Biopolym. Cell. 2015; 31(5):351-361.
Genomics, Transcriptomics and Proteomics
Expression of isgylation related genes in regenerating rat liver
1Kuklin A. V., 2Poliezhaieva T. A., 2Zhyryakova I. O., 3Ogryzko V. V., 1Obolenskaya M. Yu.
  1. Institute of Molecular Biology and Genetics, NAS of Ukraine
    150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03680
  2. Educational and Scientific Center "Institute of Biology",
    Taras Shevchenko National University of Kyiv
    64/13, Volodymyrska Str., Kyiv, Ukraine, 01601
  3. CNRS UMR 8126, Universit Paris-Sud 11, Institut Gustave Roussy
    114, rue Edouard Vaillant, Villejuif, France, 94805


Our recent studies have revealed the early up-regulated expression of interferon alpha (IFNα) in the liver, induced by partial hepatectomy. The role of this cytokine of innate immune response in liver regeneration is still controversial. Aim. To analyze expression of canonical interferon-stimulated genes Ube1l, Ube2l6, Trim25, Usp18 and Isg15 during the liver transition from quiescence to proliferation induced by partial hepatectomy, and acute phase response induced by laparotomy. These genes are responsible for posttranslational modification of proteins by ISGylation. The expression of genes encoding TATA binding protein (TBP) and 18S rRNA served as indirect general markers of transcriptional and translational activities. Methods. The abundance of investigated RNAs was assessed in total liver RNA by real time RT–qPCR. Results. Partial hepatecomy induced steady upregulation of the Tbp and 18S rRNA genes expression during 12 hours post-surgery and downregulation or no change in expression of ISGylation-related genes during the first 3 hours followed by slight upregulation at 12 hours. The level of Isg15 transcripts was permanently below that of the control during the prereplicative period. Laparotomy induced a continuous downregulation of Tbp and 18S rRNA expression and early (1–3h) upregulation of ISGylation–related transcripts followed by a sharp drop at 6 hours and slight increase/decrease at 12 hours. The changes in the abundance of Ifnα and ISGylation-related mRNAs were oppositely directed at each stage of the response to partial hepatectomy and laparotomy. Conclusion. We suggest that the expression of ISGylation-related genes does not depend on the expression of Ifnα gene after both surgeries. The indirect indices of transcription and translation as well as the expression of ISGylation-relaled genes are principally different in response to partial hepatectomy and laparotomy and argue for the high specificity of innate immune response.
Keywords: Interferon α, ISGylation, liver regeneration, acute phase response


[1] Obolenskaya MYu, Bernauer H, Tran-Thi T-A, Decker K. Levels of RNA for TNF-α and receptors during the prereplicative period of liver regeneration. Biopolym Cell. 1994;10(5):68–77.
[2] Decker KF, Obolenskaya MY. Cytokines, nitric oxide synthesis and liver regeneration. J Gastroenterol Hepatol. 1995;10 Suppl 1:S12-7.
[3] Obolenskaya MYu. Cytokines and liver regeneration. EOS. 1997; 17(2):51–8.
[4] Obolenskaya MYu. Signalling molecules in regenerating liver. Biopolym Cell. 1998;14(3):210–22.
[5] Akerman P, Cote P, Yang SQ, McClain C, Nelson S, Bagby GJ, Diehl AM. Antibodies to tumor necrosis factor-alpha inhibit liver regeneration after partial hepatectomy. Am J Physiol. 1992;263(4 Pt 1):G579-85.
[6] Cressman DE, Greenbaum LE, DeAngelis RA, Ciliberto G, Furth EE, Poli V, Taub R. Liver failure and defective hepatocyte regeneration in interleukin-6-deficient mice. Science. 1996;274(5291):1379-83.
[7] Yamada Y, Kirillova I, Peschon JJ, Fausto N. Initiation of liver growth by tumor necrosis factor: deficient liver regeneration in mice lacking type I tumor necrosis factor receptor. Proc Natl Acad Sci U S A. 1997;94(4):1441-6.
[8] Fujita J, Marino MW, Wada H, Jungbluth AA, Mackrell PJ, Rivadeneira DE, Stapleton PP, Daly JM. Effect of TNF gene depletion on liver regeneration after partial hepatectomy in mice. Surgery. 2001;129(1):48-54.
[9] Markiewski MM, DeAngelis RA, Lambris JD. Liver inflammation and regeneration: two distinct biological phenomena or parallel pathophysiologic processes? Mol Immunol. 2006;43(1-2):45-56.
[10] Hosoya S, Ikejima K, Takeda K, Arai K, Ishikawa S, Yamagata H, Aoyama T, Kon K, Yamashina S, Watanabe S. Innate immune responses involving natural killer and natural killer T cells promote liver regeneration after partial hepatectomy in mice. Am J Physiol Gastrointest Liver Physiol. 2013;304(3):G293-9.
[11] Iimuro Y, Fujimoto J. TLRs, NF-κB, JNK, and Liver Regeneration. Gastroenterol Res Pract. 2010;2010. pii: 598109.
[12] Michalopoulos GK, DeFrances M. Liver regeneration. Adv Biochem Eng Biotechnol. 2005;93:101-34.
[13] Riehle KJ, Dan YY, Campbell JS, Fausto N. New concepts in liver regeneration. J Gastroenterol Hepatol. 2011;26 Suppl 1:203-12.
[14] Fausto N, Campbell JS, Riehle KJ. Liver regeneration. J Hepatol. 2012;57(3):692-4.
[15] Perepelyuk MM, Fedorchenko DB, Rybalko SL, Obolenskaya MYu. Interferon α expression in the rat liver after partial hepatectomy. Biopolym Cell. 2006;22(4):276–82.
[16] Perepelyuk MM, Kuklin AV, Shcherba IaV, Tokovenko BT, Makogon NV, Gogler A, Szala S, Obolenskaya MYu. Interferon α and protein kinase R during rat liver restoration after partial hepatectomy. Biopolym Cell. 2009;25(2):145–9.
[17] Han H, Zhang L, Xu C. Effects of the pathogen infection-related genes on rat liver regeneration following 2. 3 hepatectomy. Life Sci J. 2007; 4(2):50–5.
[18] Chen X, Xu C, Zhang F, Ma J. Microarray approach reveals the relevance of interferon signaling pathways with rat liver restoration post 2/3 hepatectomy at cellular level. J Interferon Cytokine Res. 2010;30(7):525-39.
[19] Batusic DS, von Bargen A, Blaschke S, Dudas J, Ramadori G. Different physiology of interferon-α/-γ in models of liver regeneration in the rat. Histochem Cell Biol. 2011;136(2):131-44.
[20] Vaquero J, Riehle KJ, Fausto N, Campbell JS. Liver Regeneration after partial hepatectomy is not impaired in mice with double deficiency of Myd88 and IFNAR genes. Gastroenterol Res Pract. 2011;2011:727403.
[21] Sen GC, Sarkar SN. The interferon-stimulated genes: targets of direct signaling by interferons, double-stranded RNA, and viruses. Curr Top Microbiol Immunol. 2007;316:233-50.
[22] Zhang D, Zhang DE. Interferon-stimulated gene 15 and the protein ISGylation system. J Interferon Cytokine Res. 2011;31(1):119-30.
[23] Jeon YJ, Yoo HM, Chung CH. ISG15 and immune diseases. Biochim Biophys Acta. 2010;1802(5):485-96.
[24] Ciechanover A. Intracellular protein degradation: from a vague idea thru the lysosome and the ubiquitin-proteasome system and onto human diseases and drug targeting. Cell Death Differ. 2005;12(9):1178-90.
[25] Yuan W, Krug RM. Influenza B virus NS1 protein inhibits conjugation of the interferon (IFN)-induced ubiquitin-like ISG15 protein. EMBO J. 2001;20(3):362-71.
[26] Zhao C, Beaudenon SL, Kelley ML, Waddell MB, Yuan W, Schulman BA, Huibregtse JM, Krug RM. The UbcH8 ubiquitin E2 enzyme is also the E2 enzyme for ISG15, an IFN-alpha/beta-induced ubiquitin-like protein. Proc Natl Acad Sci U S A. 2004;101(20):7578-82.
[27] Takeuchi T, Iwahara S, Saeki Y, Sasajima H, Yokosawa H. Link between the ubiquitin conjugation system and the ISG15 conjugation system: ISG15 conjugation to the UbcH6 ubiquitin E2 enzyme. J Biochem. 2005;138(6):711-9.
[28] Durfee LA, Kelley ML, Huibregtse JM. The basis for selective E1-E2 interactions in the ISG15 conjugation system. J Biol Chem. 2008;283(35):23895-902.
[29] Inoue S, Urano T, Ogawa S, Saito T, Orimo A, Hosoi T, Ouchi Y, Muramatsu M. Molecular cloning of rat efp: expression and regulation in primary osteoblasts. Biochem Biophys Res Commun. 1999;261(2):412-8.
[30] Malakhov MP, Malakhova OA, Kim KI, Ritchie KJ, Zhang DE. UBP43 (USP18) specifically removes ISG15 from conjugated proteins. J Biol Chem. 2002;277(12):9976-81.
[31] Higgins GM, Anderson RM. Experimental pathology of the liver I. Restoration of the liver of white rat following partial surgical removal. Arch Pathol. 1931; 12:186–202.
[32] Rio DC, Ares M Jr, Hannon GJ, Nilsen TW. Removal of DNA from RNA. Cold Spring Harb Protoc. 2010;2010(6):pdb.prot5443.
[33] Webber EM, Godowski PJ, Fausto N. In vivo response of hepatocytes to growth factors requires an initial priming stimulus. Hepatology. 1994;19(2):489-97.
[34] Fausto N. Liver regeneration. J Hepatol. 2000;32(1 Suppl):19-31.
[35] Baumann H, Gauldie J. The acute phase response. Immunol Today. 1994;15(2):74-80.
[36] Cray C, Zaias J, Altman NH. Acute phase response in animals: a review. Comp Med. 2009;59(6):517-26.
[37] Basehoar AD, Zanton SJ, Pugh BF. Identification and distinct regulation of yeast TATA box-containing genes. Cell. 2004;116(5):699-709.
[38] Huisinga KL, Pugh BF. A genome-wide housekeeping role for TFIID and a highly regulated stress-related role for SAGA in Saccharomyces cerevisiae. Mol Cell. 2004;13(4):573-85.
[39] Moshonov S, Elfakess R, Golan-Mashiach M, Sinvani H, Dikstein R. Links between core promoter and basic gene features influence gene expression. BMC Genomics. 2008;9:92.
[40] Han HW, Bae SH, Jung YH, Kim JH, Moon J. Genome-wide characterization of the relationship between essential and TATA-containing genes. FEBS Lett. 2013;587(5):444-51.
[41] Obolenskaia MIu, Gerasimova TB, Bilich KM, Platonov OM. [Intracellular distribution of newly formed hepatocyte RNA in the 1st hours following partial hepatectomy]. Tsitol Genet. 1987;21(5):376-82.
[42] Chaudhuri S, Lieberman I. Control of ribosome syntesis in normal and regenerating liver. J Biol Chem. 1968;243(1):29-33.
[43] Dabeva MD, Dudov KP. Transcriptional control of ribosome production in regenerating rat liver. Biochem J. 1982;208(1):101-8.
[44] Huang J, Rudnick DA. Elucidating the metabolic regulation of liver regeneration. Am J Pathol. 2014;184(2):309-21.
[45] Kuklin A, Tokovenko B, Makogon N, Oczko-Wojciechowska M, Jarząb B, Obolenskaya M. Hepatocytes response to interferon alpha levels recorded after liver resection. J Interferon Cytokine Res. 2014;34(2):90-9.
[46] Sen GC, Sarkar SN. The interferon-stimulated genes: targets of direct signaling by interferons, double-stranded RNA, and viruses. Curr Top Microbiol Immunol. 2007;316:233-50.
[47] Shi HX, Yang K, Liu X, Liu XY, Wei B, Shan YF, Zhu LH, Wang C. Positive regulation of interferon regulatory factor 3 activation by Herc5 via ISG15 modification. Mol Cell Biol. 2010;30(10):2424-36.
[48] Feng Q, Sekula D, Guo Y, Liu X, Black CC, Galimberti F, Shah SJ, Sempere LF, Memoli V, Andersen JB, Hassel BA, Dragnev K, Dmitrovsky E. UBE1L causes lung cancer growth suppression by targeting cyclin D1. Mol Cancer Ther. 2008;7(12):3780-8.
[49] Urano T, Saito T, Tsukui T, Fujita M, Hosoi T, Muramatsu M, Ouchi Y, Inoue S. Efp targets 14-3-3 sigma for proteolysis and promotes breast tumour growth. Nature. 2002;417(6891):871-5.
[50] Okumura F, Zou W, Zhang DE. ISG15 modification of the eIF4E cognate 4EHP enhances cap structure-binding activity of 4EHP. Genes Dev. 2007;21(3):255-60.
[51] Zhang X, Bogunovic D, Payelle-Brogard B, Francois-Newton V, Speer SD, Yuan C, Volpi S, Li Z, Sanal O, Mansouri D, Tezcan I, Rice GI, Chen C, Mansouri N, Mahdaviani SA, Itan Y, Boisson B, Okada S, Zeng L, Wang X, Jiang H, Liu W, Han T, Liu D, Ma T, Wang B, Liu M, Liu JY, Wang QK, Yalnizoglu D, Radoshevich L, Uzé G, Gros P, Rozenberg F, Zhang SY, Jouanguy E, Bustamante J, García-Sastre A, Abel L, Lebon P, Notarangelo LD, Crow YJ, Boisson-Dupuis S, Casanova JL, Pellegrini S. Human intracellular ISG15 prevents interferon-α/β over-amplification and auto-inflammation. Nature. 2015;517(7532):89-93.
[52] Schneider WM, Chevillotte MD, Rice CM. Interferon-stimulated genes: a complex web of host defenses. Annu Rev Immunol. 2014;32:513-45.