Biographical Sketch

Michele Pagano is currently an Associate Professor at the New York University School of Medicine where he directs the Molecular Oncology Program of the Department of Pathology, and the Growth Control Program of the NYU Cancer Institute. He received his doctorate in Medicine with a thesis in Molecular Endocrinology in 1989 from the Federico II University in Naples, Italy. After joining Giulio Draetta’s lab (European Molecular Biology Laboratory in Heidelberg, Germany) in August 1990, his research interest has focused on the molecular control of the cell cycle in eukaryotes. As a postdoctoral fellow, he demonstrated that mammalian cyclins are required for DNA replication (Pagano et al., EMBO J. 11:961, 1992), (Pagano et al., J. Cell Biol. 121:101, 1993). He subsequently found that E2F, a transcription factor essential for the G1 to S transition, is one of the targets of Cdks that are active in S-phase (Pagano et al., Science 255:1144, 1992), (Pagano et al., Oncogene, 7:1681, 1992).

In November 1992, he became one of the scientific founders and principal investigators of Mitotix (Cambridge, Massachusetts), a biotechnology company that concentrates on the study of diseases resulting from inappropriate cell proliferation. As a team leader, his interests focused on how cyclins are involved in the cellular decision to proliferate or remain quiescent. His group was particularly interested in distinguishing the biological roles of two cyclins, Cyclin D1 and Cyclin E in normal and cancerous human cells. They found that Cyclin D1 is an essential component of the G1 checkpoint (Pagano et al., Genes & Dev. 8:2183, 1994). They then showed that in the absence of the tumor suppressor pRb, Cyclin D1 protein is expressed at low levels, is dissociated from its catalytic partner Cdk4 and is dispensable for the progression through the G1 phase of the cell cycle (Tam et al., Cancer Res. 54:5816, 1994), (Tam et al., Oncogene 9:2663, 1994). These data clearly indicated that Cyclin D1 is intimately involved in the Rb pathway. Furthermore, in collaboration with Jim Roberts’ group at the Fred Hutchinson Cancer Center in Seattle, they showed that in contrast to Cyclin D1, Cyclin E regulates the G1/S transition independent of pRb (Ohtsubo et al., Mol. Cell. Biol. 15:2612, 1995).

While studying the regulation of p27, a Cdk-inhibitor important for the regulation of the G1 phase, Pagano’s group found that p27 cellular abundance is controlled by the ubiquitin-dependent proteolytic pathway (Pagano et al., Science, 269:682, 1995). These results were the first demonstration that the levels of a mammalian Cdk subunit are regulated not by synthesis but by degradation and suggested that the specific proteolysis of p27 represents a novel mechanism for regulating the activity of Cdks. To pursue the fundamental mechanisms underlying the deregulation of cell cycle control in cancer, Michele Pagano moved in September 1996 to the New York University Medical Center as an Assistant Professor in the Department of Pathology. Currently, Pagano’s lab is studying the processes controlling the ubiquitination of cell cycle regulators, such as p21, p27, Emi1 and Cdc25A. They found that the presence of lysines is dispensable for the ubiquitination of both p21 and p27 (Bloom et al., Cell, 15:71, 2003), suggesting that the free amino group of the N-terminal methionine is a site for ubiquitinylation in vivo. Cdk-mediated phosphorylation of p27 on threonine 187 as well as a p27/cyclin/Cdk trimeric complex formation is essential for p27 ubiquitination (Montagnoli et al., Genes & Dev., 13:1181, 1999). They also found that Skp2, an F-box protein essential for entry into S phase, specifically recognizes p27 in a phosphorylation-dependent manner. Furthermore, both in vivo and in vitro, Skp2 is a rate-limiting component of the machinery that ubiquitinates and degrades phosphorylated p27 (Carrano et al., Nature Cell Biol., 1:193, 1999), (Carrano et al., J. Cell Biol.,153:1381, 2001). In collaboration with Avram Hershko at the Technion-Israel Institute of Technology, they have demonstrated that Skp2 requires an accessory protein, the cell cycle regulator Cks1, for the recognition, binding and ubiquitinylation of phosphorylated p27 (Ganoth et al., Nature Cell Biol., 3:321, 2001). Finally, in collaboration with Nikola Pavletich's group at Memorial Sloan-Kettering Cancer Center, they have solved the structure of the Skp1-Skp2 complex and shown that Skp1 recruits Skp2 through a bipartite interface involving both the F-box and the substrate recognition domain (Schulman et al., Nature, 408:381, 2000). Skp2 and its cofactor Cks1 are also unstable proteins in G1, and their degradation prevents unscheduled degradation of p21 and p27, and premature entry into S-phase. Significantly, degradation of Skp2 and Cks1 during G1 is controlled by another cell cycle-regulated ubiquitin ligase, the anaphase-promoting complex/cyclosome (APC/C) (Bashir et al., Nature, 248:190, 2004).

In collaboration with pathologists at the Dana Farber Cancer Institute and at the NYU Medical Center, Pagano's laboratory has expanded the studies on p27 to the clinic. They have found that aggressive human carcinomas contain high p27-specific degradation activity (Loda et al., Nature Med., 3:231, 1997). Additionally, they have found that the absence of p27 is a powerful prognostic marker for poor survival in patients with lymphomas, esophageal, prostate, breast, and colorectal carcinomas. Although cellular transformation has often been shown to involve the inactivation of some Cdk inhibitors, no homologous deletions or mutations of the p27 gene have been found so far in human tumors. The results from Pagano’s group suggest that low p27 expression observed in human carcinomas can result from an enhanced ubiquitin-mediated degradation of p27 rather than mutations in the p27 gene. Significantly, Skp2 levels inversely correlate with p27 expression in human breast cancers (Signoretti et al., J. of Clinincal Invest., 110:63, 2002) and lymphomas, and Skp2 cooperates with activated N-Ras in an in vivo model of lymphomagenesis (Latres et al., PNAS, 98:2515, 2001).

Using human Skp1 as a bait in a two-hybrid screen and searching DNA databases, Pagano’s group has identified a family of 26 human Fbps, 25 of which are novel (Cenciarelli et al., Current Biol. 9: 1177, 1999). Some of these Fbps contain WD-40 domains or leucine-rich repeats (LRRs) (both involved in substrate interaction), while other Fbps contain potential protein-protein interaction domains not previously identified in Fbps. Representative examples of all three classes of Fbps have been characterized and demonstrated to form novel active ubiquitin-ligase complexes in vivo which potentially target different substrates for ubiquitin-mediated degradation. They also investigated further the function of one of these F-box proteins, ß-Trcp1, by inactivating its gene in mice (Guardavaccaro et al., Dev. Cell, 4: 799–812, 2003 and Kudo et al., Mol. Cell. Biol., 24: 8184–8194, 2004). ß-Trcp1-/- males show reduced fertility correlating with an accumulation of methaphase I spermatocytes. §-Trcp1-/- MEFs display a lengthened mitosis, centrosome overduplication, multipolar metaphase spindles and misaligned chromosomes. Furthermore, they demonstrated that Emi1, an inhibitor of APC/C, is a substrate of ß-Trcp1. In contrast, stabilization of interphase substrates, such as ß-catenin (Latres et al., Oncogene, 18:849, 1999), IkB and Cdc25A (Busino et al. Nature, 426:87, 2003), does not occur in the absence of ß-Trcp1 and instead requires the additional silencing of ß-Trcp2 by siRNA. Thus ß-Trcp1 regulates the timely order of meiotic and mitotic events.