Phosphatidate Phosphatase-1 is Functionally Conserved in Lipid Synthesis and Storage from Human to Yeast

Abstract

Phosphatidate phosphatase-1 (PAP1) enzymes (yeast Pah1p/Smp2p, mammalian lipin1-3) have a key role in lipid homeostasis by controlling the relative proportions of its substrate phosphatidate (PA) and its product diacylglycerol (DAG). Recent investigation shows that mammalian lipin-1 complements phenotypes exhibited by yeast pah1Δ mutant cells, which indicates the functions of PAP1 enzymes are evolutionarily conserved. The observation was confirmed after transformation of human LPIN1 into PAH1-defective yeast, which resulted in human LPIN1-induced accumulation of triacylglycerol (TAG) and lipid droplet formation. In double mutants lacking Tgl3p and Tgl4p, overexpression of PAH1 or LPIN1-induced TAG accumulation and excessive obesity. Furthermore, the obese yeast was used as a model to study the anti-obesity effects of PAP1 activity inhibitors, including propranolol and clenbuterol. The data showed that the inhibitors significantly suppressed TAG accumulation and lipid droplets formation. These findings demonstrate that LPIN1 plays a functional role in lipid synthesis and storage, a role which is highly conserved from human to yeast. Inhibition of TAG synthesis will become an efficacious treatment strategy for obesity and our excessive obesity model will provide a very useful tool for discovery of new anti-obesity drugs in the future.

Abbreviations

ATGL:

Adipose triglyceride lipase

DAG:

Diacylglycerol

DIC:

Differential interference contrast microscopy

HSL:

Hormone-sensitive lipase

PA:

Phosphatidate

PAP:

Phosphatidate phosphatase

TAG:

Triglyceride

TLC:

Thin-layer chromatography

WT:

Wild type

References

  1. 1.

    Adeyo, O., Horn, P. J., Lee, S. K., Binns, D. D., Chandrahas, A., Chapman, K. D., Goodman, J. M. (2011) The yeast lipin orthologue Pah1p is important for biogenesis of lipid droplets. J. Cell Biol. 192, 1043–1055.

    CAS  Article  Google Scholar 

  2. 2.

    Bilyk, A., Piazza, G., Bistline, R., Jr., Haas, M. (1991) Separation of cholesterol, and fatty acylglycerols, acids and amides by thin-layer chromatography. Lipids 26, 405–406.

    CAS  Article  Google Scholar 

  3. 3.

    Brachmann, C. B., Davies, A., Cost, G. J. Caputo, E., Li, J., Hieter, P., Boeke, J. D. (1998) Designer deletion strains derived from Saccharomyces cerevisiae S288C: a useful set of strains and plasmids for PCR-mediated gene disruption and other applications. Yeast 14, 115–132.

    CAS  Article  Google Scholar 

  4. 4.

    Carman, G. M., Han, G. S. (2006) Roles of phosphatidate phosphatase enzymes in lipid metabolism. Trends Biochem. Sci. 31, 694–699.

    CAS  Article  Google Scholar 

  5. 5.

    Choi, H. S., Su, W. M., Han, G. S., Plote, D., Xu, Z., Carman, G. M. (2012) Pho85p-Pho80p phosphorylation of yeast Pah1p phosphatidate phosphatase regulates its activity, location, abundance, and function in lipid metabolism. J. Biol. Chem. 287, 11290–11301.

    CAS  Article  Google Scholar 

  6. 6.

    Csaki, L. S., Reue, K. (2010) Lipins: multifunctional lipid metabolism proteins. Ann. Rev. Nutr. 30, 257–272.

    CAS  Article  Google Scholar 

  7. 7.

    Gietz, R. D., Akio, S. (1988) New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites. Gene 74, 527–534.

    CAS  Article  Google Scholar 

  8. 8.

    Grimsey, N., Han, G. S., O’hara, L., Rochford, J. J., Carman, G. M., Siniossoglou, S. (2008) Temporal and spatial regulation of the phosphatidate phosphatases lipins 1 and 2. J. Biol. Chem. 283, 29166–29174.

    CAS  Article  Google Scholar 

  9. 9.

    Guo, Y., Cordes, K. R., Farese, R. V., Jr., Walther, T. C. (2009) Lipid droplets at a glance. J. Cell Sci. 122, 749–752.

    CAS  Article  Google Scholar 

  10. 10.

    Huang, Z., Sucgang, R. S., Lin, Y. Y., Shi, X., Boeke, J. D., Pan, X. (2008) Plasmid-chromosome shuffling for non-deletion alleles in yeast. Nat. Meth. 5, 167–169.

    CAS  Article  Google Scholar 

  11. 11.

    Kitazono, A. A. (2009) Improved gap-repair cloning method that uses oligonucleotides to target cognate sequences. Yeast 26, 497–505.

    CAS  Article  Google Scholar 

  12. 12.

    Kohlwein, S. D. (2010) Triacylglycerol homeostasis: insights from yeast. J. Biol. Chem. 285, 15663–15667.

    CAS  Article  Google Scholar 

  13. 13.

    Kurat, C. F., Natter, K., Petschnigg, J., Wolinski, H., Scheuringer, K., Scholz, H., Zimmermann, R., Leber, R., Zechner, R., Kohlwein, S. D. (2006) Obese yeast: triglyceride lipolysis is functionally conserved from mammals to yeast. J. Biol. Chem. 281, 491–500.

    CAS  Article  Google Scholar 

  14. 14.

    Nanjundan, M., Possmayer, F. (2003) Pulmonary phosphatidic acid phosphatase and lipid phosphate phosphohydrolase. Am. J. Physiol. Lung Cell. Mol. Physiol. 284, L1–L23.

  15. 15.

    O’hara, L., Han, G. S., Peak-Chew, S., Grimsey, N., Carman, G. M., Siniossoglou, S. (2006) Control of phospholipid synthesis by phosphorylation of the yeast lipin Pah1p/Smp2p Mg2+-dependent phosphatidate phosphatase. J. Biol. Chem. 281, 34537–34548.

    Article  Google Scholar 

  16. 16.

    Phan, J., Reue, K. (2005) Lipin, a lipodystrophy and obesity gene. Cell Metab. 1, 73–83.

    CAS  Article  Google Scholar 

  17. 17.

    Pyne, S., Long, J. S., Ktistakis, N. T., Pyne, N. J. (2005) Lipid phosphate phosphatases and lipid phosphate signalling. Biochem. Soc. Trans. 33, 1370–1374.

    CAS  Article  Google Scholar 

  18. 18.

    Reue, K., Dwyer, J. R. (2009) Lipin proteins and metabolic homeostasis. J. Lipid Res. 50 Suppl, S109–S114.

    Article  Google Scholar 

  19. 19.

    Santos-Rosa, H., Leung, J., Grimsey, N., Peak-Chew, S., Siniossoglou, S. (2005) The yeast lipin Smp2 couples phospholipid biosynthesis to nuclear membrane growth. EMBO J. 24, 1931–1941.

    CAS  Article  Google Scholar 

  20. 20.

    Sasser, T., Qiu, Q. S., Karunakaran, S., Padolina, M., Reyes, A., Flood, B., Smith, S., Gonzales, C., Fratti, R. A. (2012) Yeast lipin 1 orthologue pah1p regulates vacuole homeostasis and membrane fusion. J. Biol. Chem. 287, 2221–2236.

    CAS  Article  Google Scholar 

  21. 21.

    Schneiter, R., Daum, G. (2006) Extraction of yeast lipids. Methods Mol. Biol. 313, 41–45.

    CAS  PubMed  Google Scholar 

  22. 22.

    Sciorra, V. A., Morris, A. J. (2002) Roles for lipid phosphate phosphatases in regulation of cellular signaling. Biochim. Biophys. Acta 1582, 45–51.

    CAS  Article  Google Scholar 

  23. 23.

    Shahnazari, S., Yen, W. L., Birmingham, C. L., Shiu, J., Namolovan, A., Zheng, Y. T., Nakayama, K., Klionsky, D. J., Brumell, J. H. (2010) A diacylglycerol-dependent signaling pathway contributes to regulation of antibacterial autophagy. Cell Host Microbe 8, 137–146.

    CAS  Article  Google Scholar 

  24. 24.

    Siniossoglou, S. (2013) Phospholipid metabolism and nuclear function: roles of the lipin family of phosphatidic acid phosphatases. Biochim. Biophys. Acta 1831, 575–581.

    CAS  Article  Google Scholar 

  25. 25.

    Skinner, J. R., Shew, T. M., Schwartz, D. M., Tzekov, A., Lepus, C. M., Abumrad, N. A., Wolins, N. E. (2009) Diacylglycerol enrichment of endoplasmic reticulum or lipid droplets recruits perilipin 3/TIP47 during lipid storage and mobilization. J. Biol. Chem. 284, 30941–30948.

    CAS  Article  Google Scholar 

  26. 26.

    Suviolahti, E., Reue, K., Cantor, R. M., Phan, J., Gentile, M., Naukkarinen, J., Soro-Paavonen, A., Oksanen, L., Kaprio, J., Rissanen, A., Salomaa, V., Kontula, K., Taskinen, M. R., Pajukanta, P., Peltonen, L. (2006) Cross-species analyses implicate Lipin 1 involvement in human glucose metabolism. Hum. Mol. Genet. 15, 377–386.

    CAS  Article  Google Scholar 

  27. 27.

    Sztalryd, C., Xu, G., Dorward, H., Tansey, J. T., Contreras, J. A., Kimmel, A. R., Londos, C. (2003) Perilipin A is essential for the translocation of hormone-sensitive lipase during lipolytic activation. J. Cell Biol. 161, 1093–1103.

    CAS  Article  Google Scholar 

  28. 28.

    Wolinski, H., Kohlwein, S. D. (2008) Microscopic analysis of lipid droplet metabolism and dynamics in yeast. Methods Mol. Biol. 457, 151–163.

    CAS  Article  Google Scholar 

  29. 29.

    Zeharia, A., Shaag, A., Houtkooper, R. H., Hindi, T., De Lonlay, P., Erez, G., Hubert, L., Saada, A., De Keyzer, Y., Eshel, G., Vaz, F. M., Pines, O., Elpeleg, O. (2008) Mutations in LPIN1 cause recurrent acute myoglobinuria in childhood. Am. J. Hum. Genet. 83, 489–494.

    CAS  Article  Google Scholar 

  30. 30.

    Zimmermann, R., Strauss, J. G., Haemmerle, G., Schoiswohl, G., Birner-Gruenberger, R., Riederer, M., Lass, A., Neuberger, G., Eisenhaber, F., Hermetter, A., Zechner, R. (2004) Fat mobilization in adipose tissue is promoted by adipose triglyceride lipase. Science 306, 1383–1386.

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank Dr. Ophry Pines and Orly Elpeleg for their kind gift of plasmid YEp-LPIN1. We are grateful to Dr. Deeksha Vishwamitra for her kind assistance in language proofing. This work was sponsored by grants from National Natural Science Foundation of China (31100549), the Scientific Research Foundation for the Returned Overseas Chinese Scholars, State Education Ministry and Fundamental Research Funds for the Central Universities (2232014A3-03 and 222201313010), the National Special Fund for State Key Laboratory of Bioreactor Engineering (2060204).

Author information

Affiliations

Authors

Corresponding authors

Correspondence to Ping Shi or Zhiwei Huang.

Rights and permissions

This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Fang, Z., Wang, S., Du, X. et al. Phosphatidate Phosphatase-1 is Functionally Conserved in Lipid Synthesis and Storage from Human to Yeast. BIOLOGIA FUTURA 65, 481–492 (2014). https://doi.org/10.1556/ABiol.65.2014.4.11

Download citation

Keywords

  • Phosphatidate phosphatase
  • triacylglycerol
  • PAH1
  • LPIN1
  • obesity yeast model