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Weber U.,Mount Sinai School of Medicine | Gault W.J.,Mount Sinai School of Medicine | Gault W.J.,Memorial Sloan Kettering Institute | Olguin P.,Mount Sinai School of Medicine | And 2 more authors.
Genetics | Year: 2012

Planar cell polarity (PCP) is a common feature of many epithelia and epithelial organs. Although progress has been made in the dissection of molecular mechanisms regulating PCP, many questions remain. Here we describe a screen to identify novel PCP regulators in Drosophila. We employed mild gain-of-function (GOF) phenotypes of two cytoplasmic Frizzled (Fz)/PCP core components, Diego (Dgo) and Prickle (Pk), and screened these against the DrosDel genome-wide deficiency collection for dominant modifiers. Positive genomic regions were rescreened and narrowed down with smaller overlapping deficiencies from the Exelixis collection and RNAi-mediated knockdown applied to individual genes. This approach isolated new regulators of PCP, which were confirmed with loss-of-function analyses displaying PCP defects in the eye and/or wing. Furthermore, knockdown of a subset was also sensitive to dgo dosage or dominantly modified a dishevelled (dsh) GOF phenotype, supporting a role in Fz/PCP-mediated polarity establishment. Among the new "PCP" genes we identified several kinases, enzymes required for lipid modification, scaffolding proteins, and genes involved in substrate modification and/or degradation. Interestingly, one of them is a member of the Meckel-Gruber syndrome factors, associated with human ciliopathies, suggesting an important role for cell polarity in nonciliated cells. © 2012 by the Genetics Society of America.

Gustafsson K.,Uppsala University | Jamalpour M.,Uppsala University | Trinh C.,Uppsala University | Kharas M.G.,Memorial Sloan Kettering Institute | Welsh M.,Uppsala University
Journal of Hematology and Oncology | Year: 2014

Background: The Src homology-2 domain protein B (Shb) is an adapter protein operating downstream of several tyrosine kinase receptors and consequently Shb regulates various cellular responses. Absence of Shb was recently shown to reduce hematopoietic stem cell proliferation through activation of focal adhesion kinase (FAK) and thus we sought to investigate Shb's role in the progression of leukemia. Methods. Wild type and Shb knockout bone marrow cells were transformed with a retroviral BCR-ABL construct and subsequently transplanted to wild type or Shb knockout recipients. Disease latency, bone marrow and peripheral blood cell characteristics, cytokine expression, signaling characteristics and colony formation were determined by flow cytometry, qPCR, western blotting and methylcellulose colony forming assays. Results: It was observed that Shb knockout BCR-ABL-transformed bone marrow cells produced a disease with death occurring at earlier time points compared with corresponding wild type controls due to elevated proliferation of transformed bone marrow cells. Moreover, significantly elevated interleukin-6 and granulocyte colony-stimulation factor mRNA levels were observed in Shb knockout c-Kit + leukemic bone marrow cells providing a plausible explanation for the concurrent peripheral blood neutrophilia. Shb knockout leukemic bone marrow cells also showed increased ability to form colonies in methylcellulose devoid of cytokines that was dependent on the concomitantly observed increased activity of FAK. Transplanting BCR-ABL-transformed Shb knockout bone marrow cells to Shb knockout recipients revealed decreased disease latency without neutrophilia, thus implicating the importance of niche-derived cues for the increase of blood granulocytes. Conclusions: Absence of Shb accelerates disease progression by exerting dual roles in BCR-ABL-induced leukemia: increased cell expansion due to elevated FAK activity and neutrophilia in peripheral blood, the latter dependent on the genetic background of the leukemic niche. © 2014 Gustafsson et al.; licensee BioMed Central Ltd.

Jeong J.,Memorial Sloan Kettering Institute | White N.E.,University of Missouri | Loyalka S.K.,University of Missouri
Annals of Nuclear Energy | Year: 2015

"Three-dimensional transport theory: an analytical solution of an internal beam searchlight problem, I", Annals of Nuclear Energy, 36(8), 1256-1261 (2009) by Williams extends the range of analytical solutions, and the associated development of techniques, numerical results and analysis near singularities. The final integrals are not easy to evaluate as the integrands are highly oscillatory, singular and also on infinite range. We report here some further numerical evaluations of expressions of Williams, and also compare these with those of Williams and Ganapol and Kornreich. The numerical results compare very well. The disagreements are very rare, and even then in the fifth decimal place. We are also able to explore the nature of the results near singularities in conformity with the results of Williams. We also verify MCNP-5, the widely used Monte Carlo code against these analytical results. We have found that MCNP is easily able to provide results within 0.1% deviation from the "exact" results for most cases, and within 1% for almost all cases. It is challenged near the singularities, however, where the deviations are larger. © 2015 Elsevier Ltd. All rights reserved.

Pan G.,Howard Hughes Medical Institute | Feng Y.,Howard Hughes Medical Institute | Feng Y.,Memorial Sloan Kettering Institute | Ambegaonkar A.A.,Howard Hughes Medical Institute | And 4 more authors.
Development (Cambridge) | Year: 2013

The large atypical cadherin Fat is a receptor for both Hippo and planar cell polarity (PCP) pathways. Here we investigate the molecular basis for signal transduction downstream of Fat by creating targeted alterations within a genomic construct that contains the entire fat locus, and by monitoring and manipulating the membrane localization of the Fat pathway component Dachs. We establish that the human Fat homolog FAT4 lacks the ability to transduce Hippo signaling in Drosophila, but can transduce Drosophila PCP signaling. Targeted deletion of conserved motifs identifies a four amino acid C-terminal motif that is essential for aspects of Fat-mediated PCP, and other internal motifs that contribute to Fat-Hippo signaling. Fat-Hippo signaling requires the Drosophila Casein kinase 1ε encoded by discs overgrown (Dco), and we characterize candidate Dco phosphorylation sites in the Fat intracellular domain (ICD), the mutation of which impairs Fat-Hippo signaling. Through characterization of Dachs localization and directed membrane targeting of Dachs, we show that localization of Dachs influences both the Hippo and PCP pathways. Our results identify a conservation of Fat-PCP signaling mechanisms, establish distinct functions for different regions of the Fat ICD, support the correlation of Fat ICD phosphorylation with Fat-Hippo signaling, and confirm the importance of Dachs membrane localization to downstream signaling pathways. © 2013.

Hanoun M.,Yeshiva University | Zhang D.,Yeshiva University | Mizoguchi T.,Yeshiva University | Pinho S.,Yeshiva University | And 6 more authors.
Cell Stem Cell | Year: 2014

Perivascular mesenchymal stem and progenitor cells (MSPCs) are critical for forming a healthy hematopoietic stem cell (HSC) niche. However, the interactions and influence of acute myelogenous leukemia (AML) stem cells with the microenvironment remain largely unexplored. We have unexpectedly found that neuropathy of the sympathetic nervous system (SNS) promotes leukemic bone marrow infiltration in an MLL-AF9 AML model. Development of AML disrupts SNS nerves and the quiescence of Nestin+ niche cells, leading to an expansion of phenotypic MSPCs primed for osteoblastic differentiation at the expense of HSC-maintaining NG2+ periarteriolar niche cells. Adrenergic signaling promoting leukemogenesis is transduced by the β2, but not β3, adrenergic receptor expressed on stromal cells of leukemic bone marrow. These results indicate that sympathetic neuropathy may represent a mechanism for the malignancy in order to co-opt the microenvironment and suggest separate mesenchymal niche activities for malignant and healthy hematopoietic stem cells in the bone marrow. © 2014 Elsevier Inc.

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