@article{pmid26525406,

title = {SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy},

author = {Graeme Ball and Justin Demmerle and Rainer Kaufmann and Ilan Davis and Ian M Dobbie and Lothar Schermelleh},

doi = {10.1038/srep15915},

issn = {2045-2322},

year  = {2015},

date = {2015-11-01},

journal = {Sci Rep},

volume = {5},

pages = {15915},

abstract = {Three-dimensional structured illumination microscopy (3D-SIM) is a versatile and accessible method for super-resolution fluorescence imaging, but generating high-quality data is challenging, particularly for non-specialist users. We present SIMcheck, a suite of ImageJ plugins enabling users to identify and avoid common problems with 3D-SIM data, and assess resolution and data quality through objective control parameters. Additionally, SIMcheck provides advanced calibration tools and utilities for common image processing tasks. This open-source software is applicable to all commercial and custom platforms, and will promote routine application of super-resolution SIM imaging in cell biology.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26486521,

title = {Bioprinting of human pluripotent stem cells and their directed differentiation into hepatocyte-like cells for the generation of mini-livers in 3D},

author = {Alan Faulkner-Jones and Catherine Fyfe and Dirk-Jan Cornelissen and John Gardner and Jason King and Aidan Courtney and Wenmiao Shu},

doi = {10.1088/1758-5090/7/4/044102},

issn = {1758-5090},

year  = {2015},

date = {2015-10-01},

journal = {Biofabrication},

volume = {7},

number = {4},

pages = {044102},

abstract = {We report the first investigation into the bioprinting of human induced pluripotent stem cells (hiPSCs), their response to a valve-based printing process as well as their post-printing differentiation into hepatocyte-like cells (HLCs). HLCs differentiated from both hiPSCs and human embryonic stem cells (hESCs) sources were bioprinted and examined for the presence of hepatic markers to further validate the compatibility of the valve-based bioprinting process with fragile cell transfer. Examined cells were positive for nuclear factor 4 alpha and were demonstrated to secrete albumin and have morphology that was also found to be similar to that of hepatocytes. Both hESC and hiPSC lines were tested for post-printing viability and pluripotency and were found to have negligible difference in terms of viability and pluripotency between the printed and non-printed cells. hESC-derived HLCs were 3D printed using alginate hydrogel matrix and tested for viability and albumin secretion during the remaining differentiation and were found to be hepatic in nature. 3D printed with 40-layer of HLC-containing alginate structures reached peak albumin secretion at day 21 of the differentiation protocol. This work demonstrates that the valve-based printing process is gentle enough to print human pluripotent stem cells (hPSCs) (both hESCs and hiPSCs) while either maintaining their pluripotency or directing their differentiation into specific lineages. The ability to bioprint hPSCs will pave the way for producing organs or tissues on demand from patient specific cells which could be used for animal-free drug development and personalized medicine.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26071977,

title = {The evolution of structured illumination microscopy in studies of HIV},

author = {Kelly Marno and Lara Al'Zoubi and Matthew Pearson and Markus Posch and Áine McKnight and Ann P Wheeler},

doi = {10.1016/j.ymeth.2015.06.007},

issn = {1095-9130},

year  = {2015},

date = {2015-10-01},

journal = {Methods},

volume = {88},

pages = {20--27},

abstract = {The resolution limit of conventional light microscopy has proven to be limiting for many biological structures such as viruses including Human immunodeficiency virus (HIV). Individual HIV virions are impossible to study using confocal microscopy as they are well below the 200 nm resolution limit of conventional light microscopes. Structured illumination microscopy (SIM) allows a twofold enhancement in image resolution compared to standard widefield illumination and so provides an excellent tool for study of HIV. Viral capsids (CAs) vary between 110 and 146 nm so this study challenges the performance of SIM microscopes. SIM microscopy was first developed in 2000, commercialised in 2007 and rapidly developed. Here we present the changes in capabilities of the SIM microscopes for study of HIV localisation as the instrumentation for structured illumination microscopy has evolved over the past 8 years.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{nokey,

title = {Observing the Dynamics of Waterborne Pathogens for Assessing the Level of Contamination},

author = {McKenna, Isabella and Tonolini, Francesco and Tobin, Rachael and Houssineau, Jeremie and Bridle, Helen and McDougall, Craig and Schlangen, Isabel and McGrath, John S. and Jimenez, Melanie and Clark, Daniel E},

doi = {10.1109/SSPD.2015.7288514},

year  = {2015},

date = {2015-09-09},

urldate = {2015-09-09},

journal = {2015 Sensor Signal Processing for Defence (SSPD)},

pages = {1-5},

abstract = {In environments of scarce hygiene it is of primary importance to detect potentially harmful concentrations of pathogens in drinking water. In many situations, however, accurate analysis of water samples is prohibitively complex and often requires highly specialised apparatuses and technicians. In order to overcome these limitations, a method to employ video processing to assist microfluidics water filtering apparatuses is proposed. Through the automated analysis of videos captured at the output of such devices it is possible to extract useful information that could control an autonomous calibration, hence eliminating the need of an expert and possibly leading to the construction of readily employable water quality assessing devices.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26341556,

title = {A negative loop within the nuclear pore complex controls global chromatin organization},

author = {Manuel Breuer and Hiroyuki Ohkura},

doi = {10.1101/gad.264341.115},

issn = {1549-5477},

year  = {2015},

date = {2015-09-01},

journal = {Genes Dev},

volume = {29},

number = {17},

pages = {1789--1794},

abstract = {The nuclear pore complex (NPC) tethers chromatin to create an environment for gene regulation, but little is known about how this activity is regulated to avoid excessive tethering of the genome. Here we propose a negative regulatory loop within the NPC controlling the chromatin attachment state, in which Nup155 and Nup93 recruit Nup62 to suppress chromatin tethering by Nup155. Depletion of Nup62 severely disrupts chromatin distribution in the nuclei of female germlines and somatic cells, which can be reversed by codepleting Nup155. Thus, this universal regulatory system within the NPC is crucial to control large-scale chromatin organization in the nucleus.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26386247,

title = {TALPID3 controls centrosome and cell polarity and the human ortholog KIAA0586 is mutated in Joubert syndrome (JBTS23)},

author = {Louise A Stephen and Hasan Tawamie and Gemma M Davis and Lars Tebbe and Peter Nürnberg and Gudrun Nürnberg and Holger Thiele and Michaela Thoenes and Eugen Boltshauser and Steffen Uebe and Oliver Rompel and André Reis and Arif B Ekici and Lynn McTeir and Amy M Fraser and Emma A Hall and Pleasantine Mill and Nicolas Daudet and Courtney Cross and Uwe Wolfrum and Rami Abou Jamra and Megan G Davey and Hanno J Bolz},

doi = {10.7554/eLife.08077},

issn = {2050-084X},

year  = {2015},

date = {2015-09-01},

journal = {Elife},

volume = {4},

abstract = {Joubert syndrome (JBTS) is a severe recessive neurodevelopmental ciliopathy which can affect several organ systems. Mutations in known JBTS genes account for approximately half of the cases. By homozygosity mapping and whole-exome sequencing, we identified a novel locus, JBTS23, with a homozygous splice site mutation in KIAA0586 (alias TALPID3), a known lethal ciliopathy locus in model organisms. Truncating KIAA0586 mutations were identified in two additional patients with JBTS. One mutation, c.428delG (p.Arg143Lysfs*4), is unexpectedly common in the general population and may be a major contributor to JBTS. We demonstrate KIAA0586 protein localization at the basal body in human and mouse photoreceptors, as is common for JBTS proteins, and also in pericentriolar locations. We show that loss of TALPID3 (KIAA0586) function in animal models causes abnormal tissue polarity, centrosome length and orientation, and centriolar satellites. We propose that JBTS and other ciliopathies may in part result from cell polarity defects.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26387948,

title = {Visualization of the Serratia Type VI Secretion System Reveals Unprovoked Attacks and Dynamic Assembly},

author = {Amy J Gerc and Andreas Diepold and Katharina Trunk and Michael Porter and Colin Rickman and Judith P Armitage and Nicola R Stanley-Wall and Sarah J Coulthurst},

doi = {10.1016/j.celrep.2015.08.053},

issn = {2211-1247},

year  = {2015},

date = {2015-09-01},

journal = {Cell Rep},

volume = {12},

number = {12},

pages = {2131--2142},

abstract = {The Type VI secretion system (T6SS) is a bacterial nanomachine that fires toxic proteins into target cells. Deployment of the T6SS represents an efficient and widespread means by which bacteria attack competitors or interact with host organisms and may be triggered by contact from an attacking neighbor cell as a defensive strategy. Here, we use the opportunist pathogen Serratia marcescens and functional fluorescent fusions of key components of the T6SS to observe different subassemblies of the machinery simultaneously and on multiple timescales in vivo. We report that the localization and dynamic behavior of each of the components examined is distinct, revealing a multi-stage and dynamic assembly process for the T6SS machinery. We also show that the T6SS can assemble and fire without needing a cell contact trigger, defining an aggressive strategy that broadens target range and suggesting that activation of the T6SS is tailored to survival in specific niches.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26170147,

title = {Chromatin at the nuclear periphery and the regulation of genome functions},

author = {Charlene Lemaître and Wendy A Bickmore},

doi = {10.1007/s00418-015-1346-y},

issn = {1432-119X},

year  = {2015},

date = {2015-08-01},

journal = {Histochem Cell Biol},

volume = {144},

number = {2},

pages = {111--122},

abstract = {Chromatin is not randomly organized in the nucleus, and its spatial organization participates in the regulation of genome functions. However, this spatial organization is also not entirely fixed and modifications of chromatin architecture are implicated in physiological processes such as differentiation or senescence. One of the most striking features of chromatin architecture is the concentration of heterochromatin at the nuclear periphery. A closer examination of the association of chromatin at the nuclear periphery reveals that heterochromatin accumulates at the nuclear lamina, whereas nuclear pores are usually devoid of heterochromatin. After summarizing the current techniques used to study the attachment of chromatin at the nuclear lamina or the nuclear pores, we review the mechanisms underlying these attachments, their plasticity and their consequences on the regulation of gene expression, DNA repair and replication.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid25656851,

title = {Rapid formation of a supramolecular polypeptide-DNA hydrogel for in situ three-dimensional multilayer bioprinting},

author = {Chuang Li and Alan Faulkner-Jones and Alison R Dun and Juan Jin and Ping Chen and Yongzheng Xing and Zhongqiang Yang and Zhibo Li and Wenmiao Shu and Dongsheng Liu and Rory R Duncan},

doi = {10.1002/anie.201411383},

issn = {1521-3773},

year  = {2015},

date = {2015-03-01},

journal = {Angew Chem Int Ed Engl},

volume = {54},

number = {13},

pages = {3957--3961},

abstract = {A rapidly formed supramolecular polypeptide-DNA hydrogel was prepared and used for in situ multilayer three-dimensional bioprinting for the first time. By alternative deposition of two complementary bio-inks, designed structures can be printed. Based on their healing properties and high mechanical strengths, the printed structures are geometrically uniform without boundaries and can keep their shapes up to the millimeter scale without collapse. 3D cell printing was demonstrated to fabricate live-cell-containing structures with normal cellular functions. Together with the unique properties of biocompatibility, permeability, and biodegradability, the hydrogel becomes an ideal biomaterial for 3D bioprinting to produce designable 3D constructs for applications in tissue engineering.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26590168,

title = {Regulatory Domains and Their Mechanisms},

author = {Nezha S Benabdallah and Wendy A Bickmore},

doi = {10.1101/sqb.2015.80.027268},

issn = {1943-4456},

year  = {2015},

date = {2015-01-01},

journal = {Cold Spring Harb Symp Quant Biol},

volume = {80},

pages = {45--51},

abstract = {The concept of gene regulation is being refined as our understanding of the role of enhancer elements grows. Although described more than 30 years ago, the mechanisms through which these cis-regulating elements operate remain under debate. With the recognition that most of the human genetic variation contributing to common disease risk lies outside of genes and probably in enhancers, unraveling these mechanisms becomes ever more important. Originally, a popular view was to consider regulatory elements as an entry site for the transcription machinery that could scan the intervening chromatin until the cognate core promoter was located. Now, the most prominent model for distal enhancer-promoter interaction involves direct enhancer/promoter contacts with a looping out of intervening chromatin. However, a rising awareness of the importance of chromatin architecture and organization forces us to consider enhancer-promoter communication in light of the polymer folding properties of chromatin. Here, we discuss how three-dimensional chromatin folding, topological domains, and the constrained motion, plasticity, and accessibility of chromatin could offer a structural basis for regulatory domains that greatly enhances the probability of enhancer-promoter and transcription factor-promoter interactions and gene activation.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid25517944,

title = {A molecular toggle after exocytosis sequesters the presynaptic syntaxin1a molecules involved in prior vesicle fusion},

author = {Deirdre M Kavanagh and Annya M Smyth and Kirsty J Martin and Alison Dun and Euan R Brown and Sarah Gordon and Karen J Smillie and Luke H Chamberlain and Rhodri S Wilson and Lei Yang and Weiping Lu and Michael A Cousin and Colin Rickman and Rory R Duncan},

doi = {10.1038/ncomms6774},

issn = {2041-1723},

year  = {2014},

date = {2014-12-01},

journal = {Nat Commun},

volume = {5},

pages = {5774},

abstract = {Neuronal synapses are among the most scrutinized of cellular systems, serving as a model for all membrane trafficking studies. Despite this, synaptic biology has proven difficult to interrogate directly in situ due to the small size and dynamic nature of central synapses and the molecules within them. Here we determine the spatial and temporal interaction status of presynaptic proteins, imaging large cohorts of single molecules inside active synapses. Measuring rapid interaction dynamics during synaptic depolarization identified the small number of syntaxin1a and munc18-1 protein molecules required to support synaptic vesicle exocytosis. After vesicle fusion and subsequent SNARE complex disassembly, a prompt switch in syntaxin1a and munc18-1-binding mode, regulated by charge alteration on the syntaxin1a N-terminal, sequesters monomeric syntaxin1a from other disassembled fusion complex components, preventing ectopic SNARE complex formation, readying the synapse for subsequent rounds of neurotransmission.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid25359883,

title = {Eps8 controls Src- and FAK-dependent phenotypes in squamous carcinoma cells},

author = {Christina Schoenherr and Bryan Serrels and Charlotte Proby and Debbie L Cunningham and Jane E Findlay and George S Baillie and John K Heath and Margaret C Frame},

doi = {10.1242/jcs.157560},

issn = {1477-9137},

year  = {2014},

date = {2014-12-01},

journal = {J Cell Sci},

volume = {127},

number = {Pt 24},

pages = {5303--5316},

abstract = {Eps8 is an actin regulatory scaffold protein whose expression is increased in squamous cell carcinoma (SCC) cells. It forms a complex with both focal adhesion kinase (FAK, also known as PTK2) and Src in SCC cells derived from skin carcinomas induced by administration of the chemical DMBA followed by TPA (the DMBA/TPA model). Here, we describe two new roles for Eps8. Firstly, it controls the spatial distribution of active Src in a FAK-dependent manner. Specifically, Eps8 participates in, and regulates, a biochemical complex with Src and drives trafficking of Src to autophagic structures that SCC cells use to cope with high levels of active Src when FAK is absent. Secondly, when FAK is expressed in SCC cells, thereby meaning active Src becomes tethered at focal adhesion complexes, Eps8 is also recruited to focal adhesions and is required for FAK-dependent polarization and invasion. Therefore, Eps8 is a crucial mediator of Src- and FAK-regulated processes; it participates in specific biochemical complexes and promotes actin re-arrangements that determine the spatial localization of Src, and modulates the functions of Src and FAK during invasive migration.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid24027557,

title = {Imaging large cohorts of single ion channels and their activity},

author = {Katia Hiersemenzel and Euan R Brown and Rory R Duncan},

doi = {10.3389/fendo.2013.00114},

issn = {1664-2392},

year  = {2013},

date = {2013-09-01},

journal = {Front Endocrinol (Lausanne)},

volume = {4},

pages = {114},

abstract = {As calcium is the most important signaling molecule in neurons and secretory cells, amongst many other cell types, it follows that an understanding of calcium channels and their regulation of exocytosis is of vital importance. Calcium imaging using calcium dyes such as Fluo3, or FRET-based dyes that have been used widely has provided invaluable information, which combined with modeling has estimated the subtypes of channels responsible for triggering the exocytotic machinery as well as inferences about the relative distances away from vesicle fusion sites these molecules adopt. Importantly, new super-resolution microscopy techniques, combined with novel Ca(2+) indicators and imaginative imaging approaches can now define directly the nano-scale locations of very large cohorts of single channel molecules in relation to single vesicles. With combinations of these techniques the activity of individual channels can be visualized and quantified using novel Ca(2+) indicators. Fluorescently labeled specific channel toxins can also be used to localize endogenous assembled channel tetramers. Fluorescence lifetime imaging microscopy and other single-photon-resolution spectroscopic approaches offer the possibility to quantify protein-protein interactions between populations of channels and the SNARE protein machinery for the first time. Together with simultaneous electrophysiology, this battery of quantitative imaging techniques has the potential to provide unprecedented detail describing the locations, dynamic behaviors, interactions, and conductance activities of many thousands of channel molecules and vesicles in living cells.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid23929972,

title = {Transcription. Flashing a light on the spatial organization of transcription},

author = {Colin Rickman and Wendy A Bickmore},

doi = {10.1126/science.1242889},

issn = {1095-9203},

year  = {2013},

date = {2013-08-01},

journal = {Science},

volume = {341},

number = {6146},

pages = {621--622},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid23223447,

title = {Munc18-1 protein molecules move between membrane molecular depots distinct from vesicle docking sites},

author = {Annya M Smyth and Lei Yang and Kirsty J Martin and Charlotte Hamilton and Weiping Lu and Michael A Cousin and Colin Rickman and Rory R Duncan},

doi = {10.1074/jbc.M112.407585},

issn = {1083-351X},

year  = {2013},

date = {2013-02-01},

journal = {J Biol Chem},

volume = {288},

number = {7},

pages = {5102--5113},

abstract = {Four evolutionarily conserved proteins are required for mammalian regulated exocytosis: three SNARE proteins, syntaxin, SNAP-25, and synaptobrevin, and the SM protein, Munc18-1. Here, using single-molecule imaging, we measured the spatial distribution of large cohorts of single Munc18-1 molecules correlated with the positions of single secretory vesicles in a functionally rescued Munc18-1-null cellular model. Munc18-1 molecules were nonrandomly distributed across the plasma membrane in a manner not directed by mode of interaction with syntaxin1, with a small mean number of molecules observed to reside under membrane resident vesicles. Surprisingly, we found that the majority of vesicles in fully secretion-competent cells had no Munc18-1 associated within distances relevant to plasma membrane-vesicle SNARE interactions. Live cell imaging of Munc18-1 molecule dynamics revealed that the density of Munc18-1 molecules at the plasma membrane anticorrelated with molecular speed, with single Munc18-1 molecules displaying directed motion between membrane hotspots enriched in syntaxin1a. Our findings demonstrate that Munc18-1 molecules move between membrane depots distinct from vesicle morphological docking sites.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid23804033,

title = {Kindlin-1 regulates mitotic spindle formation by interacting with integrins and Plk-1},

author = {Hitesh Patel and Judith Zich and Bryan Serrels and Colin Rickman and Kevin G Hardwick and Margaret C Frame and Valerie G Brunton},

doi = {10.1038/ncomms3056},

issn = {2041-1723},

year  = {2013},

date = {2013-01-01},

journal = {Nat Commun},

volume = {4},

pages = {2056},

abstract = {Kindlin-1 binds to integrins and regulates integrin activation at cell adhesions. Here we report a new function of Kindlin-1 in regulating spindle assembly. We show that Kindlin-1 localizes to centrosomes, its concentration peaking during G2/M, where it associates with various pericentriolar material proteins, including Polo-like kinase 1. Short interfering RNA-mediated depletion of Kindlin-1 increases formation of abnormal mitotic spindles and decreases cellular survival. This effect is dependent not only on the ability of Kindlin-1 to bind integrins but also on Polo-like kinase 1-mediated Kindlin-1 phosphorylation. We demonstrate that a subcellular pool of phosphorylated Kindlin-1 is located exclusively at centrosomes. Our work identifies a novel cellular role for Kindlin-1 in ensuring mitotic spindle assembly and cellular survival that is controlled by phosphorylation via Polo-like kinase 1.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}