Tài liệu Báo cáo khoa học: Consequences of COP9 signalosome and 26S proteasome interaction doc

Consequences of COP9 signalosome and 26S proteasome

interaction

Xiaohua Huang1

, Bettina K. J. Hetfeld1

, Ulrike Seifert2

, Thilo Ka¨hne3

, Peter-Michael Kloetzel2

,

Michael Naumann3

, Dawadschargal Bech-Otschir4 and Wolfgang Dubiel1

1 Division of Molecular Biology, Department of Surgery, Charite´, Universita¨tsmedizin Berlin, Germany

2 Institute of Biochemistry, Charite´, Universita¨tsmedizin Berlin, Germany

3 Institut fu¨r Experimentelle Innere Medizin, Universita¨t Magdeburg, Germany

4 MRC Human Genetics Unit, Western General Hospital, Edinburgh, UK

The COP9 signalosome (CSN) has been discovered in

plant cells as a negative regulator of photomorphogen￾esis [1]. It occurs in all eukaryotic cells and consists of

eight core subunits, CSN1–CSN8 [2]. Six of the CSN

subunits contain PCI (proteasome, COP9 signalosome,

initiation factor 3) domains and two contain MPN

(Mpr-Pad1-N-terminal) domains [3]. These two charac￾teristic domains have been found in three protein com￾plexes: the CSN, the 26S proteasome lid complex (lid)

and the eukaryotic translation initiation factor 3

(eIF3) complex. The two domains are composed of

about 150–200 amino acids at the N- or C-terminus of

the CSN subunits. The PCI domain has been demon￾strated to be important for interactions between CSN

subunits. Thus, it might have scaffolding function

[4,5]. The MPN+ or JAMM domain of CSN5 is

responsible for an intrinsic metalloprotease activity of

the complex [6]. The function of the MPN domain of

CSN6 is unknown.

The CSN is associated with a large number of pro￾teins [7], most of which are substrates or regulators

of the ubiquitin (Ub) system. Analysis of associated

Keywords

COP9 signalosome; lid; p53; PCI domain;

26S proteasome

Correspondence

Division of Molecular Biology, Department

of Surgery, Charite´, Universita¨tsmedizin

Berlin, Monbijoustr. 2, 10117 Berlin,

Germany

Fax: +49 30 450522928

Tel: +49 30 450522305

e-mail: [email protected]

(Received 9 May 2005, accepted 6 June

2005)

doi:10.1111/j.1742-4658.2005.04807.x

The COP9 signalosome (CSN) occurs in all eukaryotic cells. It is a regula￾tory particle of the ubiquitin (Ub)⁄ 26S proteasome system. The eight sub￾units of the CSN possess sequence homologies with the polypeptides of the

26S proteasome lid complex and just like the lid, the CSN consists of six

subunits with PCI (proteasome, COP9 signalosome, initiation factor 3)

domains and two components with MPN (Mpr-Pad1-N-terminal) domains.

Here we show that the CSN directly interacts with the 26S proteasome and

competes with the lid, which has consequences for the peptidase activity

of the 26S proteasome in vitro. Flag-CSN2 was permanently expressed in

mouse B8 fibroblasts and Flag pull-down experiments revealed the forma￾tion of an intact Flag-CSN complex, which is associated with the 26S

proteasome. In addition, the Flag pull-downs also precipitated cullins indi￾cating the existence of super-complexes consisting of the CSN, the 26S pro￾teasome and cullin-based Ub ligases. Permanent expression of a chimerical

subunit (Flag-CSN2-Rpn6) consisting of the N-terminal 343 amino acids

of CSN2 and of the PCI domain of S9 ⁄Rpn6, the paralog of CSN2 in the

lid complex, did not lead to the assembly of an intact complex showing

that the PCI domain of CSN2 is important for complex formation. The

consequence of permanent Flag-CSN2 overexpression was de-novo assem￾bly of the CSN complex connected with an accelerated degradation of p53

and stabilization of c-Jun in B8 cells. The possible role of super-complexes

composed of the CSN, the 26S proteasome and of Ub ligases in the regula￾tion of protein stability is discussed.

Abbreviations

CSN, COP9 signalosome; PCI, proteasome-COP9 signalosome-initiation factor 3; MPN, Mpr-Pad1-N-terminal; Ub, ubiquitin.

FEBS Journal 272 (2005) 3909–3917 ª 2005 FEBS 3909