Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 2002 Dec 15;368(Pt 3):789–797. doi: 10.1042/BJ20020582

Functional interaction between nuclear inhibitor of protein phosphatase type 1 (NIPP1) and protein phosphatase type 1 (PP1) in Drosophila: consequences of over-expression of NIPP1 in flies and suppression by co-expression of PP1.

Louise Parker 1, Sascha Gross 1, Monique Beullens 1, Mathieu Bollen 1, Daimark Bennett 1, Luke Alphey 1
PMCID: PMC1223049  PMID: 12358598

Abstract

The catalytic subunit of type 1 Ser/Thr protein phosphatases (PP1c) forms complexes with many proteins that target it to particular subcellular locations and regulate its activity towards specific substrates. We report the identification of a Drosophila orthologue of nuclear inhibitor of PP1 (NIPP1Dm) through interaction with PP1c in the yeast two-hybrid system. NIPP1Dm shares many properties with mammalian NIPP1 including inhibition of PP1c in vitro, binding to RNA and PP1c, and localization to nuclear speckles. However, the mechanism controlling interaction of PP1c with NIPP1 is not conserved in Drosophila. NIPP1 can function independently of PP1c as a splicing factor, but the relative importance of this function is unknown. Over-expression of NIPP1Dm in Drosophila is cell-lethal in a range of tissues and developmental stages. The effects of ectopic NIPP1Dm are suppressed by co-expression of PP1c, indicating that the only effect of ectopic NIPP1Dm is to affect PP1c function. Co-expression of NIPP1Dm and PP1c does not have any detectable physiological effect in vivo, suggesting that the NIPP1Dm-PP1c holoenzyme is not normally limiting in Drosophila. These data show that NIPP1Dm and PP1c interact in vivo and suggest that NIPP1's role as a phosphatase regulator is conserved in Drosophila.

Full Text

The Full Text of this article is available as a PDF (418.1 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aggen J. B., Nairn A. C., Chamberlin R. Regulation of protein phosphatase-1. Chem Biol. 2000 Jan;7(1):R13–R23. doi: 10.1016/s1074-5521(00)00069-7. [DOI] [PubMed] [Google Scholar]
  2. Allen P. B., Kwon Y. G., Nairn A. C., Greengard P. Isolation and characterization of PNUTS, a putative protein phosphatase 1 nuclear targeting subunit. J Biol Chem. 1998 Feb 13;273(7):4089–4095. doi: 10.1074/jbc.273.7.4089. [DOI] [PubMed] [Google Scholar]
  3. Alphey L. PCR-based method for isolation of full-length clones and splice variants from cDNA libraries. Biotechniques. 1997 Mar;22(3):481-4, 486. doi: 10.2144/97223st02. [DOI] [PubMed] [Google Scholar]
  4. Alphey L., Parker L., Hawcroft G., Guo Y., Kaiser K., Morgan G. KLP38B: a mitotic kinesin-related protein that binds PP1. J Cell Biol. 1997 Jul 28;138(2):395–409. doi: 10.1083/jcb.138.2.395. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Axton J. M., Dombrádi V., Cohen P. T., Glover D. M. One of the protein phosphatase 1 isoenzymes in Drosophila is essential for mitosis. Cell. 1990 Oct 5;63(1):33–46. doi: 10.1016/0092-8674(90)90286-n. [DOI] [PubMed] [Google Scholar]
  6. Baksa K., Morawietz H., Dombrádi V., Axton M., Taubert H., Szabó G., Török I., Udvardy A., Gyurkovics H., Ször B. Mutations in the protein phosphatase 1 gene at 87B can differentially affect suppression of position-effect variegation and mitosis in Drosophila melanogaster. Genetics. 1993 Sep;135(1):117–125. doi: 10.1093/genetics/135.1.117. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bennett D., Szöor B., Alphey L. The chaperone-like properties of mammalian inhibitor-2 are conserved in a Drosophila homologue. Biochemistry. 1999 Dec 7;38(49):16276–16282. doi: 10.1021/bi9917028. [DOI] [PubMed] [Google Scholar]
  8. Beullens M., Van Eynde A., Bollen M., Stalmans W. Inactivation of nuclear inhibitory polypeptides of protein phosphatase-1 (NIPP-1) by protein kinase A. J Biol Chem. 1993 Jun 25;268(18):13172–13177. [PubMed] [Google Scholar]
  9. Beullens M., Van Eynde A., Stalmans W., Bollen M. The isolation of novel inhibitory polypeptides of protein phosphatase 1 from bovine thymus nuclei. J Biol Chem. 1992 Aug 15;267(23):16538–16544. [PubMed] [Google Scholar]
  10. Beullens M., Van Eynde A., Vulsteke V., Connor J., Shenolikar S., Stalmans W., Bollen M. Molecular determinants of nuclear protein phosphatase-1 regulation by NIPP-1. J Biol Chem. 1999 May 14;274(20):14053–14061. doi: 10.1074/jbc.274.20.14053. [DOI] [PubMed] [Google Scholar]
  11. Beullens M., Vulsteke V., Van Eynde A., Jagiello I., Stalmans W., Bollen M. The C-terminus of NIPP1 (nuclear inhibitor of protein phosphatase-1) contains a novel binding site for protein phosphatase-1 that is controlled by tyrosine phosphorylation and RNA binding. Biochem J. 2000 Dec 15;352(Pt 3):651–658. [PMC free article] [PubMed] [Google Scholar]
  12. Beullens Monique, Bollen Mathieu. The protein phosphatase-1 regulator NIPP1 is also a splicing factor involved in a late step of spliceosome assembly. J Biol Chem. 2002 Mar 21;277(22):19855–19860. doi: 10.1074/jbc.M200847200. [DOI] [PubMed] [Google Scholar]
  13. Bollen M. Combinatorial control of protein phosphatase-1. Trends Biochem Sci. 2001 Jul;26(7):426–431. doi: 10.1016/s0968-0004(01)01836-9. [DOI] [PubMed] [Google Scholar]
  14. Boudrez An, Beullens Monique, Waelkens Etienne, Stalmans Willy, Bollen Mathieu. Phosphorylation-dependent interaction between the splicing factors SAP155 and NIPP1. J Biol Chem. 2002 Jun 24;277(35):31834–31841. doi: 10.1074/jbc.M204427200. [DOI] [PubMed] [Google Scholar]
  15. Brand A. H., Manoukian A. S., Perrimon N. Ectopic expression in Drosophila. Methods Cell Biol. 1994;44:635–654. doi: 10.1016/s0091-679x(08)60936-x. [DOI] [PubMed] [Google Scholar]
  16. Ceulemans Hugo, Stalmans Willy, Bollen Mathieu. Regulator-driven functional diversification of protein phosphatase-1 in eukaryotic evolution. Bioessays. 2002 Apr;24(4):371–381. doi: 10.1002/bies.10069. [DOI] [PubMed] [Google Scholar]
  17. Claverie-Martin F., Wang M., Cohen S. N. ARD-1 cDNA from human cells encodes a site-specific single-strand endoribonuclease that functionally resembles Escherichia coli RNase E. J Biol Chem. 1997 May 23;272(21):13823–13828. doi: 10.1074/jbc.272.21.13823. [DOI] [PubMed] [Google Scholar]
  18. Cohen Patricia T. W. Protein phosphatase 1--targeted in many directions. J Cell Sci. 2002 Jan 15;115(Pt 2):241–256. doi: 10.1242/jcs.115.2.241. [DOI] [PubMed] [Google Scholar]
  19. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Doonan J. H., Morris N. R. The bimG gene of Aspergillus nidulans, required for completion of anaphase, encodes a homolog of mammalian phosphoprotein phosphatase 1. Cell. 1989 Jun 16;57(6):987–996. doi: 10.1016/0092-8674(89)90337-1. [DOI] [PubMed] [Google Scholar]
  21. Egloff M. P., Johnson D. F., Moorhead G., Cohen P. T., Cohen P., Barford D. Structural basis for the recognition of regulatory subunits by the catalytic subunit of protein phosphatase 1. EMBO J. 1997 Apr 15;16(8):1876–1887. doi: 10.1093/emboj/16.8.1876. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Fernandez A., Brautigan D. L., Lamb N. J. Protein phosphatase type 1 in mammalian cell mitosis: chromosomal localization and involvement in mitotic exit. J Cell Biol. 1992 Mar;116(6):1421–1430. doi: 10.1083/jcb.116.6.1421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Fields S., Sternglanz R. The two-hybrid system: an assay for protein-protein interactions. Trends Genet. 1994 Aug;10(8):286–292. doi: 10.1016/0168-9525(90)90012-u. [DOI] [PubMed] [Google Scholar]
  24. Hofmann K., Bucher P. The FHA domain: a putative nuclear signalling domain found in protein kinases and transcription factors. Trends Biochem Sci. 1995 Sep;20(9):347–349. doi: 10.1016/s0968-0004(00)89072-6. [DOI] [PubMed] [Google Scholar]
  25. Jagiello I., Beullens M., Stalmans W., Bollen M. Subunit structure and regulation of protein phosphatase-1 in rat liver nuclei. J Biol Chem. 1995 Jul 21;270(29):17257–17263. doi: 10.1074/jbc.270.29.17257. [DOI] [PubMed] [Google Scholar]
  26. Jagiello I., Beullens M., Vulsteke V., Wera S., Sohlberg B., Stalmans W., von Gabain A., Bollen M. NIPP-1, a nuclear inhibitory subunit of protein phosphatase-1, has RNA-binding properties. J Biol Chem. 1997 Aug 29;272(35):22067–22071. doi: 10.1074/jbc.272.35.22067. [DOI] [PubMed] [Google Scholar]
  27. Jagiello I., Van Eynde A., Vulsteke V., Beullens M., Boudrez A., Keppens S., Stalmans W., Bollen M. Nuclear and subnuclear targeting sequences of the protein phosphatase-1 regulator NIPP1. J Cell Sci. 2000 Nov;113(Pt 21):3761–3768. doi: 10.1242/jcs.113.21.3761. [DOI] [PubMed] [Google Scholar]
  28. Jin Q., Beullens M., Jagiello I., Van Eynde A., Vulsteke V., Stalmans W., Bollen M. Mapping of the RNA-binding and endoribonuclease domains of NIPP1, a nuclear targeting subunit of protein phosphatase 1. Biochem J. 1999 Aug 15;342(Pt 1):13–19. [PMC free article] [PubMed] [Google Scholar]
  29. Kreivi J. P., Trinkle-Mulcahy L., Lyon C. E., Morrice N. A., Cohen P., Lamond A. I. Purification and characterisation of p99, a nuclear modulator of protein phosphatase 1 activity. FEBS Lett. 1997 Dec 22;420(1):57–62. doi: 10.1016/s0014-5793(97)01485-3. [DOI] [PubMed] [Google Scholar]
  30. Li J., Lee G. I., Van Doren S. R., Walker J. C. The FHA domain mediates phosphoprotein interactions. J Cell Sci. 2000 Dec;113(Pt 23):4143–4149. doi: 10.1242/jcs.113.23.4143. [DOI] [PubMed] [Google Scholar]
  31. Lin Q., Buckler E. S., 4th, Muse S. V., Walker J. C. Molecular evolution of type 1 serine/threonine protein phosphatases. Mol Phylogenet Evol. 1999 Jun;12(1):57–66. doi: 10.1006/mpev.1998.0560. [DOI] [PubMed] [Google Scholar]
  32. Mermoud J. E., Cohen P. T., Lamond A. I. Regulation of mammalian spliceosome assembly by a protein phosphorylation mechanism. EMBO J. 1994 Dec 1;13(23):5679–5688. doi: 10.1002/j.1460-2075.1994.tb06906.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mermoud J. E., Cohen P., Lamond A. I. Ser/Thr-specific protein phosphatases are required for both catalytic steps of pre-mRNA splicing. Nucleic Acids Res. 1992 Oct 25;20(20):5263–5269. doi: 10.1093/nar/20.20.5263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Misteli T., Cáceres J. F., Spector D. L. The dynamics of a pre-mRNA splicing factor in living cells. Nature. 1997 May 29;387(6632):523–527. doi: 10.1038/387523a0. [DOI] [PubMed] [Google Scholar]
  35. Ohkura H., Kinoshita N., Miyatani S., Toda T., Yanagida M. The fission yeast dis2+ gene required for chromosome disjoining encodes one of two putative type 1 protein phosphatases. Cell. 1989 Jun 16;57(6):997–1007. doi: 10.1016/0092-8674(89)90338-3. [DOI] [PubMed] [Google Scholar]
  36. Parker L., Gross S., Alphey L. Vectors for the expression of tagged proteins in Drosophila. Biotechniques. 2001 Dec;31(6):1280-2, 1284, 1286. doi: 10.2144/01316st01. [DOI] [PubMed] [Google Scholar]
  37. Trinkle-Mulcahy L., Ajuh P., Prescott A., Claverie-Martin F., Cohen S., Lamond A. I., Cohen P. Nuclear organisation of NIPP1, a regulatory subunit of protein phosphatase 1 that associates with pre-mRNA splicing factors. J Cell Sci. 1999 Jan;112(Pt 2):157–168. doi: 10.1242/jcs.112.2.157. [DOI] [PubMed] [Google Scholar]
  38. Trinkle-Mulcahy L., Sleeman J. E., Lamond A. I. Dynamic targeting of protein phosphatase 1 within the nuclei of living mammalian cells. J Cell Sci. 2001 Dec;114(Pt 23):4219–4228. doi: 10.1242/jcs.114.23.4219. [DOI] [PubMed] [Google Scholar]
  39. Van Eynde A., Pérez-Callejón E., Schoenmakers E., Jacquemin M., Stalmans W., Bollen M. Organization and alternate splice products of the gene encoding nuclear inhibitor of protein phosphatase-1 (NIPP-1). Eur J Biochem. 1999 Apr;261(1):291–300. doi: 10.1046/j.1432-1327.1999.00272.x. [DOI] [PubMed] [Google Scholar]
  40. Van Eynde A., Wera S., Beullens M., Torrekens S., Van Leuven F., Stalmans W., Bollen M. Molecular cloning of NIPP-1, a nuclear inhibitor of protein phosphatase-1, reveals homology with polypeptides involved in RNA processing. J Biol Chem. 1995 Nov 24;270(47):28068–28074. doi: 10.1074/jbc.270.47.28068. [DOI] [PubMed] [Google Scholar]
  41. Vulsteke V., Beullens M., Waelkens E., Stalmans W., Bollen M. Properties and phosphorylation sites of baculovirus-expressed nuclear inhibitor of protein phosphatase-1 (NIPP-1). J Biol Chem. 1997 Dec 26;272(52):32972–32978. doi: 10.1074/jbc.272.52.32972. [DOI] [PubMed] [Google Scholar]
  42. Wang M., Cohen S. N. ard-1: a human gene that reverses the effects of temperature-sensitive and deletion mutations in the Escherichia coli rne gene and encodes an activity producing RNase E-like cleavages. Proc Natl Acad Sci U S A. 1994 Oct 25;91(22):10591–10595. doi: 10.1073/pnas.91.22.10591. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Wera S., Van Eynde A., Stalmans W., Bollen M. Inhibition of translation by mRNA encoding NIPP-1, a nuclear inhibitor of protein phosphatase-1. Eur J Biochem. 1997 Jul 1;247(1):411–415. doi: 10.1111/j.1432-1033.1997.00411.x. [DOI] [PubMed] [Google Scholar]
  44. Yeh E., Gustafson K., Boulianne G. L. Green fluorescent protein as a vital marker and reporter of gene expression in Drosophila. Proc Natl Acad Sci U S A. 1995 Jul 18;92(15):7036–7040. doi: 10.1073/pnas.92.15.7036. [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Multimedia adjunct for figure 5 (small movie)
Download video file (845.5KB, mov)
Multimedia adjunct for figure 5(large movie)
Download video file (844.5KB, mov)

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES