Focal adhesions (FAs) are dynamic protein nanostructures that form mechanical links between cytoskeletal actin fibers and the extracellular matrix. Here, we demonstrate that interferometric scattering (iSCAT) microscopy, a high-speed and time-unlimited imaging technique, can uncover the real-time dynamics of nanoscopic nascent adhesions (NAs). The high sensitivity and stability of the iSCAT signal enabled us to trace the whole life span of each NA spontaneously nucleated under a lamellipodium. Such high-throughput and long-term image data provide a unique opportunity for statistical analysis of adhesion dynamics. Moreover, we directly revealed that FAs play critical roles in both the extrusion of filopodia as nucleation sites on the leading edge and the one-dimensional transport of cargos along cytoskeletal fibers as fiber docking sites. These experimental results show that iSCAT is a sensitive tool for tracking real-time dynamics of nanoscopic objects involved in endogenous and exogenous biological processes in living cells.
53BP1 contributes to activation of the G2/M checkpoint downstream of ATM and MDC1 in response to ionizing radiation and promotes nonhomologous end-joining (NHEJ) in mammalian cells. In order to determine whether the cellular activities of 53BP1 are conserved in the model organism C. elegans, we analyzed the function of its homolog, HSR-9 in response to DNA damage. Deletion or Mos1-insertion in hsr-9 did not affect the sensitivity of worms to double strand DNA breaks (DSBs), as reflected in embryonic survival and larval development. Nevertheless, the hsr-9 mutations, as well as a lig-4 deletion, reversed the hypersensitivity of rad-54-deficient worms to DSBs. In addition, oocyte chromosomal aberrations, which were increased by rad-54 knockdown in response to DSBs, were also reduced by the hsr-9 mutations. The hsr-9 mutations did not prevent the cell cycle arrest induced by DSBs in mitotically proliferating germ cells. However, they attenuated apoptosis induced by DSBs, but not when CEP-1 (a p53 ortholog) was absent, suggesting that HSR-9 functions in the same pathway as CEP-1. We concluded that the 53BP1 homolog in C. elegans is not directly involved in cell cycle arrest in response to DSBs, but that it promotes apoptosis and also a form of NHEJ that occurs only when rad-54 is deficient.
Objective We investigated the proportions of and reclassified BRCA1/2 variants of unknown significance (VUS) in Korean patients with epithelial ovarian, tubal, and primary peritoneal cancers. Methods Data from 805 patients who underwent genetic testing for BRCA1/2 from January 1, 2006 to August 31, 2018 were included. The VUS in BRCA1/2 were reclassified using the 2015 American College of Medical Genetics and Genomics and the Association for Molecular Pathology standards and guidelines. Results A BRCA1 pathogenic variant was found in 17.0% (137/805) of the patients, and BRCA1 VUS were found in 15.9% (128/805) of the patients. Further, 8.7% (69/805) of the patients possessed a BRCA2 pathogenic variant and 18.4% (148/805) of the patients possessed BRCA2 VUS. Fifty-three specific BRCA1 VUS were found and 20 were further reclassified as benign (n=11), likely benign (n=5), likely pathogenic (n=3), and pathogenic (n=1). The remaining 33 remained classified as VUS. For BRCA2 , 55 specific VUS were detected; among these, 14 were reclassified as benign or likely benign, and 2 were reclassified as likely pathogenic. Among the 805 patients, 195 were found to have only VUS and no pathogenic variants (PV), and 41.5% (81/195) were reclassified as benign or likely benign, and 10.3% (20/195) as pathogenic or likely pathogenic variants. Conclusions Approximately 33.3% (36/108) of the specific BRCA1/2 variants analyzed in this study that were initially classified as VUS over a 13-year period were reclassified. Among these, 5.6% (6/108) were reclassified as pathogenic or likely pathogenic variants.
Werner syndrome protein (WRN) is unusual among RecQ family DNA helicases in having an additional exonuclease activity. WRN is involved in the repair of double-strand DNA breaks via the homologous recombination and nonhomologous end joining pathways, and also in the base excision repair pathway. In addition, the protein promotes the recovery of stalled replication forks. The helicase activity is thought to unwind DNA duplexes, thereby moving replication forks or Holliday junctions. The targets of the exonuclease could be the nascent DNA strands at a replication fork or the ends of double-strand DNA breaks. However, it is not clear which enzyme activities are essential for repairing different types of DNA damage. Model organisms such as mice, flies, and worms deficient in WRN homologs have been investigated to understand the physiological results of defects in WRN activity. Premature aging, the most remarkable characteristic of Werner syndrome, is also seen in the mutant mice and worms, and hypersensitivity to DNA damage has been observed in WRN mutants of all three model organisms, pointing to conservation of the functions of WRN. In the nematode Caenorhabditis elegans, the WRN homolog contains a helicase domain but no exonuclease domain, so that this animal is very useful for studying the in vivo functions of the helicase without interference from the activity of the exonuclease. Here, we review the current status of investigations of C. elegans WRN-1 and discuss its functional differences from the mammalian homologs.
PurposeThis study investigated the effectiveness of meaning-centered psychotherapy (MCP), which is known to be a helpful psychotherapeutic intervention in distressing conditions, for patients with pancreatobiliary cancer.Materials and MethodsWe recruited 37 patients with pancreatobiliary cancer from three university general hospitals and assessed their psychological characteristics. Patients who reported clinically significant emotional distress were recommended to undergo MCP. Patients who consented to MCP were provided four sessions of the therapy. Patient psychological characteristics were assessed again 2 months after MCP. For statistical comparison, outcome variables included anxiety, depression, mental adjustment to cancer, and quality of life (QoL), as well as the degree of stress and physical symptoms.ResultsSixteen patients completed the MCP and the final assessment 2 months later. In the initial assessment, the patients receiving MCP showed higher levels of anxiety and depression than those not receiving MCP, and QoL was also lower in terms of role function, emotional function, social function, and global QoL. At the 2-month follow-up, the MCP group showed a significant improvement in anxiety (p=0.007), depression (p=0.010), and anxious preoccupation (p<0.001). In addition, QoL significantly improved in the MCP group, while there was no significant change in the non-MCP group.ConclusionIn this study, MCP showed potential therapeutic benefits against emotional distress in patients with pancreatobiliary cancer, improving their QoL.
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