The picture above represents a schematic representation of the UPS, which is explained in more detail below.

20S proteasomes are cylinder-shaped, multisubunit, self-compartmentalized, multi-catalytic proteases able to degrade efficiently short peptides, however quite reluctant in the degradation of entire proteins. 20S proteasomes bind different regulatory complexes such as PA700 (11S) or PA28 (REG). The complex of the 20S proteasome with PA700 known as the 26S proteasome is of uttermost significance: it is able to degrade native proteins, most of them targeted by the prior attachement of a polyubiquitin chain. PA700 allows the recognition of substrates, their binding, unfolding, threading to the 20S catalytic core and assures the release of intact ubiquitin tags. Those processes require metabolic energy in form of ATP hydrolysis, despite the fact that the hydrolysis of a peptide bond releases free energy.

Attachement of multiple ubiquitin (Ubi) moieties (=ubiquitination) to a protein, targets that protein for degradation by the 26S proteasomes. Ubiquitination is mediated by a cascade of enzymes: E1 or ubiquitin-activating enzyme, E2 or ubiquitin-conjugating enzyme and E3 or ubiquitin ligase. Human genome encodes hundreds of different E3s, which are often multimeric modular complexes and bring upon a high specificity in the recognition of susbstrates.. The action of the E1-E2-E3 cascade is counteracted by multiple deubiquitinating enzymes (DUBs). To make things even more complicated, ubiquitin can serve non-proteolytic functions as well. Moreover, several ubiquitin-like proteins exist, which may modify proteins in a way similar to ubiquitin.

Until recently it was believed that ubiquitinated proteins are recognized directly by 26S proteasomes, however new data emerged which suggest that in the cell numerous adaptor proteins may exist, which bind on one side the 26S proteasome and on the other side the ubiquitinated substrate acting like molecular shuttles. While it is difficult to understand the rationale for the existence of such shuttles in case of cytosolic substrates, in the case of ERAD their need is more obvious. Lumenal or transmembrane ER proteins which failed to fold properly and are doomed for destruction must be first recognized by a complicated ER chaperone system, then threaded back to the cytosol, recognized as UPS susbstrates, ubiquitinated and delivered to the 26S proteasomes. The pathways of mammalian ERAD are being elucidated based on the previous studies performed in yeast. Multiple lines of evidence point to a special role played in ERAD by a protein called VCP (valosin-containing protein), p97 or Cdc48p. Studying the role of VCP in ERAD and UPS in general is a special focus of our lab.

Different cytosolic or ERAD substrates in the cytosol are usually immedioately cleared by the proteasomes, however when their burden exceeds the degradative capacity of the UPS they tend to aggregate. Back in 1996 I have described that treatment of human cells with proteasome inhibitors leads to the formation of a single perinuclear aggregate rich in ubiquitin and proteasomal antigens. Moreover, I have demonstrated that formation of such an aggregate depends on microtubule transport and new protein synthesis. Electron microscopy has revealed that this aggregate, forms out of an electron-dense material around the centrosome and in the vicinity of the Golgi. Once the aggregate is formed it is removed from the cells by autophagy. To explain this finding I have proposed that the aggregates form in a region of the cytosol which is most active in terms of UPS-dependent proteolysis, most likely enriched in some hypothetical proteasome activators, called the proteolysis center. Two years later, the group of Ron Kopito has reproduced my results, enriching them with the finding that overexpression of a transmembrane protein such as CFTR also leads to the formation of such aggregate or “aggresome”. Since then it has become obvious that formation of such aggregates mimics the situation found in vivo during various neurodegenerative disorders, such as Alzheimer’s and Parkinson’s diseases.

UPS is involved in the degradation of cyclins, cyclin-dependent kinase inhibitors, transcription factors, pro-apoptotic and anti-apoptotic molecules etc.,  therefore pharmacological inhibition of the UPS induces cell cycle block and apoptosis in various cancer cell lines, while is relatively harmless to normal cells. Together with my colleagues from the Department of Immunology of the Medical University of Warsaw, Poland we were among the first to show the anticancer effects of proteasome inhibitors in vitro (1997) and in vivo, using murine cancer models (1999). Nowadays, a potent proteasome inhibitor called Velcade has been already approved by the FDA for the treatment of multiple myeloma and other malignancies, however many unanswered questions remain regarding its mechanism of action. Moreover, despite limited success Velcade is not a magic bullet against cancer and further work is needed in order to optimize therapies based on the combination of Velcade with other drugs, cytokines and anticancer agents. Quest for such treatments is also in the focus of our research group.

Selected Publications