Identification of novel putative regulators of the major apoptotic nuclease DNA Fragmentation Factor

• Authors: Jakub Hanus , Magdalena Kalinowska-Herok , Piotr Widlak
• Languages of publication: EN

Abstracts

EN

Yeast two- and three-hybrid systems were used to screen cDNA libraries from HeLa cells and human brain tissue to identify novel protein partners of DNA Fragmentation Factor, the major apoptotic nuclease. The two-hybrid system revealed the DFF45 inhibitory subunit of the nuclease as the only identified partner of the DFF40 catalytic subunit. Similar analysis revealed several protein candidates that potentially interact with the DFF45 subunit: FBXO28, FOSL1, PGK1, PCNT, FHL1 and GFAP. Recombinant GFAP protected DFF45 against cleavage with caspase-3 and prevented activation of the DFF nuclease in vitro. In addition, three-hybrid system results revealed a putative novel protein partner of the DFF40-DFF45 heterodimer. The candidate cDNA contained two open reading frames that mapped to an intron of the GBF1 gene. Products of the candidate cDNA derived from a cell-free transcription/translation system inhibited DNA cleavage by recombinant caspase-activated DFF. This putative partner of DFF may have functional importance in regulating the apoptotic response because its RNAi silencing facilitated cleavage of the DFF45 inhibitor subunit and affected chromatin fragmentation in HeLa cells undergoing apoptosis. This hypothetical protein, named DRIG based on an acronym specifying its genomic location, could be a novel factor involved in regulation of DFF40 apoptotic nuclease.

Keywords

EN

CAD; apoptotic nuclease; HeLa cells; yeast two-hybrid system; DFF

• Publisher: Polish Biochemical Society
• Journal: Acta Biochimica Polonica
• Year: 2010
• Volume: 57
• Issue: 4
• Pages: 521-527

Dates

published

2010

received

2010-04-02

revised

2010-10-16

accepted

2010-11-22

(unknown)

2010-12-10

Contributors

author

Jakub Hanus

• Center for Translational Research and Molecular Biology of Cancer, Maria Sklodowska-Curie Memorial Cancer Center and Institute of Oncology, Gliwice, Poland

author

Magdalena Kalinowska-Herok

• Center for Translational Research and Molecular Biology of Cancer, Maria Sklodowska-Curie Memorial Cancer Center and Institute of Oncology, Gliwice, Poland

author

Piotr Widlak

• Center for Translational Research and Molecular Biology of Cancer, Maria Sklodowska-Curie Memorial Cancer Center and Institute of Oncology, Gliwice, Poland

References

• Ageichik AV, Samejima K, Kaufmann SH, Earnshaw WC (2007) Genetic analysis of the short splice variant of the inhibitor of caspase-activated DNase (ICAD-S) in chicken DT40 cells. J Biol Chem 282: 27374-27382.
• Ahn JY, Liu X, Cheng D, Peng J, Chan PK, Wade PA, Ye K (2005) Nucleophosmin/B23, a nuclear PI(3,4,5)P(3) receptor, mediates the antiapoptotic actions of NGF by inhibiting CAD. Mol Cell 18: 435-445.
• Ahn JY, Liu X, Liu Z, Pereira L, Cheng D, Peng J, Wade PA, Hamburger AW, Ye K (2006) Nuclear Akt associates with PKC-phosphorylated Ebp1, preventing DNA fragmentation by inhibition of caspase-activated DNase. EMBO J 25: 2083-2095.
• Brenner M, Johnson AB, Boespflug-Tanguy O, Rodriguez D, Goldman JE, Messing A (2001) Mutations in GFAP, encoding glial fibrillary acidic protein, are associated with Alexander disease. Nat Genet 27: 117-120.
• Cho SG, Kim JW, Lee YH, Hwang HS, Kim MS, Ryoo K, Kim MJ, Noh KT, Kim EK, Cho JH, Yoon KW, Cho EG, Park HS, Chi SW, Lee MJ, Kang SS, Ichijo H, Choi EJ (2003) Identification of a novel antiapoptotic protein that antagonizes ASK1 and CAD activities. J Cell Biol 163: 71-81.
• Durrieu F, Samejima K, Fortune JM, Kandels-Lewis S, Osheroff N, Earnshaw WC (2000) DNA topoisomerase IIalpha interacts with CAD nuclease and is involved in chromatin condensation during apoptotic execution. Curr Biol 10: 923-926.
• Enari M, Sakahira H, Yokoyama H, Okawa K, Iwamatsu A, Nagata S (1998) A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD. Nature 391: 43-50.
• Gu J, Dong RP, Zhang C, McLaughlin DF, Wu MX, Schlossman SF (1999) Functional interaction of DFF35 and DFF45 with caspase-activated DNA fragmentation nuclease DFF40. J Biol Chem 274: 20759-20762.
• Halenbeck R, MacDonald H, Roulston A, Chen TT, Conroy L, Williams LT (1998) CPAN, a human nuclease regulated by the caspase-sensitive inhibitor DFF45. Curr Biol 8: 537-540.
• Hanus J, Kalinowska-Herok M, Widłak P (2008) The major apoptotic endonuclease DFF40/CAD is a deoxyribose-specific and double-strand-specific enzyme. Apoptosis 13: 377-382.
• Jin J, Smith FD, Stark C, Wells CD, Fawcett JP, Kulkarni S, Metalnikov P, O'Donnell P, Taylor P, Taylor L, Zougman A, Woodgett JR, Langeberg LK, Scott JD, Pawson T (2004) Proteomic, functional, and domain-based analysis of in vivo 14-3-3 binding proteins involved in cytoskeletal regulation and cellular organization. Curr Biol 14: 1436-1450.
• Kalinowska-Herok M, Widłak P (2008) High Mobility Group proteins stimulate DNA cleavage by apoptotic endonuclease DFF40/CAD due to HMG-box interactions with DNA. Acta Biochim Pol 55: 21-26.
• Kanouchi H, Nishizaki H, Minatogawa Y, Toné S. (2005) Large complex formation of the inhibitor of caspase-activated DNase. Apoptosis 10, 651-656.
• Korn C, Scholz SR, Gimadutdinow O, Lurz R, Pingoud A, Meiss G (2005) Interaction of DNA fragmentation factor (DFF) with DNA reveals an unprecedented mechanism for nuclease inhibition and suggests that DFF can be activated in a DNA-bound state. J Biol Chem 280: 6005-6015.
• Lechardeur D, Drzymala L, Sharma M, Zylka D, Kinach R, Pacia J, Hicks C, Usmani H, Rommens JM, Luckacs GL (2000) Determinants of the nuclear localization of the heterodimeric DNA fragmentation factor (ICAD/CAD). J Cell Biol 150: 321-334.
• Lechardeur D, Dougaparsad S, Nemes C, Lukacs GL (2005) Oligomerization state of the DNA fragmentation factor in normal and apoptotic cells. J Biol Chem 280: 40216-40225.
• Liu X, Li P, Widłak P, Zou H, Luo X, Garrard WT, Wang X (1998) The 40-kDa subunit of DNA fragmentation factor induces DNA fragmentation and chromatin condensation during apoptosis. Proc Natl Acad Sci USA 95: 8461-8466.
• Liu QL, Kishi H, Ohtsuka K, Muraguchi A (2003) Heat shock protein 70 binds caspase-activated DNase and enhances its activity in TCR-stimulated T cells. Blood 102: 1788-1796.
• Liu X, Zou H, Slaughter C, Wang X (1997) DFF, a heterodimeric protein that functions downstream of caspase-3 to trigger DNA fragmentation during apoptosis. Cell 89: 175-184.
• Liu X, Zou H, Widłak P, Garrard WT, Wang X (1999) Activation of the apoptotic endonuclease DFF40 (Caspase-activated DNase or Nuclease): Oligomerization and direct interaction with histone H1. J Biol Chem 274: 13836-13840.
• Lu C, Zhu F, Cho YY, Tang F, Zykova T, Ma WY, Bode AM, Dong Z (2006) Cell apoptosis: requirement of H2AX in DNA ladder formation, but not for the activation of caspase-3. Mol Cell 23: 121-132.
• Lugovskoy AA, Zhou P, Chou JJ, McCarty JS, Li P, Wagner G (1999) Solution structure of the CIDE-N domain of CIDE-B and a model for CIDE-N/CIDE-N interactions in the DNA fragmentation pathway of apoptosis. Cell 99: 747-755.
• Mouser PE, Head E, Ha K-H, Rohn TT (2006) Caspase-mediated cleavage of glial fibillary acidic protein within degenerating astrocytes of the Alzheimer's disease brain. Am J Pathol 168: 936-947.
• Neimanis S, Albig W, Doenecke D, Kahle J (2007) Sequence elements in both subunits of the DNA fragmentation factor are essential for its nuclear transport. J Biol Chem 282: 35821-35830.
• Oberhammer F, Wilson JW, Dive C, Morris ID, Hickman JA, Wakeling AE, Walker PR, Sikorska M (1993) Apoptotic death in epithelial cells: cleavage of DNA to 300 and/or 50 kb fragments prior to or in the absence of internucleosomal fragmentation. EMBO J 12: 3679-3684.
• Quinlan RA, Brenner M, Goldman JE, Messing A (2007) GFAP and its role in Alexander Disease. Exp Cell Res 313: 2077-2087.
• Sakahira H, Nagata S (2002) Co-translational folding of caspase-activated DNase with Hsp70, Hsp40, and inhibitor of caspase-activated DNase. J Biol Chem 277: 3364-3370.
• Samejima K, Earnshaw WC (1998) ICAD/DFF regulator of apoptotic nuclease is nuclear. Exp Cell Res 243: 453-459.
• Samejima K, Earnshaw WC (2000) Differential localization of ICAD-L and ICAD-S in cells due to removal of a C-terminal NLS from ICAD-L by alternative splicing. Exp Cell Res 255: 314-320.
• Slane JM, Lee HS, Vorhees CV, Zhang J, Xu M (2000) DNA fragmentation factor 45 deficient mice exhibit enhanced spatial learning and memory compared to wild-type control mice. Brain Res 867: 70-79.
• Widłak P (2000) DFF40/CAD hypersensitive sites are potentially involved in high molecular weight DNA fragmentation during apoptosis. Cell Mol Biol Lett 5: 373-379.
• Widłak P, Garrard WT (2006a) The apoptotic endonuclease DFF40/CAD is inhibited by RNA, heparin and other polyanions. Apoptosis 11: 1331-1337.
• Widłak P, Garrard WT (2006b) Unique features of the apoptotic endonucleaase DFF40/CAD relative to micrococcal nuclease as a structural probe for chromatin. Biochem Cell Biol 84: 405-410.
• Widłak P, Li P, Wang X, Garrard WT (2000) Cleavage preferences of the apoptotic endonuclease DFF40 (Caspase-activated DNase or Nuclease) on naked DNA and chromatin substrates. J Biol Chem 275: 8226-8232.
• Widłak P, Lanuszewska J, Cary RB, Garrard WT (2003) Subunit structures and stoichiometries of human DFF proteins before and after induction of apoptosis. J Biol Chem 278: 26915-26922.
• Widłak P, Kalinowska M, Parseghian MH, Lu X, Hansen JC, Garrard WT (2005) The histone H1 C-terminal domain binds to the apoptotic nuclease, DNA Fragmentation Factor (DFF40/CAD) and stimulates DNA cleavage. Biochemistry 44: 7871-7878.
• Widłak P, Garrard WT (2009) Roles of the major apoptotic nuclease - DNA Fragmentation Factor - in biology and disease. Cell Mol Life Sci 66: 263-274.
• Woo EJ, Kim YG, Kim MS, Han WD, Shin S, Robinson H, Park SY, Oh BH (2004) Structural mechanism for inactivation and activation of CAD/DFF40 in the apoptotic pathway. Mol Cell 14: 531-539.
• Wyllie AH (1980) Glucocorticoid-induced thymocyte apoptosis is associated with endogenous endonuclease activation. Nature 284: 555-556.