosphatases to terminate TNFalpha-induced IKKbeta-NF-kappaB activation. Cell Signal. 2009 Jan;21(1):95-102. DOI:10.1016/j.cellsig.2008.09.012 |
3. Wang Y, Dow EC, Liang YY, Ramakrishnan R, Liu H, Sung TL, Lin X, and Rice AP. Phosphatase PPM1A regulates phosphorylation of Thr-186 in the Cdk9 T-loop. J Biol Chem. 2008 Nov 28;283(48):33578-84. DOI:10.1074/jbc.M807495200 |
4. Hishiya A, Ohnishi M, Tamura S, and Nakamura F. Protein phosphatase 2C inactivates F-actin binding of human platelet moesin. J Biol Chem. 1999 Sep 17;274(38):26705-12. DOI:10.1074/jbc.274.38.26705 |
5. Cheng A, Kaldis P, and Solomon MJ. Dephosphorylation of human cyclin-dependent kinases by protein phosphatase type 2C alpha and beta 2 isoforms. J Biol Chem. 2000 Nov 3;275(44):34744-9. DOI:10.1074/jbc.M006210200 |
6. Cheng A, Ross KE, Kaldis P, and Solomon MJ. Dephosphorylation of cyclin-dependent kinases by type 2C protein phosphatases. Genes Dev. 1999 Nov 15;13(22):2946-57. DOI:10.1101/gad.13.22.2946 |
7. Flajolet M, Rakhilin S, Wang H, Starkova N, Nuangchamnong N, Nairn AC, and Greengard P. Protein phosphatase 2C binds selectively to and dephosphorylates metabotropic glutamate receptor 3. Proc Natl Acad Sci U S A. 2003 Dec 23;100(26):16006-11. DOI:10.1073/pnas.2136600100 |
8. Lu X, An H, Jin R, Zou M, Guo Y, Su PF, Liu D, Shyr Y, and Yarbrough WG. PPM1A is a RelA phosphatase with tumor suppressor-like activity. Oncogene. 2014 May 29;33(22):2918-27. DOI:10.1038/onc.2013.246 |
9. Yoshizaki T, Maegawa H, Egawa K, Ugi S, Nishio Y, Imamura T, Kobayashi T, Tamura S, Olefsky JM, and Kashiwagi A. Protein phosphatase-2C alpha as a positive regulator of insulin sensitivity through direct activation of phosphatidylinositol 3-kinase in 3T3-L1 adipocytes. J Biol Chem. 2004 May 21;279(21):22715-26. DOI:10.1074/jbc.M313745200 |
10. Takekawa M, Maeda T, and Saito H. Protein phosphatase 2Calpha inhibits the human stress-responsive p38 and JNK MAPK pathways. EMBO J. 1998 Aug 17;17(16):4744-52. DOI:10.1093/emboj/17.16.4744 |
11. Strovel ET, Wu D, and Sussman DJ. Protein phosphatase 2Calpha dephosphorylates axin and activates LEF-1-dependent transcription. J Biol Chem. 2000 Jan 28;275(4):2399-403. DOI:10.1074/jbc.275.4.2399 |
12. Srivastava J, Goris J, Dilworth SM, and Parker PJ. Dephosphorylation of PKCdelta by protein phosphatase 2Ac and its inhibition by nucleotides. FEBS Lett. 2002 Apr 10;516(1-3):265-9. DOI:10.1016/s0014-5793(02)02500-0 |
13. Lin X, Duan X, Liang YY, Su Y, Wrighton KH, Long J, Hu M, Davis CM, Wang J, Brunicardi FC, Shi Y, Chen YG, Meng A, and Feng XH. PPM1A functions as a Smad phosphatase to terminate TGFbeta signaling. Cell. 2006 Jun 2;125(5):915-28. DOI:10.1016/j.cell.2006.03.044 |
14. Duan X, Liang YY, Feng XH, and Lin X. Protein serine/threonine phosphatase PPM1A dephosphorylates Smad1 in the bone morphogenetic protein signaling pathway. J Biol Chem. 2006 Dec 1;281(48):36526-32. DOI:10.1074/jbc.M605169200 |
15. Dai F, Shen T, Li Z, Lin X, and Feng XH. PPM1A dephosphorylates RanBP3 to enable efficient nuclear export of Smad2 and Smad3. EMBO Rep. 2011 Oct 28;12(11):1175-81. DOI:10.1038/embor.2011.174 |
16. Ofek P, Ben-Meir D, Kariv-Inbal Z, Oren M, and Lavi S. Cell cycle regulation and p53 activation by protein phosphatase 2C alpha. J Biol Chem. 2003 Apr 18;278(16):14299-305. DOI:10.1074/jbc.M211699200 |
18. Hanada M, Ninomiya-Tsuji J, Komaki K, Ohnishi M, Katsura K, Kanamaru R, Matsumoto K, and Tamura S. Regulation of the TAK1 signaling pathway by protein phosphatase 2C. J Biol Chem. 2001 Feb 23;276(8):5753-9. DOI:10.1074/jbc.M007773200 |
19. Zhao Y, Liang L, Fan Y, Sun S, An L, Shi Z, Cheng J, Jia W, Sun W, Mori-Akiyama Y, Zhang H, Fu S, and Yang J. PPM1B negatively regulates antiviral response via dephosphorylating TBK1. Cell Signal. 2012 Nov;24(11):2197-204. DOI:10.1016/j.cellsig.2012.06.017 |
20. Abraham S, Paknikar R, Bhumbra S, Luan D, Garg R, Dressler GR, and Patel SR. The Groucho-associated phosphatase PPM1B displaces Pax transactivation domain interacting protein (PTIP) to switch the transcription factor Pax2 from a transcriptional activator to a repressor. J Biol Chem. 2015 Mar 13;290(11):7185-94. DOI:10.1074/jbc.M114.607424 |
21. Tasdelen I, van Beekum O, Gorbenko O, Fleskens V, van den Broek NJ, Koppen A, Hamers N, Berger R, Coffer PJ, Brenkman AB, and Kalkhoven E. The serine/threonine phosphatase PPM1B (PP2Cβ) selectively modulates PPARγ activity. Biochem J. 2013 Apr 1;451(1):45-53. DOI:10.1042/BJ20121113 |
22. Yien YY and Bieker JJ. Functional interactions between erythroid Krüppel-like factor (EKLF/KLF1) and protein phosphatase PPM1B/PP2Cβ. J Biol Chem. 2012 May 4;287(19):15193-204. DOI:10.1074/jbc.M112.350496 |
23. Choi HK, Park SY, Oh HJ, Han EJ, Lee YH, Lee J, Jun WJ, Choi KC, and Yoon HG. PKA negatively regulates PP2Cβ to activate NF-κB-mediated inflammatory signaling. Biochem Biophys Res Commun. 2013 Jul 5;436(3):473-7. DOI:10.1016/j.bbrc.2013.05.129 |
24. Takeuchi T, Kobayashi T, Tamura S, and Yokosawa H. Negative regulation of protein phosphatase 2Cbeta by ISG15 conjugation. FEBS Lett. 2006 Aug 7;580(18):4521-6. DOI:10.1016/j.febslet.2006.07.032 |
25. Baril C and Therrien M. Alphabet, a Ser/Thr phosphatase of the protein phosphatase 2C family, negatively regulates RAS/MAPK signaling in Drosophila. Dev Biol. 2006 Jun 1;294(1):232-45. DOI:10.1016/j.ydbio.2006.02.046 |
25. Baril C, Sahmi M, Ashton-Beaucage D, Stronach B, and Therrien M. The PP2C Alphabet is a negative regulator of stress-activated protein kinase signaling in Drosophila. Genetics. 2009 Feb;181(2):567-79. DOI:10.1534/genetics.108.096461 |
27. Marley AE, Kline A, Crabtree G, Sullivan JE, and Beri RK. The cloning expression and tissue distribution of human PP2Cbeta. FEBS Lett. 1998 Jul 10;431(1):121-4. DOI:10.1016/s0014-5793(98)00708-x |