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Lalonde M.E.,Laval University | Cote J.,Laval University | Yang X.J.,Rosalind and Morris Goodman Cancer Research Center
Epigenetics : official journal of the DNA Methylation Society | Year: 2014

The MOZ/MORF complexes represent an example of a chromatin-binding assembly whose recruitment to specific genomic regions and activity can be fine-tuned by posttranslational modifications of histones. Here we detail the structures and biological functions of epigenetic readers present in the four core subunits of the MOZ/MORF complexes, highlight the imperative role of combinatorial readout by the multiple readers, and discuss new research directions to advance our understanding of histone acetylation.


Li W.,Wuxi Shunye Technology Co. | Yang J.,Wuxi Shunye Technology Co. | Yang J.,McGill University | Yang J.,Rosalind and Morris Goodman Cancer Research Center | And 3 more authors.
Materials Research Express | Year: 2015

We report a simple route for the synthesis of several morphologies of self-assembling hierarchical Ni (OH)2 nanostructures, by the reaction of NiSO4 and NH4OH in aqueous solution, at a constant temperature, using neither surfactant nor template. Both morphology and microstructure depend on the concentrations of the reactants, the reaction temperature and the anions (Cl-, .., NO3 - and SO4 2-) present. The nanostructures have been characterized by scanning electron microscopy (SEM) and x-ray diffraction (XRD). When SO4 2-is used, irrespective of the presence of other anions, only microspheres of hierarchical Ni(OH)2 nanosheets are present, suggesting that this anion plays a critical role in microsphere formation. Electrochemical characterizations of Ni(OH)2 nanosheets show good supercapacitor performance, with relatively high capacity and excellent rate capability, indicating that these hierarchical Ni(OH)2 nanosheets are serious candidates for energy storage applications. The growth mechanism for nanosheet formation is discussed, based on SEM observations under different preparation conditions, detailing the transition from nanoparticles to nanowires to nanosheets. The specific surface area and the thickness of our Ni(OH)2 nanosheets have been determined to be 149.6 m2 g-1 and 20-30 nm, respectively. © 2015 IOP Publishing Ltd.


Li L.,Rosalind and Morris Goodman Cancer Research Center | Yang X.-J.,Rosalind and Morris Goodman Cancer Research Center | Yang X.-J.,McGill University
Cellular and Molecular Life Sciences | Year: 2015

Microtubules have important functions ranging from maintenance of cell morphology to subcellular transport, cellular signaling, cell migration, and formation of cell polarity. At the organismal level, microtubules are crucial for various biological processes, such as viral entry, inflammation, immunity, learning and memory in mammals. Microtubules are subject to various covalent modifications. One such modification is tubulin acetylation, which is associated with stable microtubules and conserved from protists to humans. In the past three decades, this reversible modification has been studied extensively. In mammals, its level is mainly governed by opposing actions of α-tubulin acetyltransferase 1 (ATAT1) and histone deacetylase 6 (HDAC6). Knockout studies of the mouse enzymes have yielded new insights into biological functions of tubulin acetylation. Abnormal levels of this modification are linked to neurological disorders, cancer, heart diseases and other pathological conditions, thereby yielding important therapeutic implications. This review summarizes related studies and concludes that tubulin acetylation is important for regulating microtubule architecture and maintaining microtubule integrity. Together with detyrosination, glutamylation and other modifications, tubulin acetylation may form a unique 'language' to regulate microtubule structure and function. © 2015 Springer Basel.


Tahmasebi S.,Rosalind and Morris Goodman Cancer Research Center | Ghorbani M.,Rosalind and Morris Goodman Cancer Research Center | Savage P.,Rosalind and Morris Goodman Cancer Research Center | Gocevski G.,Rosalind and Morris Goodman Cancer Research Center | And 2 more authors.
Stem Cells | Year: 2014

Sumoylation adds a small ubiquitin-like modifier (SUMO) polypeptide to the ε-amino group of a lysine residue. Reminiscent of ubiquitination, sumoylation is catalyzed by an enzymatic cascade composed of E1, E2, and E3. For sumoylation, this cascade uses Ubc9 (ubiquitin conjugating enzyme 9, now officially named ubiquitin conjugating enzyme E2I [UBE2I]) as the sole E2 enzyme. Here, we report that expression of endogenous Ubc9 increases during reprogramming of mouse embryonic fibroblasts (MEFs) into induced pluripotent stem (iPS) cells. In addition, this E2 enzyme is required for reprogramming as its suppression dramatically inhibits iPS cell induction. While Ubc9 knockdown does not affect survival of MEFs and immortalized fibroblasts, Ubc9 is essential for embryonic stem cell (ESC) survival. In addition, we have found that Ubc9 knockdown stimulates apoptosis in ESCs but not in MEFs. Furthermore, the knockdown decreases the expression of the well-known pluripotency marker Nanog and the classical reprogramming factors Klf4, Oct4, and Sox2 in ESCs. Together, these observations indicate that while dispensable for fibroblast survival, the sole SUMO E2 enzyme Ubc9 plays a critical role in reprogramming fibroblasts to iPS cells and maintaining ESC pluripotency. © AlphaMed Press 2013.


Pla-Roca M.,McGill University | Leulmi R.F.,McGill University | Tourekhanova S.,McGill University | Bergeron S.,McGill University | And 11 more authors.
Molecular and Cellular Proteomics | Year: 2012

DNA microarrays were rapidly scaled up from 256 to 6.5 million targets, and although antibody microarrays were proposed earlier, sensitive multiplex sandwich assays have only been scaled up to a few tens of targets. Cross-reactivity, arising because detection antibodies are mixed, is a known weakness of multiplex sandwich assays that is mitigated by lengthy optimization. Here, we introduce (1) vulnerability as a metric for assays. The vulnerability of multiplex sandwich assays to cross-reactivity increases quadratically with the number of targets, and together with experimental results, substantiates that scaling up of multiplex sandwich assays is unfeasible. We propose (2) a novel concept for multiplexing without mixing named antibody colocalization microarray (ACM). In ACMs, both capture and detection antibodies are physically colocalized by spotting to the same two-dimensional coordinate. Following spotting of the capture antibodies, the chip is removed from the arrayer, incubated with the sample, placed back onto the arrayer and then spotted with the detection antibodies. ACMs with up to 50 targets were produced, along with a binding curve for each protein. The ACM was validated by comparing it to ELISA and to a small-scale, conventional multiplex sandwich assay (MSA). Using ACMs, proteins in the serum of breast cancer patients and healthy controls were quantified, and six candidate biomarkers identified. Our results indicate that ACMs are sensitive, robust, and scalable. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc.


Archibald A.,Rosalind and Morris Goodman Cancer Research Center | Al-Masri M.,Rosalind and Morris Goodman Cancer Research Center | Liew-Spilger A.,Rosalind and Morris Goodman Cancer Research Center | McCaffrey L.,Rosalind and Morris Goodman Cancer Research Center | McCaffrey L.,McGill University
Molecular Biology of the Cell | Year: 2015

Epithelial cells are major sites of malignant transformation. Atypical protein kinase C (aPKC) isoforms are overexpressed and activated in many cancer types. Using normal, highly polarized epithelial cells (MDCK and NMuMG), we report that aPKC gain of function overcomes contact inhibited growth and is sufficient for a transformed epithelial phenotype. In 2D cultures, aPKC induced cells to grow as stratified epithelia, whereas cells grew as solid spheres of nonpolarized cells in 3D culture. aPKC associated with Mst1/2, which uncoupled Mst1/2 from Lats1/2 and promoted nuclear accumulation of Yap1. Of importance, Yap1 was necessary for aPKC-mediated overgrowth but did not restore cell polarity defects, indicating that the two are separable events. In MDCK cells, Yap1 was sequestered to cell-cell junctions by Amot, and aPKC overexpression resulted in loss of Amot expression and a spindle-like cell phenotype. Reexpression of Amot was sufficient to restore an epithelial cobblestone appearance, Yap1 localization, and growth control. In contrast, the effect of aPKC on Hippo/Yap signaling and overgrowth in NMuMG cells was independent of Amot. Finally, increased expression of aPKC in human cancers strongly correlated with increased nuclear accumulation of Yap1, indicating that the effect of aPKC on transformed growth by deregulating Hippo/ Yap1 signaling may be clinically relevant. © 2015 Mount and Wolin.


Walkinshaw D.R.,Rosalind and Morris Goodman Cancer Research Center | Weist R.,Rosalind and Morris Goodman Cancer Research Center | Weist R.,McGill University | Xiao L.,Rosalind and Morris Goodman Cancer Research Center | And 5 more authors.
Journal of Biological Chemistry | Year: 2013

Background: Nucleocytoplasmic trafficking of class IIa histone deacetylases is crucial for various biological processes. Results: cAMP induces dephosphorylation at an SP motif conserved in HDAC4, HDAC5, and HDAC9 but not in HDAC7. Conclusion: Dephosphorylation at the SP motif governs cAMP sensitivity and nuclear localization of class IIa histone deacetylases. Significance: Cellular signaling pathways may act upon cAMP to promote dephosphorylation and nuclear localization of class IIa histone deacetylases. © 2013 by The American Society for Biochemistry and Molecular Biology, Inc.


PubMed | Rosalind and Morris Goodman Cancer Research Center
Type: Journal Article | Journal: The Journal of biological chemistry | Year: 2013

-Tubulin acetylation at Lys-40, located on the luminal side of microtubules, has been widely studied and used as a marker for stable microtubules in the cilia and other subcellular structures, but the functional consequences remain perplexing. Recent studies have shown that Mec-17 and its paralog are responsible for -tubulin acetylation in Caenorhabditis elegans. There is one such protein known as Atat1 (-tubulin acetyltransferase 1) per higher organism. Zebrafish Atat1 appears to govern embryo development, raising the intriguing possibility that Atat1 is also critical for development in mammals. In addition to Atat1, three other mammalian acetyltransferases, ARD1-NAT1, ELP3, and GCN5, have been shown to acetylate -tubulin in vitro, so an important question is how these four enzymes contribute to the acetylation in vivo. We demonstrate here that Atat1 is a major -tubulin acetyltransferase in mice. It is widely expressed in mouse embryos and tissues. Although Atat1-null animals display no overt phenotypes, -tubulin acetylation is lost in sperm flagella and the dentate gyrus is slightly deformed. Furthermore, human ATAT1 colocalizes on bundled microtubules with doublecortin. These results thus suggest that mouse Atat1 may regulate advanced functions such as learning and memory, thereby shedding novel light on the physiological roles of -tubulin acetylation in mammals.


PubMed | Rosalind and Morris Goodman Cancer Research Center
Type: Journal Article | Journal: Metabolites | Year: 2014

Mitochondria are a focal point in metabolism, given that they play fundamental roles in catabolic, as well as anabolic reactions. Alterations in mitochondrial functions are often studied in whole cells, and metabolomics experiments using 13C-labeled substrates, coupled with mass isotopomer distribution analyses, represent a powerful approach to study global changes in cellular metabolic activities. However, little is known regarding the assessment of metabolic activities in isolated mitochondria using this technology. Studies on isolated mitochondria permit the evaluation of whether changes in cellular metabolic activities are due to modifications in the intrinsic properties of the mitochondria. Here, we present a streamlined approach to accurately determine 13C, as well as 12C enrichments in isolated mitochondria from mammalian tissues or cultured cells by GC/MS. We demonstrate the relevance of this experimental approach by assessing the effects of drugs perturbing mitochondrial functions on the mass isotopomer enrichment of metabolic intermediates. Furthermore, we investigate 13C and 12C enrichments in mitochondria isolated from cancer cells given the emerging role of metabolic alterations in supporting tumor growth. This original method will provide a very sensitive tool to perform metabolomics studies on isolated mitochondria.


PubMed | Rosalind and Morris Goodman Cancer Research Center
Type: Journal Article | Journal: Cellular and molecular life sciences : CMLS | Year: 2015

Microtubules have important functions ranging from maintenance of cell morphology to subcellular transport, cellular signaling, cell migration, and formation of cell polarity. At the organismal level, microtubules are crucial for various biological processes, such as viral entry, inflammation, immunity, learning and memory in mammals. Microtubules are subject to various covalent modifications. One such modification is tubulin acetylation, which is associated with stable microtubules and conserved from protists to humans. In the past three decades, this reversible modification has been studied extensively. In mammals, its level is mainly governed by opposing actions of -tubulin acetyltransferase 1 (ATAT1) and histone deacetylase 6 (HDAC6). Knockout studies of the mouse enzymes have yielded new insights into biological functions of tubulin acetylation. Abnormal levels of this modification are linked to neurological disorders, cancer, heart diseases and other pathological conditions, thereby yielding important therapeutic implications. This review summarizes related studies and concludes that tubulin acetylation is important for regulating microtubule architecture and maintaining microtubule integrity. Together with detyrosination, glutamylation and other modifications, tubulin acetylation may form a unique language to regulate microtubule structure and function.

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