Subcellular localization of proteasomes and their regulatory complexes in mammalian cells

Author:

BROOKS Paul1,FUERTES Graciela2,MURRAY Rachael Z.1,BOSE Suchira1,KNECHT Erwin2,RECHSTEINER Martin C.3,HENDIL Klavs B.4,TANAKA Keiji5,DYSON Julian6,RIVETT A. Jennifer1

Affiliation:

1. Department of Biochemistry, University of Bristol, School of Medical Sciences, Bristol BS8 1TD, U.K.

2. Instituto de Investigaciones Citológicas, Amadeo de Saboya 4, 46010-Valencia, Spain

3. Department of Biochemistry, University of Utah Medical Center, 50 North Medical Drive, Salt Lake City, UT 84132, U.S.A.

4. August Krogh Institute, University of Copenhagen, Universitetsparken 13, DK-2100, Copenhagen O, Denmark

5. The Tokyo Metropolitan Institute of Medical Science 18-22, Honkomagome 3-chome, Bunkyo-ku, Tokyo 113, Japan

6. MRC Clinical Sciences Centre, Royal Postgraduate Medical School, Hammersmith Hospital, DuCane Road, London W12 0NN, U.K.

Abstract

Proteasomes can exist in several different molecular forms in mammalian cells. The core 20S proteasome, containing the proteolytic sites, binds regulatory complexes at the ends of its cylindrical structure. Together with two 19S ATPase regulatory complexes it forms the 26S proteasome, which is involved in ubiquitin-dependent proteolysis. The 20S proteasome can also bind 11S regulatory complexes (REG, PA28) which play a role in antigen processing, as do the three variable γ-interferon-inducible catalytic β-subunits (e.g. LMP7). In the present study, we have investigated the subcellular distribution of the different forms of proteasomes using subunit specific antibodies. Both 20S proteasomes and their 19S regulatory complexes are found in nuclear, cytosolic and microsomal preparations isolated from rat liver. LMP7 was enriched approximately two-fold compared with core α-type proteasome subunits in the microsomal preparations. 20S proteasomes were more abundant than 26S proteasomes, both in liver and cultured cell lines. Interestingly, some significant differences were observed in the distribution of different subunits of the 19S regulatory complexes. S12, and to a lesser extent p45, were found to be relatively enriched in nuclear fractions from rat liver, and immunofluorescent labelling of cultured cells with anti-p45 antibodies showed stronger labelling in the nucleus than in the cytoplasm. The REG was found to be localized predominantly in the cytoplasm. Three- to six-fold increases in the level of REG were observed following γ-interferon treatment of cultured cells but γ-interferon had no obvious effect on its subcellular distribution. These results demonstrate that different regulatory complexes and subpopulations of proteasomes have different distributions within mammalian cells and, therefore, that the distribution is more complex than has been reported for yeast proteasomes.

Publisher

Portland Press Ltd.

Subject

Cell Biology,Molecular Biology,Biochemistry

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