Cyclins, Cdks, E2f, Skp2, and More at the First International RB Tumor Suppressor Meeting

Published OnlineFirst July 7, 2010; DOI: 10.1158/0008-5472.CAN-10-0358 Published OnlineFirst on July 7, 2010 as 10.1158/0008-5472.CAN-10-0358

Meeting Report Cancer Research Cyclins, Cdks, E2f, Skp2, and More at the First International RB Tumor Suppressor Meeting

Rod Bremner1,3,4 and Eldad Zacksenhaus2,4,5,6

Abstract The RB1 gene was cloned because its inactivation causes the childhood ocular tumor, retinoblastoma. It is widely expressed, inactivated in most human malignancies, and present in diverse organisms from mammals to plants. Initially, retinoblastoma protein (pRB) was linked to cell cycle regulation, but it also regulates senescence, apoptosis, autophagy, differentiation, genome stability, immunity, telomere function, stem cell biology, and embryonic development. In the 23 years since the gene was cloned, a formal international symposium focused on the RB pathway has not been held. The “First International RB Tumor Suppressor Meeting” (Toronto, Canada, November 19-21, 2009) established a biennial event to bring experts in the field together to discuss how the RB family (“pocket proteins”), as well as its regulators and effectors, influence biology and human disease. We summarize major new breakthroughs and emerging trends presented at the meeting. Cancer Res; 70(15); OF1–5. ©2010 AACR.

Introduction critical for growth, thus their prominent expression could sensitize human cone precursors to RB1 mutation and un- Twenty speakers gave presentations on an astonishingly derlie a cone precursor origin of retinoblastoma (3). Studies diverse array of topics, underscoring the central role of reti- to date do not rule out an alternative possibility, that tumors noblastoma protein (pRB) in human biology (1). We have might convert to a cone-like epigenetic state during neoplas- grouped these topics into the general areas of cancer, E2f tic transformation (2). Irrespective, the data raise exciting functions, other pRB effectors, and pRB regulators. Refer- new avenues for treatment. The role of nuclear receptors ences are limited to work published in the past year. or Mdm2 in mouse retinoblastoma is unknown, but these tumors arise from amacrine cells (or horizontal cells that convert to amacrine cells during neoplastic transformation), RB1 in Cancer which exhibit naturally high resistance to apoptosis. Mouse data exist on Mycn: David Macpherson (Carnegie Institution, Retinoblastoma tumors carry homozygous RB1 mutations Baltimore, Maryland) reported gain in ∼30 to 50% of tumors (see below for an intriguing exception), and can be modeled from Rb/p107 or Rb/p130 null retina, respectively, and focal, in mice by deleting Rb1 and one of its relatives p107 or p130 high-level amplifications in a smaller subset, suggesting nat- (2). pRB mediates cell cycle exit in differentiating retinal neu- ural Mycn levels in mouse retina are insufficient for transfor- rons, and in its absence E2f1 delays cell cycle exit. Which mation. Brenda Gallie (Ontario Cancer Institute, Toronto, neuron is tumor-prone, and why, is debated. David Cobrinik Canada) has found that 1.5% of retinoblastoma tumors lack (Memorial Sloan-Kettering Cancer Center, New York, New RB1 mutations, but show high MYCN amplification. Thus, York) showed that human retinoblastoma exploits cone pho- RB1 loss plus naturally high MYCN, or high MYCN alone toreceptor features, including high expression (not amplifica- (plus as yet unknown other changes), are oncogenic in the tion) of the proto-oncogenes MYCN and MDM2,andthe human retina, and Rb1 plus Rb1-relative loss and Mycn gain nuclear receptors TRB2 and RXRG (3). All four factors are are oncogenic in mouse retina. Retinoblastoma exhibits common post-RB1 genetic events. Authors' Affiliations: 1Toronto Western Research Institute, 2Toronto General Research Institute, 3University Health Network, Departments of Gallie found that retinoma, a senescent benign tumor replete Ophthalmology and Vision Science, 4Laboratory Medicine and with p16Ink4a and p130 protein and evident in ∼15% of retino- Pathobiology, 5Medicine, 6Medical Biophysics, University of Toronto, RB1 Toronto, Ontario, Canada blastoma samples, has a subset of post- events present in malignant tumors, arguing that the latter derives from the for- Corresponding Authors: Rod Bremner, Toronto Western Research Insti- tute, University Health Network, 399 Bathurst Street, Room Mc6-424, mer. A common region of gain on human 1q contains KIF14 Toronto, Ontario, Canada, M5T 2S8. Phone: 416-603-5865; Fax: 416- and MDM4, and Macpherson showed that the syntenic region, 603-5126; E-mail: [email protected] and Eldad Zacksenhaus, Toronto General Research Institute, University Health Network, 67 College also on mouse chromosome 1q, undergoes copy number in- Street, Room 407, Toronto, Ontario, Canada, M5G 2M1. Phone: 416-340- crease in retinoblastoma models. Removing one allele of the 4800-5106; Fax: 416-340-3453; E-mail: [email protected]. Mdm2 inhibitor p19ARF accelerated mouse retinoblastoma, doi: 10.1158/0008-5472.CAN-10-0358 and approximately half of these malignancies showed loss of ©2010 American Association for Cancer Research. heterozygosity. These and other studies suggest an important

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role for the MDM2-p53 pathway in retinoblastoma progres- disorder in the hematopoietic system, but not leukemia. sion, even though p53 itself is never mutated. In summary, TKO mouse embryonic stem cells could differentiate into MYCN, the p14/19ARF-MDM2/MDM4-p53 axis, and KIF14 embryoid bodies, and only 5 to 10% of cells showed BrdU in- are key players after RB1 loss in both mouse and human corporation. In these differentiated TKO cells, some E2f tar- retinoblastoma. gets remained elevated, but others and Myc genes were Typically, RB1± individuals develop multiple tumors in down-regulated as effectively as in wild-type cells. TKO em- both eyes, but low penetrance mutations that partially dis- bryos survived until midgestation in the presence of a wild- able pRB cause fewer tumors. R661W is the best studied mu- type placenta. Thus, multiple TKO cell types have potent tation, and David Goodrich (Roswell Park Cancer Institute, backup mechanisms, likely explaining why oncogenic viral Buffalo, New York) is employing the knock-in equivalent he proteins bind many factors other than pocket proteins. developed (R654W) to model cancer in mice. For example, Laura Buttitta (Fred Hutchinson Cancer Research Center, R654W/− − − the Rb :p53 / prostate is better protected against tu- Seattle, Washington) also discussed extensive feedback con- −/− − − mor progression than the Rb :p53 / tissue. Some E2f tar- trols that prevent cell cycle abnormalities. Robust ectopic di- gets are equally deregulated in both genotypes, thus vision in differentiating Drosophila wing or eye cells requires protection must be through distinct E2f targets and/or E2f- perturbation of both Rb-E2f and Cdk2 pathways. Removing independent mechanisms. Goodrich has also developed an inhibitors or expressing activators from either side of the T epitope-tagged knock-in Rb1 allele that will be useful in equation has minimal effect, which is explained by feedback mapping the protein and genome-wide chromatin targets mechanisms, for example, whereas E2f induces cyclin A, it of pRB in vivo (4). also induces APC/C, which degrades cyclin A. Loss of heterozygosity and RB1 down-regulation are common in basal-like breast cancer, a highly aggressive subtype, as well as in luminal tumors. Eldad Zacksenhaus (Toronto Surprising In vivo Functions of Activating E2fs in General Research Institute, Toronto, Canada) described a Dividing Cells new mouse model for basal-like breast cancer, based on conditional Rb1 inactivation in mammary epithelium.7 Interest- Rod Bremner (Toronto Western Research Institute, Toronto, ingly, loss of Rb1 alone (without loss of Rb1 relatives) is Canada) and Gustavo Leone (Ohio State University, Columbus, sufficient to induce mammary tumors, and p53 status dic- Ohio) discussed in vivo functions of the three activating E2fs tates tumor subtype. (E2f1-3), downstream targets of pRB. Typically, E2fs are RB1 is also inactivated in hepatocellular carcinoma (HCC). viewed as indispensable for cell division. Surprisingly, howev- Erik Knudsen (Kimmel Cancer Center, Philadelphia, Pennsyl- er, many cell types divide in vivo without E2f1-3 (7, 8). In retina, vania) discussed the unexpected finding that conditional Rb1 this finding was explained by the ability of Mycn to maintain deletion does not perturb quiescence in normal or regenerat- expression of E2f targets and repress Cdkn1a (p21Cip1; ref. 7). ing mouse hepatocytes (even adding p107 or p130 absence; Unexpectedly, E2f1-3 have a prosurvival role in dividing ref. 5). Although Rb1 loss also does not cause HCC, it en- cells, which seems counterintuitive given that high E2f1-3 le- hances chemically induced tumorigenesis (see ref. 5 and re- vels drive apoptosis through p53 and p53-independent ferences therein). The expression profile of pRB target genes means. In retina, survival is linked to E2f-dependent expres- predicts disease outcome in human liver cancer, and animal sion of Sirt1 and other deacetylases that constrain p53 (7). models are revealing the impact of cell cycle inhibitory com- Death of E2f1-3 null crypt progenitors is p53-independent pounds on HCC.8 pRB blocks aflotoxin-induced HCC through (8). In addition to promoting survival, E2f1-3 also prevent a unique role in limiting E2f1 levels and ectopic DNA synthe- DNA damage in dividing cells (8), consistent with their sis. Aflotoxin has no effect on apoptosis, whether Rb1 is in- induction of genes that support nucleotide synthesis, replica- tact or not. In some other cell types, for example, gut villi, tion, and repair. Thus, E2f activity must be balanced careful- pRB potentiates activation of proapoptotic E2f targets and ly. Conceivably, low E2f activity could be oncogenic in cells in cell death in response to genotoxic agents (6). which Myc is sufficient for division. Although Rb1/p107 double knockout (DKO) liver does not “Activating” E2fsarealsolinkedtogenerepression,but develop spontaneous tumors (5), Julien Sage (Stanford Uni- in vivo relevance is unclear. Leone revealed that E2f1-3 mediate versity School of Medicine, Stanford, California) showed that repression in postmitotic differentiating cells (8). In dividing Rb1/p107/p130 triple knockout (TKO) causes HCC within 4 to small intestine crypt cells, pRB is phosphorylated and not 6 months. Molecular analysis connected pocket protein loss E2f-bound. Conversely, in differentiating postmitotic villi, pRB with Notch signaling, extending observations made in neural is dephosphorylated and forms pRB-E2f repressor complexes. progenitors. Pocket protein loss causes a myeloproliferative In E2f1-3 TKO, differentiating villi derepression increases E2f − − targets, and Rb1 / null villi show E2f1-3-dependent superacti- vation of these targets and ectopic division. 7 Z. Jiang et al. Rb deletion in mouse mammary progenitors induces luminal-B To summarize, although abnormal proliferation in Rb1- or basal like/EMT tumor subtypes depending on p53 status. J. Clinical Inves- null-differentiating cells is E2f-dependent, in normal prolifer- tigation, in press. ating cells, activating E2fs promote survival and genome 8 D.B. Rivandeira et al. Proliferative suppression by CDK4/6 inhibition: complex function of the retinoblastoma pathway in liver tissue and hepatoma cells. integrity rather than division. In postmitotic differentiating Gastroenterology, in press. cells, E2f1-3 form essential repressor complexes with pRB.

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pRB Effectors: E2f and Beyond Hein te Riele (Netherlands Cancer Institute, Amsterdam, The Netherlands) also described DNA damage in Rb1/p107/ pRB binds >100 proteins through various surfaces. Sibylle p130 TKO MEFs. These cells lack G1 control and are refrac- Mittnacht (Chester Beatty Laboratories, London, UK) showed tory to Ras(V12)-induced senescence. Serum-starved TKO that the N-terminal domain is structurally related to the A-B MEFs ignore the serum-dependent G1-S restriction point pocket domain, and they interact. Peptides that bind to the and enter S-phase, but die at G2/M. Bcl2 expression blocks LxCxE cleft of the A-B pocket disrupt N-terminal-A-B pocket apoptosis and exposes G2/M arrest mediated by p21/p27- interaction. Mittnacht also deduced the effect of tumorigenic inhibition of cyclin A/B-Cdk1. Readding serum down-regulates mutations on pRB function and distinguished these defects p21/p27, and mitotic entry occurs after ∼24 hours. Mitogen from functionally silent codon variants. must be added for at least 6 hours, exposing a second serum- Srikanth Talluri, from Fred Dick's group (University of dependent “restriction point” (“R2”). Although reentry is pos- Western Ontario, London, Canada), discussed their work with sible, chromosomal breaks and “railroad chromosomes” lack ΔL/ΔL Rb knock-in mice, which lack the LxCxE-binding groove. centromeric constriction. This damage might be related to ΔL/ΔL Rb murine embryonic fibroblasts (MEF) have abnormal E2f-dependent and/or -independent chromatin regulation pericentric heterochromatin and histone H4K20 trimethylation, mentioned above. Kay Macleod (University of Chicago, Chica- yet cell cycle exit in differentiating neurons and muscle fibers is go, Illinois) has shown that erythroblasts lacking E2f2, the normal, and the mice are viable. However, senescence-induced major activating E2f in these cells, also have chromosome exit in MEFs and transforming growth factor β (TGFβ-induced condensation defects, potentially related to deregulation of 9 exit in mammary epithelium are defective, correlating with the E2f targets Aurora B, caspase-3, and Rad21. failure to down-regulate E2f targets (9, 10). Thus, LxCxE pro- Prior studies proposed that pRB facilitates differentiation teins are required for context-specific repression by E2f-pRB. by enhancing the function of lineage-specific transcription pRB also has E2f-independent functions. When phos- factors. However, more recent work attributes several differ- phorylated on Thr187, the CDK inhibitor p27Kip1 is targeted entiation defects to deregulated E2f activity; for example, de- −/− for degradation by Skp2. Liang Zhu (Albert Einstein College leting E2f1 blocks ectopic division in Rb retina, rescuing of Medicine, New York, New York) showed that pRB binds all but one differentiation defect, and even this cell-cycle- Skp2, blocking binding to Thr187-phosphorylated p27. Nick independent abnormality is E2f3a-dependent. Ruth Slack Dyson (Massachusetts General Hospital, Boston, Massachusetts) (University of Ottawa, Ottawa, Canada) and Macleod dis- −/− found pRB and Skp2 in a larger complex with APCCdh1,driving cussed E2f-dependent defects in Rb1 cortical neurons SKP2 ubiquitylation and degradation. pRB also represses Skp2 and erythroblasts. E2f7, a repressive family member, does − − transcription through E2f. Thus, in Rb1 / cells, Skp2 levels in- not bind pRB and is induced by activating E2fs. Slack showed −/− crease and p27Kip1 is degraded. Indeed, Skp2 is required for pitu- E2f7 induction in the Rb1 cortex and repression of Dlx, a itary tumorigenesis in Rb1± mice (11). Skp2 inactivation and homeobox gene that regulates migration. Macleod showed −/− p27Kip1 stabilization causes apoptosis rather than arrest of Rb1- that Rb1 erythrocyte defects are E2f2-dependent. In normal null melanotrophs. Apoptosis is p27Kip1-dependent because a erythrocytes, pRB/E2f2 complexes repress E2f1 and other E2f- −/− p27T187A allele, which cannot bind Skp2, mimics Skp2 loss. This target genes, ensuring G1 arrest and enucleation. In Rb1 is the first case showing that p27T187A phenocopies Skp2 absence. erythrocytes, deregulated E2f2 drives S-phase, increasing Zhu and Cobrinik showed that down-regulating SKP2 or expres- reactive oxygen species, single-strand damage, and DNA- sing p27T187A kills human retinoblastoma cells. Thus, SKP2 and damage-induced arrest in G2 (also discussed above), explain- Thr187 ing why these cells fail to enucleate. The Rb-E2f pathway also p27 phosphorylation represent excellent therapeutic targets − − Kip1 Rb1 / in pRB-deficient tumors. How p27 induces death in the regulates muscle development (12). Indeed, muscle absence of Skp2 is unclear. cells exhibit endoreduplication, incomplete differentiation, Dyson described an E2f-independent function of pRB fam- and degeneration. Apoptosis in unfused myoblasts is mediat- ily proteins in promoting chromatin condensation and segre- ed by noncanonical cell death. Zacksenhaus showed that Rb1 gation, through interaction with dCAP-D3/Condensin II. This induction of several survival pathways allows -deficient interaction, together with a recently discovered defect in sis- myocytes to fuse and form functional myotubes despite con- 10 Rb1 ter chromatin cohesion, may explain the increase in chromo- tinued DNA synthesis. Thus, promotes survival of dif- some missegregation and aneuploidy in pRB-deficient ferentiated muscle and may not be critical to stimulate tumors. Binding requires the pRB LxCxE binding groove. Fred activity of differentiation factors. ΔL/ΔL Dick showed that RB MEFs exhibit reduced H4K20 trimethylation in pericentric heterochromatin, which may also How Is pRB Regulated? lead to centromere fusions, chromosome missegregation, and genomic instability. The latter might be E2f-dependent, as Several posttranslational modifications regulate pRB. pRB recruits the H4K20me-binding protein L3MBTL1 to E2f Cyclin/Cdk complexes negatively regulate RB and are often targets. Decrease in H4K20 trimethylation and increased telomere length (shown previously by the Blasco laboratory), seen in double and triple Rb-p107-p130 TKO MEFs, together 9 Y. Zhou, A.M. Dirlam, K.F. Macleod, unpublished observations. with the chromosome segregation defects described above, 10 G. Ciavarra and E. Zacksenhaus. Rescue of Rb-deficient myogenic defect by may underline genomic instability in human cancer. survival factors, submitted for publication.

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overexpressed and/or activated in cancer. In some contexts, defects. Using new Flag-HA-cyclin-D1 knock-in mice, proteo- active pRB is used in cancer cells, likely to block E2f1-induced mics, and chromatin immunoprecipitation coupled to DNA apoptosis. David Park (University of Ottawa, Ottawa, Canada) microarray (ChIP-chip) Sicinski's laboratory showed that discussed the role of pRB phosphorylation in postmitotic cyclin D1 binds to promoter regions of several genes (15). In neurons after stroke. Cdk4 activity is upregulated following the retina, this activity is required for D1-mediated transcrip- − − stroke and is critical for death following ischemia. Cdk4 acts tion of Notch1. In D1 / retina, Notch1 is down-regulated, thus upstream of conserved death pathways in vitro and in vivo. causing premature cell cycle exit. Remarkably, transduction of Pim1 and Chk1 kinases act reciprocally to regulate the phos- activated Notch partially rescues the division defects. Thus, − − phatase Cdc25, which in turn regulates Cdk4 activation after hypocellularity of the D1 / retinaisduetobothreduced neuronal injury. division rate and early cell cycle exit linked to lower Cdk4/6 Seth Rubin (University of California, Santa Cruz, Califor- and Notch1 activity, respectively. nia) discussed structural aspects of how phosphorylation affects pRB-E2f interaction. pRB forms two contacts with E2f: Future Perspectives the pRB C-terminus binds E2f/Dp1 heterodimer, and the pRB pocket binds the E2f trans-activation domain (TD). Phos- Despite the more than two decades that have passed since phorylation of Ser/Thr sites in the pRB spacer or N-terminus its molecular secrets began to unfold, the RB pathway con- inhibits Rb-pocket/E2f-TD interaction. Moreover, the phos- tinues to surprise. Retinoblastoma is still yielding fundamen- phatase PP1 binds C-terminal residues 870 to 882, and not tal insights into cancer, emphasizing the value of studying only dephosphorylates pRB, but prevents cyclin-Cdk binding. this rare but, relatively speaking, simpler class of neoplasia. Thus, catalytically dead PP1 could block Cdk2-mediated New animal models underscore the importance of the RB phosphorylation of pRB. family in other cancers, but also raise questions about how pRB is phosphorylated sequentially by cyclin D1/2/3-Cdk4/ cells cope with Rb1 loss, why some cell types are more sus- 6 and cyclin E1/2-Cdk2 during G1/S transition, and then cy- ceptible than others, and when and where RB family action is clin A1/2-Cdk2/1. Peter Sicinski (Dana-Farber Cancer Insti- most critical: Is it primarily in the cell cycle and cell survival? tute, Boston, Massachusetts) described redundancy in this Or are its effects on genome stability and other aspects of system. Cyclin D1/D2/D3 TKO embryos survive to midgesta- cellular metabolism just as critical? And if they are, how is tion and exhibit normal proliferation in most compartments. the pathway controlling these key determinants? Many ques- TKO MEFs proliferate relatively normally, but are hypersen- tions also remain about where and when RB effectors are sitive to Cdk2-inhibition. When TKO cells exit quiescence, critical for its action in vivo. Clearly, E2f and Skp2 are major pRB and p107 phosphorylation, presumably via Cdk2-cyclinE, RB targets, and key goals are: (a) to revisit the plethora of is slightly delayed. Likewise, MEFs lacking cyclin E1/E2 pro- phenotypes linked with Rb family loss, and define those that liferate, although they cannot reenter the cell cycle from qui- are E2f- and/or Skp2-dependent or -independent; and (b) to escence. Thus, dividing cells do not require sequential D-E-A define the relevance of other RB protein-protein interactions kinase pRB phosphorylation, but adapt and use different (for example, LxCxE partners). At the atomic level, we still cyclin/Cdk combinations. Indeed, combined ablation of all need a fuller understanding of the details of how RB is mod- A- and E-type cyclins in fibroblasts causes arrest, which is ified, and how, exactly, this affects its partners. Surprising not cell cycle phase-specific (13). D+E or D+A knockout stud- new functions for activating E2fs in promoting survival and ies will also be interesting. Parallel studies by others show DNA integrity, and for cyclin D1 in supporting Notch action, that CDK1 is the only essential CDK, whereas CDK2-6 are suggest that similar genetic and biochemical studies with dispensable for cell cycle progression. these and other members of the RB pathway will yield more In addition to posttranslational modifications, Rb1 is also exciting discoveries. It will also be important to define regulated transcriptionally. This finding is most evident dur- whether the links between low E2f and DNA damage, and ing embryogenesis, in which Rb1 is abundant in the nervous − − between cyclin D1 and Notch signaling, are important in hu- system, muscle, liver, and the tissues affected in Rb1 / or − − man cancer and other disorders in which the RB pathway is placental-rescued Rb1 / mutant embryos. Zacksenhaus commonly perturbed. Where the field will take us in the next showed previously that 6 kb of upstream sequence is suffi- few decades is impossible to predict, but we are certain it will cient for expression in the nervous system, but not skeletal continue to have a broad impact on our understanding of muscle. Using a green fluorescent protein (GFP) reporter biology and human health. knocked-in into a 200-kb bacterial artificial chromosome en- compassing most of Rb1, Sage observes expression in several tissues including nervous tissue and skeletal muscle, but not Disclosure of Potential Conflicts of Interest liver, thus Rb1 regulation involves multiple remote enhancers No potential conflicts of interest were disclosed. (14). Vectors that permit appropriate expression will facili- tate structure-function analysis of pRB in vivo. Sicinski also revealed Cdk-independent cyclin D1 functions. Acknowledgments Previously, Hinds showed that knock-in mice expressing K112E −/− We thank the speakers for sharing unpublished data and critiquing the D1 , which does not bind Cdk4/6, retain D1 phenotypes manuscript; Lucy Fuccillo and Kristina Matevski for coordinating the −/− related to tumorigenicity, but lack D1 developmental meeting; Greg Clarke for webpage design; and Bob Philips for advice.

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Health Network (UHN), the Department of Ophthalmology and Vision Grant Support Sciences University of Toronto, Cedarlane, ThermoFisher Scientific, Sarstedt, and Diamed provided sponsorship. The Canadian Institute for Health Research (CIHR) and the Terry Fox Foundation provided peer-reviewed funding. The Ontario Institute for Cancer Received 02/02/2010; revised 05/13/2010; accepted 05/15/2010; published Research (OICR), the Toronto General Research Institute (TGRI), University OnlineFirst 07/06/2010.

References 1. First International Rb meeting. Toronto (Canada): University Health 9. Talluri S, Isaac CE, Ahmad M, et al. A G1 checkpoint mediated by the Network; 2009. Available from: http://www.rbmeeting.ca/. retinoblastoma protein that is dispensable in terminal differentiation 2. Bremner R. Retinoblastoma, an inside job. Cell 2009;137:992–4. but essential for senescence. Mol Cell Biol 2010;30:948–60. 3. Xu XL, Fang Y, Lee TC, et al. Retinoblastoma has properties of a 10. Francis SM, Bergsied J, Isaac CE, et al. A functional connection be- cone precursor tumor and depends upon cone-specific MDM2 sig- tween pRB and transforming growth factor beta in growth inhibition naling. Cell 2009;137:1018–31. and mammary gland development. Mol Cell Biol 2009;29:4455–66. 4. Xiao H, Zhang X, Goodrich DW. Construction of a dual affinity tagged 11. Wang H, Bauzon F, Ji P, et al. Skp2 is required for survival of aber- allele of the Rb1 tumor suppressor gene in the mouse. Genesis 2009; rantly proliferating Rb1-deficient cells and for tumorigenesis in 48:121–6. Rb1(±) mice. Nat Genet 2010;42:83–8. 5. Reed CA, Mayhew CN, McClendon AK, Knudsen ES. Unique impact 12. Asp P, Acosta-Alvear D, Tsikitis M, van Oevelen C, Dynlacht BD. of RB loss on hepatic proliferation: tumorigenic stresses uncover dis- E2f3b plays an essential role in myogenic differentiation through iso- tinct pathways of cell cycle control. J Biol Chem 2010;285:1089–96. form-specific gene regulation. Genes Dev 2009;23:37–53. 6. Ianari A, Natale T, Calo E, et al. Proapoptotic function of the retino- 13. Kalaszczynska I, Geng Y, Iino T, et al. Cyclin A is redundant in fibro- blastoma tumor suppressor protein. Cancer Cell 2009;15:184–94. blasts but essential in hematopoietic and embryonic stem cells. Cell 7. Chen D, Pacal M, Wenzel P, Knoepfler PS, Leone G, Bremner R. Di- 2009;138:352–65. vision and apoptosis of E2f-deficient retinal progenitors. Nature 14. Burkhart DL, Ngai LK, Roake CM, et al. Regulation of Rb transcrip- 2009;462:925–9. tion in vivo by Rb family members. Mol Cell Biol 2010;30:1729–45. 8. Chong JL, Wenzel PL, Saenz-Robles MT, et al. E2f1–3 switch from 15. Bienvenu F, Jirawatnotai S, Elias JE, et al. Transcriptional role of activators in progenitor cells to repressors in differentiating cells. cyclin D1 in development revealed by a genetic-proteomic screen. Nature 2009;462:930–4. Nature 2010;463:374–8.

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Cyclins, Cdks, E2f, Skp2, and More at the First International RB Tumor Suppressor Meeting

Rod Bremner and Eldad Zacksenhaus

Cancer Res Published OnlineFirst July 7, 2010.

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