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Dive into the research topics where Paul S. Danielian is active.

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Featured researches published by Paul S. Danielian.


Nature | 2010

Rb regulates fate choice and lineage commitment in vivo

Eliezer Calo; Jose A. Quintero-Estades; Paul S. Danielian; Simona Nedelcu; Seth D. Berman; Jacqueline A. Lees

Mutation of the retinoblastoma gene (RB1) tumour suppressor occurs in one-third of all human tumours and is particularly associated with retinoblastoma and osteosarcoma. Numerous functions have been ascribed to the product of the human RB1 gene, the retinoblastoma protein (pRb). The best known is pRb’s ability to promote cell-cycle exit through inhibition of the E2F transcription factors and the transcriptional repression of genes encoding cell-cycle regulators. In addition, pRb has been shown in vitro to regulate several transcription factors that are master differentiation inducers. Depending on the differentiation factor and cellular context, pRb can either suppress or promote their transcriptional activity. For example, pRb binds to Runx2 and potentiates its ability to promote osteogenic differentiation in vitro. In contrast, pRb acts with E2F to suppress peroxisome proliferator-activated receptor γ subunit (PPAR-γ), the master activator of adipogenesis. Because osteoblasts and adipocytes can both arise from mesenchymal stem cells, these observations suggest that pRb might play a role in the choice between these two fates. However, so far, there is no evidence for this in vivo. Here we use mouse models to address this hypothesis in mesenchymal tissue development and tumorigenesis. Our data show that Rb status plays a key role in establishing fate choice between bone and brown adipose tissue in vivo.


Proceedings of the National Academy of Sciences of the United States of America | 2008

Metastatic osteosarcoma induced by inactivation of Rb and p53 in the osteoblast lineage.

Seth D. Berman; Eliezer Calo; Allison S. Landman; Paul S. Danielian; Emily S. Miller; Julie C. West; Borel Djouedjong Fonhoue; Alicia M. Caron; Roderick T. Bronson; Mary L. Bouxsein; Siddhartha Mukherjee; Jacqueline A. Lees

Mutation of the RB-1 and p53 tumor suppressors is associated with the development of human osteosarcoma. With the goal of generating a mouse model of this disease, we used conditional and transgenic mouse strains to inactivate Rb and/or p53 specifically in osteoblast precursors. The resulting Rb;p53 double mutant (DKO) animals are viable but develop early onset osteosarcomas with complete penetrance. These tumors display many of the characteristics of human osteosarcomas, including being highly metastatic. We established cell lines from the DKO osteosarcomas to further investigate their properties. These immortalized cell lines are highly proliferative and they retain their tumorigenic potential, as judged by their ability to form metastatic tumors in immunocompromised mice. Moreover, they can be induced to differentiate and, depending on the inductive signal, will adopt either the osteogenic or adipogenic fate. Consistent with this multipotency, a significant portion of these tumor cells express Sca-1, a marker that is typically associated with stem cells/uncommitted progenitors. By assaying sorted cells in transplant assays, we demonstrate that the tumorigenicity of the osteosarcoma cell lines correlates with the presence of the Sca-1 marker. Finally, we show that loss of Rb and p53 in Sca-1-positive mesenchymal stem/progenitor cells is sufficient to yield transformed cells that can initiate osteosarcoma formation in vivo.


Genes & Development | 2010

A vertebrate gene, ticrr, is an essential checkpoint and replication regulator

Christopher L. Sansam; Nelly M. Cruz; Paul S. Danielian; Adam Amsterdam; Melissa L. Lau; Nancy Hopkins; Jacqueline A. Lees

Eukaryotes have numerous checkpoint pathways to protect genome fidelity during normal cell division and in response to DNA damage. Through a screen for G2/M checkpoint regulators in zebrafish, we identified ticrr (for TopBP1-interacting, checkpoint, and replication regulator), a previously uncharacterized gene that is required to prevent mitotic entry after treatment with ionizing radiation. Ticrr deficiency is embryonic-lethal in the absence of exogenous DNA damage because it is essential for normal cell cycle progression. Specifically, the loss of ticrr impairs DNA replication and disrupts the S/M checkpoint, leading to premature mitotic entry and mitotic catastrophe. We show that the human TICRR ortholog associates with TopBP1, a known checkpoint protein and a core component of the DNA replication preinitiation complex (pre-IC), and that the TICRR-TopBP1 interaction is stable without chromatin and requires BRCT motifs essential for TopBP1s replication and checkpoint functions. Most importantly, we find that ticrr deficiency disrupts chromatin binding of pre-IC, but not prereplication complex, components. Taken together, our data show that TICRR acts in association with TopBP1 and plays an essential role in pre-IC formation. It remains to be determined whether Ticrr represents the vertebrate ortholog of the yeast pre-IC component Sld3, or a hitherto unknown metazoan replication and checkpoint regulator.


Proceedings of the National Academy of Sciences of the United States of America | 2003

The role of E2F4 in adipogenesis is independent of its cell cycle regulatory activity

Rebecca L. Landsberg; Julia E. Sero; Paul S. Danielian; Tina L. Yuan; Eunice Y. Lee; Jacqueline A. Lees

The E2F and pocket protein families are known to play an important role in the regulation of both cellular proliferation and terminal differentiation. In this study, we have used compound E2F and pocket protein mutant mouse embryonic fibroblasts to dissect the role of these proteins in adipogenesis. This analysis shows that loss of E2F4 allows cells to undergo spontaneous differentiation. The ability of E2F4 to prevent adipogenesis seems to be quite distinct from the known properties of E2F. First, it can be separated from any change in either E2F-responsive gene expression or cell cycle regulation. Second, it is a specific property of E2F4, and not other E2Fs, and it occurs independently of E2F4s ability to interact with pocket proteins. In addition, E2F4 loss does not override the differentiation defect resulting from pRB loss even though it completely suppresses the proliferation defect of Rb−/− mouse embryonic fibroblasts. This finding definitively separates the known, positive role of pRB in adipogenesis from its cell cycle function and shows that this pocket protein is required to act downstream of E2F4 in the differentiation process.


Oncogene | 2008

E2f3a and E2f3b make overlapping but different contributions to total E2f3 activity

Paul S. Danielian; Lb Friesenhahn; Am Faust; Julie C. West; Alicia M. Caron; Roderick T. Bronson; Jacqueline A. Lees

The E2f transcription factors are key downstream targets of the retinoblastoma protein tumor suppressor that control cell proliferation. E2F3 has garnered particular attention because it is amplified in various human tumors. E2f3 mutant mice typically die around birth and E2f3-deficient cells have a proliferation defect that correlates with impaired E2f target gene activation and also induction of p19Arf and p53. The E2f3 locus encodes two isoforms, E2f3a and E2f3b, which differ in their N-termini. However, it is unclear how E2f3a versus E2f3b contributes to E2f3s requirement in either proliferation or development. To address this, we use E2f3a- and E2f3b-specific knockouts. We show that inactivation of E2f3a results in a low penetrance proliferation defect in vitro whereas loss of E2f3b has no effect. This proliferation defect appears insufficient to disrupt normal development as E2f3a and E2f3b mutant mice are both fully viable and have no detectable defects. However, when combined with E2f1 mutation, inactivation of E2f3a, but not E2f3b, causes significant proliferation defects in vitro, neonatal lethality and also a striking cartilage defect. Thus, we conclude that E2f3a and E2f3b have largely overlapping functions in vivo and that E2f3a can fully substitute for E2f1 and E2f3 in most murine tissues.


Genes & Development | 2015

Proteomic analysis of pRb loss highlights a signature of decreased mitochondrial oxidative phosphorylation

Brandon N. Nicolay; Paul S. Danielian; Filippos Kottakis; John D. Lapek; Ioannis Sanidas; Wayne O. Miles; Mantre Dehnad; Katrin Tschöp; Jessica J. Gierut; Amity L. Manning; Robert Morris; Kevin M. Haigis; Nabeel Bardeesy; Jacqueline A. Lees; Wilhelm Haas; Nicholas J. Dyson

The retinoblastoma tumor suppressor (pRb) protein associates with chromatin and regulates gene expression. Numerous studies have identified Rb-dependent RNA signatures, but the proteomic effects of Rb loss are largely unexplored. We acutely ablated Rb in adult mice and conducted a quantitative analysis of RNA and proteomic changes in the colon and lungs, where Rb(KO) was sufficient or insufficient to induce ectopic proliferation, respectively. As expected, Rb(KO) caused similar increases in classic pRb/E2F-regulated transcripts in both tissues, but, unexpectedly, their protein products increased only in the colon, consistent with its increased proliferative index. Thus, these protein changes induced by Rb loss are coupled with proliferation but uncoupled from transcription. The proteomic changes in common between Rb(KO) tissues showed a striking decrease in proteins with mitochondrial functions. Accordingly, RB1 inactivation in human cells decreased both mitochondrial mass and oxidative phosphorylation (OXPHOS) function. RB(KO) cells showed decreased mitochondrial respiratory capacity and the accumulation of hypopolarized mitochondria. Additionally, RB/Rb loss altered mitochondrial pyruvate oxidation from (13)C-glucose through the TCA cycle in mouse tissues and cultured cells. Consequently, RB(KO) cells have an enhanced sensitivity to mitochondrial stress conditions. In summary, proteomic analyses provide a new perspective on Rb/RB1 mutation, highlighting the importance of pRb for mitochondrial function and suggesting vulnerabilities for treatment.


Proceedings of the National Academy of Sciences of the United States of America | 2009

Mutation of p107 exacerbates the consequences of Rb loss in embryonic tissues and causes cardiac and blood vessel defects

Seth D. Berman; Julie C. West; Paul S. Danielian; Alicia M. Caron; James R. Stone; Jacqueline A. Lees

The retinoblastoma tumor-suppressor protein, pRb, is a member of the pocket protein family that includes p107 and p130. These proteins have well-defined roles in regulating entry into and exit from the cell cycle and also have cell cycle–independent roles in facilitating differentiation. Here we investigate the overlap between pocket proteins function during embryonic development by using conditional mutant alleles to generate Rb;p107 double-mutant embryos (DKOs) that develop in the absence of placental defects. These DKOs die between e13.5 and e14.5, much earlier than either the conditional Rb or the germline p107 single mutants, which survive to birth or are largely viable, respectively. Analyses of the e13.5 DKOs shows that p107 mutation exacerbates the phenotypes resulting from pRb loss in the central nervous system and lens, but not in the peripheral nervous system. In addition, these embryos exhibit novel phenotypes, including increased proliferation of blood vessel endothelial cells, and heart defects, including double-outlet right ventricle (DORV). The DORV is caused, at least in part, by a defect in blood vessel endothelial cells and/or heart mesenchymal cells. These findings demonstrate novel, overlapping functions for pRb and p107 in numerous murine tissues.


Cell Cycle | 2016

E2f4 and E2f5 are essential for the development of the male reproductive system

Paul S. Danielian; Rex A. Hess; Jacqueline A. Lees

ABSTRACT The E2F transcription factors are primarily implicated in the regulation of entry and exit from the cell cycle. However, in vivo studies have established additional roles for E2Fs during organ development and homeostasis. With the goal of addressing the intestinal requirements of E2f4 and E2f5, we crossed mice carrying Vil-cre, E2f4 conditional and E2f5 germline alleles. E2f4 deletion had no detectable effect on intestinal development. However, E2f4f/f;E2f5+/−;Vil-cre males, but not E2f4f/f;Vil-cre littermates, were unexpectedly sterile. This defect was not due to defective spermatogenesis. Instead, the seminiferous tubules and rete testes showed significant dilation, and spermatozoa accumulated aberrantly in the rete testis and efferent ducts. Our data show that these problems result from defective efferent ducts, a tissue whose primary function is to concentrate sperm through fluid absorption. First, Vil-cre expression, and consequent E2F4 loss, was specific to the efferent ducts and not other reproductive tract tissues. Second, the E2f4f/f;E2f5+/−;Vil-cre efferent ducts had completely lost multiciliated cells and greatly reduced levels of critical absorptive cell proteins: aquaporin1, a water channel protein, and clusterin, an endocytic marker. Collectively, the observed testis phenotypes suggest a fluid flux defect. Remarkably, we observed rete testis dilation prior to the normal time of seminiferous fluid production, arguing that the efferent duct defects promote excessive secretory activity within the reproductive tract. Finally, we also detect key aspects of these testis defects in E2f5−/− mice. Thus, we conclude that E2f4 and E2f5 display overlapping roles in controlling the normal development of the male reproductive system.


Cancer and Metabolism | 2014

Metabolic analysis of the loss of Rb1 in vivo

Brandon N. Nicolay; Paul S. Danielian; Wilhelm Haas; Gregory Stephanopoulos; Jacqueline A. Lees; Nicholas J. Dyson

Inactivation of the p16/pRB axis is a rate-limiting step in tumorigenesis. Loss of these tumor suppressors is well known to disrupt control of cell proliferation but, additionally, these mutations cause changes in cell metabolism. To investigate the metabolic changes resulting from pRB inactivation we have exploited a Drosophila model. Metabolomics and in vivo flux analysis revealed that RBF-deficient tissues have increased nucleotide production and struggle to maintain sufficient pools of reduced glutathione; ultimately leading to increased sensitivity to oxidative stress. We have now extended these studies to mammalian cell culture and mice. Depletion of pRB in non-tumorigenic hTERT-BJ cells leads to an increased cell population in S-phase and slight increases in the cell population doubling time. Acute loss of Rb1-/-in the mouse colon, but not the lung, leads to a robust increase in Ki67 staining within 96hrs. We therefore asked to what extent metabolic changes associated with proliferation increases are conserved between pRB-deficient cells in culture and acute loss of Rb1-/-in vivo. We performed isotopic tracer analyses with both glucose and glutamine and found that while loss of pRB in cell culture promotes increased isotopic enrichment from U13C-glutamine in TCA cycle intermediates, little to no change was seen in isotopic enrichment from U13C-glucose. In stark contrast, acute loss of Rb1 in the mouse colon leads to robust isotopic enrichment from U13C-glucose in TCA cycle intermediates and the pyrimidine precursor aspartate; while little to no change in isotopic enrichment from U13C-glutamine was found in all TCA cycle intermediates but alpha-ketoglutarate and no significant differences was seen in aspartate. Collectively these results reveal that loss of the tumor suppressor pRB has metabolic consequences in vivo.


Developmental Biology | 2009

E2F4 cooperates with pRB in the development of extra-embryonic tissues.

Eunice Y. Lee; Tina L. Yuan; Paul S. Danielian; Julie C. West; Jacqueline A. Lees

The retinoblastoma gene, RB-1, was the first identified tumor suppressor. Rb(-/-) mice die in mid-gestation with defects in proliferation, differentiation and apoptosis. The activating E2F transcription factors, E2F1-3, contribute to these embryonic defects, indicating that they are key downstream targets of the retinoblastoma protein, pRB. E2F4 is the major pRB-associated E2F in vivo, yet its role in Rb(-/-) embryos is unknown. Here we establish that E2f4 deficiency reduced the lifespan of Rb(-/-) embryos by exacerbating the Rb mutant placental defect. We further show that this reflects the accumulation of trophectoderm-like cells in both Rb and Rb;E2f4 mutant placentas. Thus, Rb and E2f4 play cooperative roles in placental development. We used a conditional mouse model to allow Rb(-/-);E2f4(-/-) embryos to develop in the presence of Rb wild-type placentas. Under these conditions, Rb(-/-);E2f4(-/-) mutants survived to birth. These Rb(-/-);E2f4(-/-) embryos exhibited all of the defects characteristic of the Rb and E2f4 single mutants and had no novel defects. Taken together, our data show that pRB and E2F4 cooperate in placental development, but play largely non-overlapping roles in the development of many embryonic tissues.

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Jacqueline A. Lees

Massachusetts Institute of Technology

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Julie C. West

Massachusetts Institute of Technology

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Alicia M. Caron

Massachusetts Institute of Technology

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Eunice Y. Lee

Massachusetts Institute of Technology

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Seth D. Berman

Massachusetts Institute of Technology

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Tina L. Yuan

Massachusetts Institute of Technology

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