Identification of FBXO25-interacting proteins using an integrated proteomics approach.
FBXO25 is one of the 68 human F-box proteins that serve as specificity factors for a family of ubiquitin ligases composed of s-phase-kinase associated protein 1, really interesting new gene-box 1, Cullin 1, and F-box protein (SCF1) that are involved in targeting proteins for destruction across the ubiquitin proteasome system. We recently reported that the FBXO25 protein accumulates in novel subnuclear structures named FBXO25-associated nuclear domains (FAND). Combining two-step affinity purification followed by MS with a classical two-hybrid screen, we identified 132 novel potential FBXO25 interacting partners. One of the identified proteins, beta-actin, physically interacts through its N-terminus with FBXO25 and is enriched in the FBXO25 nuclear compartments. Inhibitors of actin polymerization promote a significant disruption of FAND, indicating that they are compartments influenced by the organizational state of actin in the nucleus. Furthermore, FBXO25 antibodies interfered with RNA polymerase II transcription in vitro. Our results open new perspectives for the understanding of this novel compartment and its nuclear functions.
Teixeira
Felipe R
FR
Department of Biochemistry and Immunology, University of São Paulo, Ribeirão Preto, Brazil.
Yokoo
Sami
S
Gartner
Carlos A
CA
Manfiolli
Adriana O
AO
Baqui
Munira M A
MM
Assmann
Eliana M
EM
Maragno
Ana Leticia G C
AL
Yu
Huijun
H
de Lanerolle
Primal
P
Kobarg
Jörg
J
Gygi
Steven P
SP
Gomes
Marcelo Damário
MD
D000199:Actins / Q000032:analysis / Q000378:metabolism; D000595:Amino Acid Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D044783:F-Box Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D020543:Proteome / Q000737:chemistry / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*
D000199:Actins; D044783:F-Box Proteins; C510759:FBXO25 protein, human; D009419:Nerve Tissue Proteins; D020543:Proteome
Mol Cell Proteomics. 2006 Jul;5(7):1326-37
16635985
Nat Rev Mol Cell Biol. 2004 Sep;5(9):739-51
15340381
J Am Soc Mass Spectrom. 1994 Nov;5(11):976-89
24226387
J Cell Sci. 1999 Mar;112 ( Pt 6):797-809
10036230
Eur J Hum Genet. 2004 Jul;12(7):551-60
15100712
Biochem Soc Trans. 1996 Aug;24(3):893-6
8878870
J Cell Biol. 1987 Oct;105(4):1473-8
3312229
Nucleic Acids Res. 1997 Sep 1;25(17):3389-402
9254694
Mol Cell. 2002 Dec;10(6):1519-26
12504026
J Cell Biol. 2006 Mar 27;172(7):967-71
16549500
Proc Natl Acad Sci U S A. 2008 Aug 5;105(31):10955-60
18667692
Proc Natl Acad Sci U S A. 2001 Dec 4;98(25):14440-5
11717410
Nat Methods. 2007 Mar;4(3):207-14
17327847
Mol Cell Biol. 2001 Jan;21(1):330-42
11113207
J Biol Chem. 2006 Apr 14;281(15):9869-81
16484223
Nat Rev Mol Cell Biol. 2004 May;5(5):410-5
15122354
J Cell Biol. 2006 Feb 13;172(4):495-6
16476772
Methods Enzymol. 2005;398:143-58
16275326
Nat Struct Mol Biol. 2005 Sep;12(9):742-6
16142228
Mol Cell. 2002 Dec;10(6):1511-8
12504025
J Cell Biol. 2008 Jul 14;182(1):77-88
18606852
Bioessays. 2005 Apr;27(4):388-96
15770676
FEBS Lett. 1987 Mar 23;213(2):316-8
3556584
Mol Biol Cell. 2008 May;19(5):1848-61
18287534
Nat Cell Biol. 2004 Nov;6(11):1094-101
15502823
Int Rev Cell Mol Biol. 2009;272:107-47
19121817
Nature. 1989 Jul 20;340(6230):245-6
2547163
Anal Chem. 1996 Mar 1;68(5):850-8
8779443
Mol Cancer Res. 2006 Oct;4(10):695-707
17050664
Biochim Biophys Acta. 2006 Jun;1760(6):966-72
16714087
Mol Biol Cell. 2005 Nov;16(11):5055-60
16148048
Exp Cell Res. 2005 Feb 15;303(2):229-39
15652338
Nat Rev Cancer. 2008 Jun;8(6):438-49
18500245
Exp Cell Res. 2006 Nov 15;312(19):3796-807
17022973
Cell. 2008 Sep 19;134(6):995-1006
18805092
Annu Rev Biochem. 1998;67:425-79
9759494
Genes Dev. 2004 Nov 1;18(21):2573-80
15520277