@article {69219, title = {More than just {\textquoteright}added value{\textquoteright}: The perils of not establishing shared core facilities in resource-constrained communities.}, journal = {Journal of Microscopy}, year = {2024}, month = {2024 Feb 20}, abstract = {

The accelerating pace of technological advancements necessitates specialised expertise and cutting-edge instruments to maintain competitive research in life sciences. Core facilities - collaborative laboratories equipped with state-of-the-art tools and staffed by expert personnel - are vital resources that support diverse scientific endeavours. However, their adoption in lower-income communities has been comparatively stagnant due to both financial and cultural challenges. This paper explores the perils of not supporting core facilities on national research enterprises, underscoring the need for balanced investments in discovery science and crucial infrastructure support. We explore the implications from the perspectives of funders, university leaders and lab heads. We advocate for a paradigm shift to recognise these facilities as essential components of national research efforts. Core facilities are positioned not as optional but as strategic investments that can catalyse breakthroughs, particularly in environments with limited resources.

}, issn = {1365-2818}, doi = {10.1111/jmi.13277}, author = {Rahmoon, Mai A and Hobson, Chad M and Aaron, Jesse S and Balasubramanian, Harikrushnan and Chew, Teng-Leong} } @article {69321, title = {More than just {\textquoteright}added value{\textquoteright}: The perils of not establishing shared core facilities in resource-constrained communities.}, journal = {J Microsc}, year = {2024}, month = {2024 Feb 20}, abstract = {

The accelerating pace of technological advancements necessitates specialised expertise and cutting-edge instruments to maintain competitive research in life sciences. Core facilities - collaborative laboratories equipped with state-of-the-art tools and staffed by expert personnel - are vital resources that support diverse scientific endeavours. However, their adoption in lower-income communities has been comparatively stagnant due to both financial and cultural challenges. This paper explores the perils of not supporting core facilities on national research enterprises, underscoring the need for balanced investments in discovery science and crucial infrastructure support. We explore the implications from the perspectives of funders, university leaders and lab heads. We advocate for a paradigm shift to recognise these facilities as essential components of national research efforts. Core facilities are positioned not as optional but as strategic investments that can catalyse breakthroughs, particularly in environments with limited resources.

}, issn = {1365-2818}, doi = {10.1111/jmi.13277}, author = {Rahmoon, Mai A and Hobson, Chad M and Aaron, Jesse S and Balasubramanian, Harikrushnan and Chew, Teng-Leong} } @article {67963, title = {Myosin VI regulates the spatial organisation of mammalian transcription initiation.}, journal = {Nature Communications}, volume = {13}, year = {2022}, month = {2022 Mar 15}, pages = {1346}, abstract = {

During transcription, RNA Polymerase II (RNAPII) is spatially organised within the nucleus into clusters that correlate with transcription activity. While this is a hallmark of genome regulation in mammalian cells, the mechanisms concerning the assembly, organisation and stability remain unknown. Here, we have used combination of single molecule imaging and genomic approaches to explore the role of nuclear myosin VI (MVI) in the nanoscale organisation of RNAPII. We reveal that MVI in the nucleus acts as the molecular anchor that holds RNAPII in high density clusters. Perturbation of MVI leads to the disruption of RNAPII localisation, chromatin organisation and subsequently a decrease in gene expression. Overall, we uncover the fundamental role of MVI in the spatial regulation of gene expression.

}, issn = {2041-1723}, doi = {10.1038/s41467-022-28962-w}, author = {Hari-Gupta, Yukti and Fili, Natalia and Dos Santos, {\'A}lia and Cook, Alexander W and Gough, Rosemarie E and Reed, Hannah C W and Wang, Lin and Aaron, Jesse and Venit, Tomas and Wait, Eric and Grosse-Berkenbusch, Andreas and Gebhardt, J Christof M and Percipalle, Piergiorgio and Chew, Teng-Leong and Martin-Fernandez, Marisa and Toseland, Christopher P} } @article {49382, title = {Macropinosome formation by tent pole ruffling in macrophages.}, journal = {The Journal of Cell Biology}, volume = {217}, year = {2018}, month = {2018 Aug 27}, pages = {3873-85}, abstract = {

Pathogen-mediated activation of macrophages arms innate immune responses that include enhanced surface ruffling and macropinocytosis for environmental sampling and receptor internalization and signaling. Activation of macrophages with bacterial lipopolysaccharide (LPS) generates prominent dorsal ruffles, which are precursors for macropinosomes. Very rapid, high-resolution imaging of live macrophages with lattice light sheet microscopy (LLSM) reveals new features and actions of dorsal ruffles, which redefine the process of macropinosome formation and closure. We offer a new model in which ruffles are erected and supported by F-actin tent poles that cross over and twist to constrict the forming macropinosomes. This process allows for formation of large macropinosomes induced by LPS. We further describe the enrichment of active Rab13 on tent pole ruffles and show that CRISPR deletion of Rab13 results in aberrant tent pole ruffles and blocks the formation of large LPS-induced macropinosomes. Based on the exquisite temporal and spatial resolution of LLSM, we can redefine the ruffling and macropinosome processes that underpin innate immune responses.

}, issn = {1540-8140}, doi = {10.1083/jcb.201804137}, author = {Condon, Nicholas D and Heddleston, John M and Chew, Teng-Leong and Luo, Lin and McPherson, Peter S and Ioannou, Maria S and Hodgson, Louis and Stow, Jennifer L and Wall, Adam A} } @article {49051, title = {Measuring integrin conformational change on the cell surface with super-resolution microscopy.}, journal = {Cell Reports}, volume = {22}, year = {2018}, month = {2018 Feb 13}, pages = {1903-1912}, abstract = {

We use super-resolution interferometric photoactivation and localization microscopy (iPALM) and a constrained photoactivatable fluorescent protein integrin fusion to measure the displacement of the head of integrin lymphocyte function-associated 1 (LFA-1) resulting from integrin conformational change on the cell surface. We demonstrate that the distance of the LFA-1 head increases substantially between basal and ligand-engaged conformations, which can only be explained at the molecular level by integrin extension. We further demonstrate that one class of integrin antagonist maintains the bent conformation, while another antagonist class induces extension. Our molecular scale measurements on cell-surface LFA-1 are in excellent agreement with distances derived from crystallographic and electron microscopy structures of bent and extended integrins. Our distance measurements are also in excellent agreement with a previous model of LFA-1 bound to ICAM-1 derived from the orientation of LFA-1 on the cell surface measured using fluorescence polarization microscopy.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2018.01.062}, author = {Moore, Travis I and Aaron, Jesse and Chew, Teng-Leong and Springer, Timothy A} } @article {49085, title = {Model-free quantification and visualization of colocalization in fluorescence images.}, journal = {Cytometry. Part A : the journal of the International Society for Analytical Cytology}, year = {2018}, month = {2018 Mar 13}, abstract = {

The spatial association between fluorescently tagged biomolecules in situ provides valuable insight into their biological relationship. Within the limits of diffraction, such association can be measured using either Pearson{\textquoteright}s Correlation Coefficient (PCC) or Spearman{\textquoteright}s Rank Coefficient (SRC), which are designed to measure linear and monotonic correlations, respectively. However, the relationship between real biological signals is often more complex than these measures assume, rendering their results difficult to interpret. Here, we have adapted methods from the field of information theory to measure the association between two probes{\textquoteright} concentrations based on their statistical dependence. Our approach is mathematically more general than PCC or SRC, making no assumptions about the type of relationship between the probes. We show that when applied to biological images, our measures provide more intuitive results that are also more robust to outliers and the presence of multiple relationships than PCC or SRC. We also devise a display technique to highlight regions in the input images where the probes{\textquoteright} association is higher versus lower. We expect that our methods will allow biologists to more accurately and robustly quantify and visualize the association between two probes in a pair of fluorescence images. {\textcopyright} 2018 International Society for Advancement of Cytometry.

}, issn = {1552-4930}, doi = {10.1002/cyto.a.23356}, author = {Taylor, Aaron B and Ioannou, Maria S and Aaron, Jesse and Chew, Teng-Leong} } @article {48936, title = {A moving source of matrix components is essential for de novo basement membrane formation.}, journal = {Current Biology : CB}, volume = {27}, year = {2017}, month = {2017 Nov 02}, pages = {3526-34}, abstract = {

The basement membrane (BM) is a thin layer of extracellular matrix (ECM) beneath nearly all epithelial cell types that is critical for cellular and tissue function. It is composed of numerous components conserved among all bilaterians [1]; however, it is unknown how all of these components are generated and subsequently constructed to form a fully mature BM in the living animal. Although BM formation is thought to simply involve a process of self-assembly [2], this concept suffers from a number of logistical issues when considering its construction in\ vivo. First, incorporation of BM components appears to be hierarchical [3-5], yet it is unclear whether their production during embryogenesis must also be regulated in a temporal fashion. Second, many BM proteins are produced not only by the cells residing on the BM but also by surrounding cell types [6-9], and it is unclear how large, possibly insoluble protein complexes [10] are delivered into the matrix. Here we exploit our ability to live image and genetically dissect de novo BM formation during Drosophila development. This reveals that there is a temporal hierarchy of BM protein production that is essential for proper component incorporation. Furthermore, we show that BM components require secretion by migrating macrophages (hemocytes) during their developmental dispersal, which is critical for embryogenesis. Indeed, hemocyte migration is essential to deliver a subset of ECM components evenly throughout the embryo. This reveals that de novo BM construction requires a combination of both production and distribution logistics allowing for the timely delivery of core components.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2017.10.001}, author = {Matsubayashi, Yutaka and Louani, Adam and Dragu, Anca and S{\'a}nchez-S{\'a}nchez, Besaiz J and Serna-Morales, Eduardo and Yolland, Lawrence and Gyoergy, Attila and Vizcay, Gema and Fleck, Roland A and Heddleston, John M and Chew, Teng-Leong and Siekhaus, Daria E and Stramer, Brian M} }