DYNAMICS OF PROTEASOME DISTRIBUTION IN LIVING CELLS

Proteasomes are proteolytic complexes involved in nonlysosomal degrada tion which are localized in both the cytoplasm and the nucleus, The dy namics of proteasomes in living cells is unclear, as is their targetin g to proteins destined for degradation, To investigate the intracellul ar distribution and mobility of proteasomes in vivo, we generated a fu sion protein of the proteasome subunit LMP2 and the green fluorescent protein (GFP). The LMP2-GFP chimera was quantitatively incorporated in to catalytically active proteasomes. The GFP-tagged proteasomes were l ocated within both the cytoplasm and the nucleus, Within these two com partments, proteasomes diffused rapidly, and bleaching experiments dem onstrated that proteasomes were transported slowly and unidirectionall y from the cytoplasm into the nucleus, During mitosis, when the nuclea r envelope has disintegrated, proteasomes diffused rapidly throughout the dividing cell without encountering a selective barrier, Immediatel y after cell division, the restored nuclear envelope formed a new barr ier for the diffusing proteasomes, Thus, proteasomes can be transporte d unidirectionally over the nuclear membrane, but can also enter the n ucleus upon reassembly during cell division. Since proteasomes diffuse rapidly in the cytoplasm and nucleus, they may perform quality contro l by continuous conclusion with intracellular proteins, and degrading those proteins that are properly tagged or misfolded.