Subcellular localization of proteasomes and their regulatory complexes in mammalian cells

Proteasomes can exist in several different molecular forms in mammalian cel ls. The con 20S proteasome, containing the proteolytic sites, binds regulat ory complexes at the ends of its cylindrical structure. Together with two 1 9S ATPase regulatory complexes it forms the 26S proteasome, which is involv ed in ubiquitin-dependent proteolysis. The 20S proteasome can also bind IIS regulatory complexes (REG, PA28) which play a role in antigen processing, as do the three variable gamma-interferon-inducible catalytic beta-subunits (e.g. LMP7). In the present study, we have investigated the subcellular di stribution of the different forms of proteasomes using subunit specific ant ibodies. 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 alpha-type prot easome subunits in the microsomal preparations. 20S proteasomes were more a bundant than 26S proteasomes, both in liver and cultured cell lines. Intere stingly, some significant differences were observed in the distribution of different subunits of the 19S regulatory complexes. S12, and to a lesser ex tent p45, were found to be relatively enriched in nuclear fractions from ra t liver, and immunofluorescent labelling of cultured cells with anti-p45 an tibodies showed stranger labelling in the nucleus than in the cytoplasm. Th e REG was found to be localized predominantly in the cytoplasm. Three- to s ix-fold increases in the level of REG were observed following gamma-interfe ron treatment of cultured cells but gamma-interferon had no obvious effect on its subcellular distribution. These results demonstrate that different r egulatory complexes and subpopulations of proteasomes have different distri butions within mammalian cells and, therefore, that the distribution is mor e complex than has been reported for yeast proteasomes.