24 results found
Mahen R, 2021, The structure and function of centriolar rootlets, JOURNAL OF CELL SCIENCE, Vol: 134, ISSN: 0021-9533
Lee M, Shorthouse D, Mahen R, et al., 2021, Cancer-causing BRCA2 missense mutations disrupt an intracellular protein assembly mechanism to disable genome maintenance, NUCLEIC ACIDS RESEARCH, Vol: 49, Pages: 5588-5604, ISSN: 0305-1048
Nicolai S, Mahen R, Raschella G, et al., 2020, ZNF281 is recruited on DNA breaks to facilitate DNA repair by non-homologous end joining, ONCOGENE, Vol: 39, Pages: 754-766, ISSN: 0950-9232
Mahen R, Schulte R, 2019, Cell-cell Fusion of Genome Edited Cell Lines for Perturbation of Cellular Structure and Function, JOVE-JOURNAL OF VISUALIZED EXPERIMENTS, ISSN: 1940-087X
Sharma P, Mahen R, Rossmann M, et al., 2019, A cryptic hydrophobic pocket in the polo-box domain of the polo-like kinase PLK1 regulates substrate recognition and mitotic chromosome segregation, SCIENTIFIC REPORTS, Vol: 9, ISSN: 2045-2322
Mahen RWJ, 2018, Stable centrosomal roots disentangle to allow interphase centriole independence, PLoS Biology, Vol: 16, ISSN: 1544-9173
The centrosome is a non-membrane bound cellular compartment consisting of two centrioles surrounded by a proteincoat termed the pericentriolar material (PCM). Centrioles generally remain physically associated together (a phenomenoncalled centrosome cohesion), yet how this occurs in the absence of a bounding lipid membrane is unclear. One modelposits that pericentriolar fibres formed from rootletin protein directly link centrioles, yet little is known about thestructure, biophysical properties or assembly kinetics of such fibres. Here I combine live cell imaging of endogenouslytagged rootletin with cell fusion, and find previously unrecognised plasticity in centrosome cohesion. Rootletin formslarge, diffusionally stable, bifurcating fibres, which amass slowly on mature centrioles over many hours from anaphase.Nascent centrioles (procentrioles) in contrast do not form roots, and must be licensed to do so through polo-like kinase 1(PLK1) activity. Transient separation of roots accompanies centriolar repositioning during the interphase, suggesting thatcentrioles organize as independent units, each containing discrete roots. Indeed, forced induction of duplicate centriolepairs allows independent re-shuffling of individual centrioles between the pairs. Thus collectively, these findings suggestthat progressively nucleated polymers mediate the dynamic association of centrioles as either one or two interphasecentrosomes, with implications for the understanding of how non-membrane bound organelles self-organise.
Isokane M, Walter T, Mahen R, et al., 2016, ARHGEF17 is an essential spindle assembly checkpoint factor that targets Mps1 to kinetochores, The Journal of Cell Biology, Vol: 212, Pages: 647-659, ISSN: 0021-9525
To prevent genome instability, mitotic exit is delayed until all chromosomes are properly attached to the mitotic spindle by the spindle assembly checkpoint (SAC). In this study, we characterized the function of ARHGEF17, identified in a genome-wide RNA interference screen for human mitosis genes. Through a series of quantitative imaging, biochemical, and biophysical experiments, we showed that ARHGEF17 is essential for SAC activity, because it is the major targeting factor that controls localization of the checkpoint kinase Mps1 to the kinetochore. This mitotic function is mediated by direct interaction of the central domain of ARHGEF17 with Mps1, which is autoregulated by the activity of Mps1 kinase, for which ARHGEF17 is a substrate. This mitosis-specific role is independent of ARHGEF17’s RhoGEF activity in interphase. Our study thus assigns a new mitotic function to ARHGEF17 and reveals the molecular mechanism for a key step in SAC establishment.
Wachsmuth M, Conrad C, Bulkescher J, et al., 2015, High-throughput fluorescence correlation spectroscopy enables analysis of proteome dynamics in living cells, NATURE BIOTECHNOLOGY, Vol: 33, Pages: 384-U91, ISSN: 1087-0156
Mahen R, Koch B, Wachsmuth M, et al., 2014, Comparative assessment of fluorescent transgene methods for quantitative imaging in human cells, MOLECULAR BIOLOGY OF THE CELL, Vol: 25, Pages: 3610-3618, ISSN: 1059-1524
Keller D, Orpinell M, Olivier N, et al., 2014, Mechanisms of HsSAS-6 assembly promoting centriole formation in human cells, JOURNAL OF CELL BIOLOGY, Vol: 204, Pages: 697-712, ISSN: 0021-9525
Mahen R, Hattori H, Lee M, et al., 2013, A-Type Lamins Maintain the Positional Stability of DNA Damage Repair Foci in Mammalian Nuclei, PLOS ONE, Vol: 8, ISSN: 1932-6203
Bolderson E, Savage KI, Mahen R, et al., 2012, Kruppel-associated Box (KRAB)-associated co-repressor (KAP-1) Ser-473 phosphorylation regulates heterochromatin protein 1β (HP1-β) mobilization and DNA repair in heterochromatin., J Biol Chem, Vol: 287, Pages: 28122-28131
The DNA damage response encompasses a complex series of signaling pathways that function to regulate and facilitate the repair of damaged DNA. Recent studies have shown that the repair of transcriptionally inactive chromatin, named heterochromatin, is dependent upon the phosphorylation of the co-repressor, Krüppel-associated box (KRAB) domain-associated protein (KAP-1), by the ataxia telangiectasia-mutated (ATM) kinase. Co-repressors, such as KAP-1, function to regulate the rigid structure of heterochromatin by recruiting histone-modifying enzymes, such HDAC1/2, SETDB1, and nucleosome-remodeling complexes such as CHD3. Here, we have characterized a phosphorylation site in the HP1-binding domain of KAP-1, Ser-473, which is phosphorylated by the cell cycle checkpoint kinase Chk2. Expression of a nonphosphorylatable S473A mutant conferred cellular sensitivity to DNA-damaging agents and led to defective repair of DNA double-strand breaks in heterochromatin. In addition, cells expressing S473A also displayed defective mobilization of the HP1-β chromodomain protein. The DNA repair defect observed in cells expressing S473A was alleviated by depletion of HP1-β, suggesting that phosphorylation of KAP-1 on Ser-473 promotes the mobilization of HP1-β from heterochromatin and subsequent DNA repair. These results suggest a novel mechanism of KAP-1-mediated chromatin restructuring via Chk2-regulated HP1-β exchange from heterochromatin, promoting DNA repair.
Mahen R, Venkitaraman AR, 2012, Pattern formation in centrosome assembly, CURRENT OPINION IN CELL BIOLOGY, Vol: 24, Pages: 14-23, ISSN: 0955-0674
James JR, McColl J, Oliveira MI, et al., 2011, The T Cell Receptor Triggering Apparatus Is Composed of Monovalent or Monomeric Proteins, JOURNAL OF BIOLOGICAL CHEMISTRY, Vol: 286, Pages: 31993-32001
Mahen R, Jeyasekharan AD, Barry NP, et al., 2011, Continuous polo-like kinase 1 activity regulates diffusion to maintain centrosome self-organization during mitosis, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, Vol: 108, Pages: 9310-9315, ISSN: 0027-8424
Amorim MJ, Bruce EA, Read EKC, et al., 2011, A Rab11-and Microtubule-Dependent Mechanism for Cytoplasmic Transport of Influenza A Virus Viral RNA, JOURNAL OF VIROLOGY, Vol: 85, Pages: 4143-4156, ISSN: 0022-538X
Jeyasekharana AD, Ayoub N, Mahen R, et al., 2010, DNA damage regulates the mobility of Brca2 within the nucleoplasm of living cells, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, Vol: 107, Pages: 21937-21942, ISSN: 0027-8424
Ayoub N, Rajendra E, Su X, et al., 2009, The carboxyl terminus of Brca2 links the disassembly of Rad51 complexes to mitotic entry., Curr Biol, Vol: 19, Pages: 1075-1085
BACKGROUND: The Rad51 recombinase assembles on DNA to execute homologous DNA recombination (HR). This process is essential to repair replication-associated genomic lesions before cells enter mitosis, but how it is started and stopped during the cell cycle remains poorly understood. Rad51 assembly is regulated by the breast cancer suppressor Brca2, via its evolutionarily conserved BRC repeats, and a distinct carboxy (C)-terminal motif whose biological function is uncertain. Using "hit-and-run" gene targeting to insert single-codon substitutions into the avian Brca2 locus, we report here a previously unrecognized role for the C-terminal motif. RESULTS: We show that the avian C-terminal motif is functionally cognate with its human counterpart and identify point mutations that either abolish or enhance Rad51 binding. When these mutations are introduced into Brca2, we find that they affect neither the assembly of Rad51 into nuclear foci on damaged DNA nor DNA repair by HR. Instead, foci disassemble more rapidly in a point mutant that fails to bind Rad51, associated with faster mitotic entry. Conversely, the slower disassembly of foci in a point mutant that constitutively binds Rad51 correlates with delayed mitosis. Indeed, Rad51 foci do not persist in mitotic cells even after G2 checkpoint suppression, suggesting that their disassembly is a prerequisite for chromosome segregation. CONCLUSIONS: We conclude that Rad51 binding by the C-terminal Brca2 motif is dispensable for the execution of HR but instead links the disassembly of Rad51 complexes to mitotic entry. This mechanism may ensure that HR terminates before chromosome segregation. Our findings assign a biological function for the C-terminal Brca2 motif in a mechanism that coordinates DNA repair with the cell cycle.
Palmer CP, Mahen R, Schnell E, et al., 2007, Sigma-1 receptors bind cholesterol and remodel lipid rafts in breast cancer cell lines, CANCER RESEARCH, Vol: 67, Pages: 11166-11175, ISSN: 0008-5472
Mahen R, A DNA-based voltmeter for organelles
<jats:title>Abstract</jats:title><jats:p>Centrioles are non-membrane bound organelles that participate in fundamental cellular processes through their ability to form physical contacts with other structures. During interphase, two mature centrioles can associate to form a single centrosome - a phenomenon known as centrosome cohesion. Centrosome cohesion is important for processes such as cell migration, and yet how it is maintained is unclear. Current models indicate that pericentriolar fibres termed rootlets, also known as the centrosome linker, entangle to maintain centriole proximity. Here, I uncover a new centriole-centriole contact site and mechanism of centrosome cohesion, based on coalescence of the proximal centriole component cNap1. Using live-cell imaging of endogenously tagged cNap1, I show that proximal centrioles form dynamic contacts in response to physical force from the cytoskeleton. Expansion microscopy reveals that cNap1 bridges between these contact sites, physically linking proximal centrioles on the nanoscale. When ectopically tethered to organelles such as lysosomes, cNap1 forms viscous and cohesive condensates that promote organelle spatial proximity. Conversely, cNap1 mutants with reduced viscosity are unable to maintain centrosome cohesion. These results define a previously unrecognised mechanism of centrosome cohesion by cNap1 assemblies at the proximal centriole and illustrate how a non-membrane bound organelle forms dynamic organelle contact sites.</jats:p>
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