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Cambridge, United Kingdom

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Blasius M.,The Gurdon Institute | Blasius M.,University of Cambridge | Blasius M.,Danish Cancer Society | Wagner S.A.,Novo Nordisk AS | And 6 more authors.
Genes and Development | Year: 2014

RNA metabolism is altered following DNA damage, but the underlying mechanisms are not well understood. Through a 14-3-3 interaction screen for DNA damageinduced protein interactions in human cells, we identified protein complexes connected to RNA biology. These include the nuclear exosome targeting (NEXT) complex that regulates turnover of noncoding RNAs termed promoter upstream transcripts (PROMPTs).We show that the NEXT subunit RBM7 is phosphorylated upon DNA damage by the MAPKAPK2 kinase and establish that this mediates 14-3-3 binding and decreases PROMPT binding. These findings and our observation that cells lacking RBM7 display DNA damage hypersensitivity link PROMPT turnover to the DNA damage response. © 2014 Blasius et al.


Florindo C.,Instituto Gulbenkian Of Ciencia | Florindo C.,University of Algarve | Perdigao J.,Instituto Gulbenkian Of Ciencia | Fesquet D.,Montpellier University | And 4 more authors.
Journal of Cell Science | Year: 2012

The completion of cytokinesis requires abscission of the midbody, a microtubule-rich cytoplasmic bridge that connects the daughter cells before their final separation. Although it has been established that both the midbody structure and membrane fusion are essential for abscission, the biochemical machinery and the cellular processes of abscission remain ill-defined. Here we report that human Mob1A and Mob1B proteins are involved in the regulation of abscission of the intercellular bridge. The Mob family is a group of highly conserved proteins in eukaryotes, described as binding partners as well as co-activators of protein kinases of the Ndr family, and as members of the Hippo pathway. We show that depletion of Mob1A and Mob1B by RNAi causes abscission failure as a consequence of hyper-stabilization of microtubules in the midbody region. Interestingly, depleting Mob1 also increases cell motility after cytokinesis, and induces prolonged centriole separation in G1 phase. In contrast, centrosomes fail to split when either Mob1A or Mob1B is overexpressed. Our findings indicate that human Mob1 proteins are involved in the regulation of microtubule stability at the midbody. We conclude that Mob1A and Mob1B are needed for cell abscission and centriole re-joining after telophase and cytokinesis. © 2012.


Bergstralh D.T.,The Gurdon Institute | Bergstralh D.T.,University of Cambridge | St Johnston D.,The Gurdon Institute | St Johnston D.,University of Cambridge
Seminars in Cell and Developmental Biology | Year: 2014

The angle of cell division is critical in at least two contexts. It can determine cell fate, as it does in developing neural tissue. It can also dictate tissue architecture, as it does in many epithelia. One way to ensure the correct angle of cell division is through controlled orientation of the spindle at metaphase. What happens when that control is lost? Ongoing work suggests that the consequence of metaphase spindle misorientation may be significant, but multiple mechanisms exist to protect the cell and the tissue. We speculate that one such mechanism involves a recently identified anaphase activity for two of the key players at metaphase: NuMA (Mud, LIN-5) and dynein. © 2014 The Authors.Published by Elsevier Ltd.


Wieser S.,The Gurdon Institute | Pines J.,The Gurdon Institute
Cold Spring Harbor Perspectives in Biology | Year: 2015

In this article, wewill discuss the biochemistryof mitosis in eukaryotic cells. We will focus on conserved principles that, importantly, are adapted to the biologyof the organism. It is vital to bear in mind that the structural requirements for division in a rapidly dividing syncytial Drosophila embryo, for example, are markedly different from those in a unicellular yeast cell. Nevertheless, division in both systems is driven by conserved modules of antagonistic protein kinases and phosphatases, underpinned by ubiquitin-mediated proteolysis, which create molecular switches to drive each stage of division forward. These conserved control modules combine with the self-organizing properties of the subcellular architecture to meet the specific needs of the cell. Our discussion will draw on discoveries in several model systems that have been important in the long history of research on mitosis, and we will try to point out those principles that appear to apply to all cells, compared with those in which the biochemistry has been specifically adapted in a particular organism. © 2015 Cold Spring Harbor Laboratory Press. All rights reserved.


PubMed | The Gurdon Institute
Type: Journal Article | Journal: The Journal of cell biology | Year: 2010

In Caenorhabditis elegans, five proteins are required for centriole duplication: SPD-2, ZYG-1, SAS-5, SAS-6, and SAS-4. Functional orthologues of all but SAS-5 have been found in other species. In Drosophila melanogaster and humans, Sak/Plk4, DSas-6/hSas-6, and DSas-4/CPAP-orthologues of ZYG-1, SAS-6, and SAS-4, respectively-are required for centriole duplication. Strikingly, all three fly proteins can induce the de novo formation of centriole-like structures when overexpressed in unfertilized eggs. Here, we find that of eight candidate duplication factors identified in cultured fly cells, only two, Ana2 and Asterless (Asl), share this ability. Asl is now known to be essential for centriole duplication in flies, but no equivalent protein has been found in worms. We show that Ana2 is the likely functional orthologue of SAS-5 and that it is also related to the vertebrate STIL/SIL protein family that has been linked to microcephaly in humans. We propose that members of the SAS-5/Ana2/STIL family of proteins are key conserved components of the centriole duplication machinery.


PubMed | The Gurdon Institute
Type: Journal Article | Journal: Cell | Year: 2010

Modifications on histones or on DNA recruit proteins that regulate chromatin function. Here, we use nucleosomes methylated on DNA and on histone H3 in an affinity assay, in conjunction with a SILAC-based proteomic analysis, to identify crosstalk between these two distinct classes of modification. Our analysis reveals proteins whose binding to nucleosomes is regulated by methylation of CpGs, H3K4, H3K9, and H3K27 or a combination thereof. We identify the origin recognition complex (ORC), including LRWD1 as a subunit, to be a methylation-sensitive nucleosome interactor that is recruited cooperatively by DNA and histone methylation. Other interactors, such as the lysine demethylase Fbxl11/KDM2A, recognize nucleosomes methylated on histones, but their recruitment is disrupted by DNA methylation. These data establish SILAC nucleosome affinity purifications (SNAP) as a tool for studying the dynamics between different chromatin modifications and provide a modification binding profile for proteins regulated by DNA and histone methylation.


PubMed | The Gurdon Institute
Type: Journal Article | Journal: Cold Spring Harbor perspectives in biology | Year: 2015

In this article, we will discuss the biochemistry of mitosis in eukaryotic cells. We will focus on conserved principles that, importantly, are adapted to the biology of the organism. It is vital to bear in mind that the structural requirements for division in a rapidly dividing syncytial Drosophila embryo, for example, are markedly different from those in a unicellular yeast cell. Nevertheless, division in both systems is driven by conserved modules of antagonistic protein kinases and phosphatases, underpinned by ubiquitin-mediated proteolysis, which create molecular switches to drive each stage of division forward. These conserved control modules combine with the self-organizing properties of the subcellular architecture to meet the specific needs of the cell. Our discussion will draw on discoveries in several model systems that have been important in the long history of research on mitosis, and we will try to point out those principles that appear to apply to all cells, compared with those in which the biochemistry has been specifically adapted in a particular organism.

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