Diabetologia

pp 1–15 | Cite as

Both conditional ablation and overexpression of E2 SUMO-conjugating enzyme (UBC9) in mouse pancreatic beta cells result in impaired beta cell function

  • Xiaoyu He
  • Qiaohong Lai
  • Cai Chen
  • Na Li
  • Fei Sun
  • Wenting Huang
  • Shu Zhang
  • Qilin Yu
  • Ping Yang
  • Fei Xiong
  • Zhishui Chen
  • Quan Gong
  • Boxu Ren
  • Jianping Weng
  • Décio L. Eizirik
  • Zhiguang Zhou
  • Cong-Yi Wang
Article

Abstract

Aims/hypothesis

Post-translational attachment of a small ubiquitin-like modifier (SUMO) to the lysine (K) residue(s) of target proteins (SUMOylation) is an evolutionary conserved regulatory mechanism. This modification has previously been demonstrated to be implicated in the control of a remarkably versatile regulatory mechanism of cellular processes. However, the exact regulatory role and biological actions of the E2 SUMO-conjugating enzyme (UBC9)-mediated SUMOylation function in pancreatic beta cells has remained elusive.

Methods

Inducible beta cell-specific Ubc9 (also known as Ube2i) knockout (KO; Ubc9Δbeta) and transgenic (Ubc9Tg) mice were employed to address the impact of SUMOylation on beta cell viability and functionality. Ubc9 deficiency or overexpression was induced at 8 weeks of age using tamoxifen. To study the mechanism involved, we closely examined the regulation of the transcription factor nuclear factor erythroid 2-related factor 2 (NRF2) through SUMOylation in beta cells.

Results

Upon induction of Ubc9 deficiency, Ubc9Δbeta islets exhibited a 3.5-fold higher accumulation of reactive oxygen species (ROS) than Ubc9f/f control islets. Islets from Ubc9Δbeta mice also had decreased insulin content and loss of beta cell mass after tamoxifen treatment. Specifically, at day 45 after Ubc9 deletion only 40% of beta cell mass remained in Ubc9Δbeta mice, while 90% of beta cell mass was lost by day 75. Diabetes onset was noted in some Ubc9Δbeta mice 8 weeks after induction of Ubc9 deficiency and all mice developed diabetes by 10 weeks following tamoxifen treatment. In contrast, Ubc9Tg beta cells displayed an increased antioxidant ability but impaired insulin secretion. Unlike Ubc9Δbeta mice, which spontaneously developed diabetes, Ubc9Tg mice preserved normal non-fasting blood glucose levels without developing diabetes. It was noted that SUMOylation of NRF2 promoted its nuclear expression along with enhanced transcriptional activity, thereby preventing ROS accumulation in beta cells.

Conclusions/interpretation

SUMOylation function is required to protect against oxidative stress in beta cells; this mechanism is, at least in part, carried out by the regulation of NRF2 activity to enhance ROS detoxification. Homeostatic SUMOylation is also likely to be essential for maintaining beta cell functionality.

Keywords

Diabetes Insulin content Insulin secretion NRF2 Oxidative stress Pancreatic beta cell SUMOylation UBC9 

Abbreviation

ARE

Antioxidant response element

ChIP

Chromatin immunoprecipitation

CHX

Cycloheximide

DCFH-DA

2′,7′-Dichlorodihydrofluorescein diacetate

EMSA

Electrophoretic mobility shift assay

HA

Haemagglutinin

KO

Knockout

mu

Mutated

NAC

N-acetylcysteine

NRF2

Nuclear factor erythroid 2-related factor 2

NTg

Tamoxifen-induced Ubc9 transgenic mouse model (controls)

ROS

Reactive oxygen species

SENP1

Sentrin-specific protease 1

STZ

Streptozocin

SUMO

Small ubiquitin-like modifier

UBC9

E2 SUMO-conjugating enzyme

Ubc9Δbeta

Beta cell-specific Ubc9 knockout mouse model

Ubc9CAT-Tg

Inducible Ubc9 transgenic mouse model

Ubc9f/f

Ubc9flox mice

Ubc9Tg

Beta cell-specific Ubc9 transgenic mouse model

WT

Wild-type

Notes

Acknowledgements

We are grateful to D. Melton (Harvard University, Cambridge, MA, USA) for providing the Rip-CreER mice, and Q. Yang (University of Alabama, Birmingham, AL, USA) for providing the pCAG-CAT-X vector.

Data availability

All data generated or analysed during this study are included in this published article (and its ESM files).

Duality of interest

The authors declare that there is no duality of interest associated with this manuscript.

Contribution statement

CYW conceived the project. XH, QL, CC, NL, FS, WH, SZ and PY contributed to the acquisition of data, analysis and interpretation. XH wrote the manuscript. CYW, SZ, QY and FX were involved in designing the strategy to generate and genotype the Ubc9Δbeta and Ubc9Tg mouse lines. SZ, ZC, QG, BR, JW, DLE and ZZ contributed to analysis and interpretation of the data. CYW, DLE and ZZ contributed to the study design and manuscript preparation. All authors were involved in drafting the article or revising it critically, and all authors gave their approval for the final manuscript to be published. CYW is the guarantor of this work and takes full responsibility for the content of the manuscript.

Supplementary material

125_2017_4523_MOESM1_ESM.pdf (553 kb)
ESM (PDF 552 kb)

References

  1. 1.
    Yang P, Hu S, Yang F et al (2014) Sumoylation modulates oxidative stress relevant to the viability and functionality of pancreatic beta cells. Am J Transl Res 6:353–360PubMedPubMedCentralGoogle Scholar
  2. 2.
    Flotho A, Melchior F (2013) Sumoylation: a regulatory protein modification in health and disease. Annu Rev Biochem 82:357–385CrossRefPubMedGoogle Scholar
  3. 3.
    Li M, Guo D, Isales CM et al (2005) SUMO wrestling with type 1 diabetes. J Mol Med (Berlin, Germany) 83:504–513CrossRefGoogle Scholar
  4. 4.
    Hajmrle C, Ferdaoussi M, Plummer G et al (2014) SUMOylation protects against IL-1beta-induced apoptosis in INS-1 832/13 cells and human islets. Am J Phys Endocrinol Metab 307:E664–E673CrossRefGoogle Scholar
  5. 5.
    Pandey D, Chen F, Patel A et al (2011) SUMO1 negatively regulates reactive oxygen species production from NADPH oxidases. Arterioscler Thromb Vasc Biol 31:1634–1642CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Guo D, Han J, Adam BL et al (2005) Proteomic analysis of SUMO4 substrates in HEK293 cells under serum starvation-induced stress. Biochem Biophys Res Commun 337:1308–1318CrossRefPubMedGoogle Scholar
  7. 7.
    Kishi A, Nakamura T, Nishio Y, Maegawa H, Kashiwagi A (2003) Sumoylation of Pdx1 is associated with its nuclear localization and insulin gene activation. Am J Phys Endocrinol Metab 284:E830–E840CrossRefGoogle Scholar
  8. 8.
    Mziaut H, Trajkovski M, Kersting S et al (2006) Synergy of glucose and growth hormone signalling in islet cells through ICA512 and STAT5. Nat Cell Biol 8:435–445CrossRefPubMedGoogle Scholar
  9. 9.
    Shao C, Cobb MH (2009) Sumoylation regulates the transcriptional activity of MafA in pancreatic beta cells. J Biol Chem 284:3117–3124CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Dai XQ, Kolic J, Marchi P, Sipione S, Macdonald PE (2009) SUMOylation regulates Kv2.1 and modulates pancreatic beta-cell excitability. J Cell Sci 122:775–779CrossRefPubMedGoogle Scholar
  11. 11.
    Dai XQ, Plummer G, Casimir M et al (2011) SUMOylation regulates insulin exocytosis downstream of secretory granule docking in rodents and humans. Diabetes 60:838–847CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Aukrust I, Bjorkhaug L, Negahdar M et al (2013) SUMOylation of pancreatic glucokinase regulates its cellular stability and activity. J Biol Chem 288:5951–5962CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Ferdaoussi M, Dai X, Jensen MV et al (2015) Isocitrate-to-SENP1 signaling amplifies insulin secretion and rescues dysfunctional beta cells. J Clin Investig 125:3847–3860CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Rajan S, Torres J, Thompson MS, Philipson LH (2012) SUMO downregulates GLP-1-stimulated cAMP generation and insulin secretion. Am J Phys Endocrinol Metab 302:E714–E723CrossRefGoogle Scholar
  15. 15.
    Rajan S, Dickson LM, Mathew E et al (2015) Chronic hyperglycemia downregulates GLP-1 receptor signaling in pancreatic beta-cells via protein kinase A. Mol Metab 4:265–276CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Ferdaoussi M, Fu J, Dai X et al (2017) SUMOylation and calcium control syntaxin-1A and secretagogin sequestration by tomosyn to regulate insulin exocytosis in human β cells. Sci Rep 7:248CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Chen L, Fan C, Zhang Y et al (2013) Beneficial effects of inhibition of soluble epoxide hydrolase on glucose homeostasis and islet damage in a streptozotocin-induced diabetic mouse model. Prostaglandins & Other Lipid Mediators 104–105:42–48CrossRefGoogle Scholar
  18. 18.
    Cheng J, Song J, He X et al (2016) Loss of Mbd2 protects mice against high fat diet-induced obesity and insulin resistance by regulating the homeostasis of energy storage and expenditure. Diabetes 65:3384–3395CrossRefPubMedGoogle Scholar
  19. 19.
    Yang P, Li M, Guo D et al (2008) Comparative analysis of the islet proteome between NOD/Lt and ALR/Lt mice. Ann N Y Acad Sci 1150:68–71CrossRefPubMedGoogle Scholar
  20. 20.
    He L, Sun F, Wang Y et al (2016) HMGB1 exacerbates bronchiolitis obliterans syndrome via RAGE/NF-kappaB/HPSE signaling to enhance latent TGF-beta release from ECM. Am J Transl Res 8:1971–1984PubMedPubMedCentralGoogle Scholar
  21. 21.
    Han J, Zhong J, Wei W et al (2008) Extracellular high-mobility group box 1 acts as an innate immune mediator to enhance autoimmune progression and diabetes onset in NOD mice. Diabetes 57:2118–2127CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Ran L, Yu Q, Zhang S et al (2015) Cx3cr1 deficiency in mice attenuates hepatic granuloma formation during acute schistosomiasis by enhancing the M2-type polarization of macrophages. Dis Model Mech 8:691–700CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Wang J, Wei Q, Wang CY, Hill WD, Hess DC, Dong Z (2004) Minocycline up-regulates Bcl-2 and protects against cell death in mitochondria. J Biol Chem 279:19948–19954CrossRefPubMedGoogle Scholar
  24. 24.
    Zhang M, Guo Y, Fu H et al (2015) Chop deficiency prevents UUO-induced renal fibrosis by attenuating fibrotic signals originated from Hmgb1/TLR4/NFkappaB/IL-1beta signaling. Cell Death Dis 6:e1847CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Yao Y, Wang Y, Zhang Z et al (2016) Chop deficiency protects mice against bleomycin-induced pulmonary fibrosis by attenuating M2 macrophage production. Mol Ther J Am Soc Gene Ther 24:915–925CrossRefGoogle Scholar
  26. 26.
    Hu S, Zhang Y, Zhang M et al (2015) Aloperine protects mice against ischemia reperfusion (IR)-induced renal injury by regulating PI3K/AKT/mTOR signaling and AP-1 activity. Mol Med.  https://doi.org/10.2119/molmed.2015.00056
  27. 27.
    Liang H, Yin B, Zhang H et al (2008) Blockade of tumor necrosis factor (TNF) receptor type 1-mediated TNF-alpha signaling protected Wistar rats from diet-induced obesity and insulin resistance. Endocrinology 149:2943–2951CrossRefPubMedGoogle Scholar
  28. 28.
    Jakobs A, Koehnke J, Himstedt F et al (2007) Ubc9 fusion-directed SUMOylation (UFDS): a method to analyze function of protein SUMOylation. Nat Methods 4:245–250CrossRefPubMedGoogle Scholar
  29. 29.
    Wei W, Yang P, Pang J et al (2008) A stress-dependent SUMO4 sumoylation of its substrate proteins. Biochem Biophys Res Commun 375:454–459CrossRefPubMedGoogle Scholar
  30. 30.
    Wang CY, Yang P, Li M, Gong F (2009) Characterization of a negative feedback network between SUMO4 expression and NFkappaB transcriptional activity. Biochem Biophys Res Commun 381:477–481CrossRefPubMedGoogle Scholar
  31. 31.
    Pi J, Collins S (2010) Reactive oxygen species and uncoupling protein 2 in pancreatic beta-cell function. Diabetes Obes Metab 12(Suppl 2):141–148CrossRefPubMedGoogle Scholar
  32. 32.
    Lei XG, Vatamaniuk MZ (2011) Two tales of antioxidant enzymes on beta cells and diabetes. Antioxid Redox Signal 14:489–503CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Bhakkiyalakshmi E, Shalini D, Sekar TV, Rajaguru P, Paulmurugan R, Ramkumar KM (2014) Therapeutic potential of pterostilbene against pancreatic beta-cell apoptosis mediated through Nrf2. Br J Pharmacol 171:1747–1757CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Yagishita Y, Fukutomi T, Sugawara A et al (2014) Nrf2 protects pancreatic beta-cells from oxidative and nitrosative stress in diabetic model mice. Diabetes 63:605–618CrossRefPubMedGoogle Scholar
  35. 35.
    Cunha DA, Cito M, Carlsson PO et al (2016) Thrombospondin 1 protects pancreatic beta-cells from lipotoxicity via the PERK-NRF2 pathway. Cell Death Differ 23:1995–2006CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Ramani K, Tomasi ML, Yang H, Ko K, Lu SC (2012) Mechanism and significance of changes in glutamate-cysteine ligase expression during hepatic fibrogenesis. J Biol Chem 287:36341–36355CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Lau A, Tian W, Whitman SA, Zhang DD (2013) The predicted molecular weight of Nrf2: it is what it is not. Antioxid Redox Signal 18:91–93CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Hourihan JM, Moronetti Mazzeo LE, Fernandez-Cardenas LP, Blackwell TK (2016) Cysteine sulfenylation directs IRE-1 to activate the SKN-1/Nrf2 antioxidant response. Mol Cell 63:553–566CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Ngo HKC, Kim DH, Cha YN, Na HK, Surh YJ (2017) Nrf2 mutagenic activation drives hepatocarcinogenesis. Cancer Res 77:4797–4808CrossRefPubMedGoogle Scholar
  40. 40.
    Soares MA, Cohen OD, Low YC et al (2016) Restoration of Nrf2 signaling normalizes the regenerative niche. Diabetes 65:633–646CrossRefPubMedGoogle Scholar
  41. 41.
    Saito T, Ichimura Y, Taguchi K et al (2016) p62/Sqstm1 promotes malignancy of HCV-positive hepatocellular carcinoma through Nrf2-dependent metabolic reprogramming. 7:12030Google Scholar
  42. 42.
    Rabbani PS, Zhou A, Borab ZM et al (2017) Novel lipoproteoplex delivers Keap1 siRNA based gene therapy to accelerate diabetic wound healing. Biomaterials 132:1–15CrossRefPubMedGoogle Scholar
  43. 43.
    Hayes JD, Dinkova-Kostova AT (2014) The Nrf2 regulatory network provides an interface between redox and intermediary metabolism. Trends Biochem Sci 39:199–218CrossRefPubMedGoogle Scholar
  44. 44.
    Itoh K, Wakabayashi N, Katoh Y et al (1999) Keap1 represses nuclear activation of antioxidant responsive elements by Nrf2 through binding to the amino-terminal Neh2 domain. Genes Dev 13:76CrossRefPubMedPubMedCentralGoogle Scholar
  45. 45.
    Uruno A, Furusawa Y, Yagishita Y et al (2013) The Keap1-Nrf2 system prevents onset of diabetes mellitus. Mol Cell Biol 33:2996–3010CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Xiaoyu He
    • 1
  • Qiaohong Lai
    • 1
  • Cai Chen
    • 1
  • Na Li
    • 1
  • Fei Sun
    • 1
  • Wenting Huang
    • 1
  • Shu Zhang
    • 1
  • Qilin Yu
    • 1
  • Ping Yang
    • 1
  • Fei Xiong
    • 1
  • Zhishui Chen
    • 1
  • Quan Gong
    • 2
  • Boxu Ren
    • 2
  • Jianping Weng
    • 3
  • Décio L. Eizirik
    • 4
  • Zhiguang Zhou
    • 5
  • Cong-Yi Wang
    • 1
  1. 1.The Center for Biomedical Research, Key Laboratory of Organ Transplantation, Ministry of Education and Ministry of Health, Tongji Hospital, Tongji Medical CollegeHuazhong University of Science and TechnologyWuhanPeople’s Republic of China
  2. 2.Medical College of Yangtze UniversityJingzhouPeople’s Republic of China
  3. 3.Department of Endocrinology and Metabolism, The Third Affiliated HospitalSun Yat-Sen UniversityGuangzhouPeople’s Republic of China
  4. 4.ULB Center for Diabetes Research, Université Libre de BruxellesBrusselsBelgium
  5. 5.Diabetes Center, The Second Xiangya Hospital, Institute of Metabolism and EndocrinologyCentral South UniversityChangshaPeople’s Republic of China

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