Errors during meiotic resumption in oocytes can result in chromosome missegregation and infertility. Several cell cycle kinases have been linked with roles in coordinating events during meiotic resumption, including polo-like kinases (PLK). Mammals express four kinase-proficient PLKs, (PLK1-4). Previous studies assessing the role of PLK1 have relied on RNA knockdown and kinase inhibition approaches, as Plk1 null mutations are embryonically lethal. To further assess the roles of PLK1 during meiotic resumption, we developed a Plk1 conditional knockout (cKO) mouse to specifically mutate Plk1 in oocytes. Despite normal oocyte numbers and follicle maturation, Plk1 cKO mice were infertile. From analysis of meiotic resumption, Plk1 cKO oocytes underwent nuclear envelope breakdown with the same timing as control oocytes. However, Plk1 cKO oocytes failed to form compact bivalent chromosomes, and localization of cohesin and condensin were defective. Furthermore, Plk1 cKO oocytes either failed to organize α-tubulin or developed an abnormally small bipolar spindle. These abnormalities were attributed to aberrant release of microtubule organizing center (MTOC) linker protein, C-NAP1, and the failure to recruit MTOC components and liquid-like spindle domain factors (LISD). Ultimately, these defects result in meiosis I arrest prior to homologous chromosome segregation.Forces generated by heart muscle contraction must be balanced by adhesion to the extracellular matrix (ECM) and to other cells for proper heart function. Decades of data have suggested that cell-ECM adhesions are important for sarcomere assembly. However, the relationship between cell-ECM adhesions and sarcomeres assembling de novo remains untested. Sarcomeres arise from muscle stress fibers that are translocating on the top (dorsal) surface of cultured cardiomyocytes. Using an array of tools to modulate cell-ECM adhesion, we established a strong positive correlation between the extent of cell-ECM adhesion and sarcomere assembly. On the other hand, we found a strong negative correlation between the extent of cell-ECM adhesion and the rate of MSF translocation; a phenomenon also observed in non-muscle cells. We further find a conserved network architecture that also exists in non-muscle cells. Taken together, our results show that cell-ECM adhesions mediate coupling between the substrate and muscle stress fibers, allowing their maturation into sarcomere-containing myofibrils. [Media see text] [Media see text].The Coronavirus Disease 2019 (COVID-19) pandemic initially presented in the United States in the greater Seattle area, and has rapidly progressed across the nation in the past 2 months, with the United States having the highest number of cases in the world. Radiology departments play a critical role in policy and guideline development both for the department and for the institutions, specifically in planning diagnostic screening, triage, and management of patients. In addition, radiology workflows, volumes and access must be optimized in preparation for the expected COVID-19 patient surges. This article discusses the processes that have been implemented at the University of Washington in managing the COVID-19 pandemic as well in preparing for patient surges, which may provide important guidance for other radiology departments who are in the early stages of preparation and management.Eukaryotic cells are compartmentalized into organelles by intracellular membranes. While the organelles are distinct, many of them make intimate contact with one another. https://www.selleckchem.com/products/TW-37.html These contacts were first observed in the 1950's, but only recently have the functions of these contact sites begun to be understood. In yeast, the ER makes extensive inter-membrane contacts with the plasma membrane (PM), covering ∼40% of the PM. Many functions of ER-PM contacts have been proposed, including non-vesicular lipid trafficking, ion transfer, and as signaling hubs. Surprisingly, cells that lack ER-PM contacts grow well, indicating that alternative pathways may be compensating for the loss of ER-PM contact. In order to better understand the function of ER-PM contact sites we used saturating transposon mutagenesis to identify synthetic lethal mutants in a yeast strain lacking ER-PM contact sites. The strongest hits were components of the ESCRT complexes. The synthetic lethal mutants have low levels of some lipid species but accumulate free fatty acids and lipid droplets. We found that only ESCRT-III components are synthetic lethal, indicating that Vps4 and other ESCRT complexes do not function in this pathway. These data suggest that ESCRT-III proteins and ER-PM contact sites act in independent pathways to maintain lipid homeostasis.The multi-component cytoplasmic dynein transports cellular cargoes with the help of another multi-component complex dynactin, but we do not know enough about factors that may affect the assembly and functions of these proteins. From a genetic screen for mutations affecting early-endosome distribution in Aspergillus nidulans, we identified the prp40AL438* mutation in Prp40A, a homolog of Prp40, an essential RNA-splicing factor in the budding yeast. Prp40A is not essential for splicing, although it associates with the nuclear splicing machinery. The prp40AL438* mutant is much healthier than the ∆prp40A mutant, but both mutants exhibit similar defects in dynein-mediated early endosome transport and nuclear distribution. In the prp40AL438* mutant, the frequency but not the speed of dynein-mediated early endosome transport is decreased, which correlates with a decrease in the microtubule plus-end accumulations of dynein and dynactin. Within the dynactin complex, the actin-related protein Arp1 forms a mini-filament. In a pulldown assay, the amount of Arp1 pulled-down with its pointed-end protein Arp11 is lowered in the prp40AL438* mutant. In addition, we found from published interactome data that a mammalian Prp40 homolog PRPF40A interacts with Arp1. Thus, Prp40 homologs may regulate the assembly or function of dynein-dynactin and their mechanisms deserve to be further studied.Nuclei are often under external stress - be it during migration through tight constrictions or compressive pressure by the actin cap - and the mechanical properties of nuclei govern their subsequent deformations. Both altered mechanical properties of nuclei and abnormal nuclear morphologies are hallmarks of a variety of disease states. Little work, however, has been done to link specific changes in nuclear shape to external forces. Here, we utilize a combined atomic force microscope and light sheet microscope (AFM-LS) to show SKOV3 nuclei exhibit a two-regime force response that correlates with changes in nuclear volume and surface area, allowing us to develop an empirical model of nuclear deformation. Our technique further decouples the roles of chromatin and lamin A/C in compression, showing they separately resist changes in nuclear volume and surface area respectively; this insight was not previously accessible by Hertzian analysis. A two-material finite element model supports our conclusions. We also observed that chromatin decompaction leads to lower nuclear curvature under compression, which is important for maintaining nuclear compartmentalization and function.