Rho1 Directs Formin-Mediated Actin Ring Assembly during Budding Yeast Cytokinesis

AbstractIn eukaryotic cells, dynamic rearrangement of the actin cytoskeleton is critical for cell division. In the yeast Saccharomyces cerevisiae, three main structures constitute the actin cytoskeleton: cortical actin patches, cytoplasmic actin cables, and the actin-based cytokinetic ring [1–4]. The conserved Arp2/3 complex and a WASP-family protein mediate actin patch formation, whereas the yeast formins (Bni1 and Bnr1) promote assembly of actin cables [5–9]. However, the mechanism of actin ring formation is currently unclear. Here, we show that actin filaments are required for cytokinesis in S. cerevisiae, and that the actin ring is a highly dynamic structure that undergoes constant turnover. Assembly of the actin ring requires the formin-like proteins and profilin, but is not Arp2/3-mediated. Furthermore, the formin-dependent actin ring assembly pathway is regulated by the Rho-type GTPase Rho1 but not Cdc42. Finally, we show that the formins are not required for localization of Cyk1/Iqg1, an IQGAP-like protein previously shown to be required for actin ring formation, suggesting that formin-like proteins and Cyk1 act synergistically but independently in assembly of the actin ring

“A team around the child” professionals’ experiences of unmet needs, access and expectations in children’s palliative care services, a phenomenological study in the UK

This study aims to understand the experiences of professionals involved in caring for families of children with life-limiting illnesses to ascertain unmet needs, access issues and expectations of services. A phenomenological approach comprising semi-structured interviews with professionals from various services was used. Interviews that happened between July and November 2018 were audio-recorded, transcribed verbatim and thematically analysed based on descriptive phenomenology. In total, 29 interviews were conducted. Findings showed how essential collaborative working was for becoming a team around the child. Barriers to this include complex working relationships, overprotectiveness of families, roles and responsibilities and use of independent care records. The pressures of waiting times and the impact of staffing shortages affected the experiences of providing care. The reality of meeting families' expectations was shaped by family networking, online research and previous services resistance, which was influential in more challenging interactions. Expectations of care were also impacted by misunderstandings and anxieties surrounding access to services. Overall, professionals were concerned about families being used as a bottomless caring resource and stressed the need for short breaks to alleviate parents. Interventions that educate families and professionals on these services and how they can benefit the child and family would be well received

ChemBank: a small-molecule screening and cheminformatics resource database

ChemBank (http://chembank.broad.harvard.edu/) is a public, web-based informatics environment developed through a collaboration between the Chemical Biology Program and Platform at the Broad Institute of Harvard and MIT. This knowledge environment includes freely available data derived from small molecules and small-molecule screens and resources for studying these data. ChemBank is unique among small-molecule databases in its dedication to the storage of raw screening data, its rigorous definition of screening experiments in terms of statistical hypothesis testing, and its metadata-based organization of screening experiments into projects involving collections of related assays. ChemBank stores an increasingly varied set of measurements derived from cells and other biological assay systems treated with small molecules. Analysis tools are available and are continuously being developed that allow the relationships between small molecules, cell measurements, and cell states to be studied. Currently, ChemBank stores information on hundreds of thousands of small molecules and hundreds of biomedically relevant assays that have been performed at the Broad Institute by collaborators from the worldwide research community. The goal of ChemBank is to provide life scientists unfettered access to biomedically relevant data and tools heretofore available primarily in the private sector

Cyclin D1-CDK4 Controls Glucose Metabolism Independently of Cell Cycle Progression

Insulin constitutes a major evolutionarily conserved hormonal axis for maintaining glucose homeostasis1-3; dysregulation of this axis causes diabetes2,4. PGC-1α links insulin signaling to the expression of glucose and lipid metabolic genes5-7. GCN5 acetylates PGC-1α and suppresses its transcriptional activity, whereas SIRT1 deacetylates and activates PGC-1α8,9. Although insulin is a mitogenic signal in proliferative cells10,11, whether components of the cell cycle machinery contribute to insulin’s metabolic action is poorly understood. Herein, we report that insulin activates cyclin D1-CDK4, which, in turn, increases GCN5 acetyltransferase activity and suppresses hepatic glucose production independently of cell cycle progression. Through a cell-based high throughput chemical screen, we identified a CDK4 inhibitor that potently decreases PGC-1α acetylation. Insulin/GSK3β signaling induces cyclin D1 protein stability via sequestering cyclin D1 in the nucleus. In parallel, dietary amino acids increase hepatic cyclin D1 mRNA transcripts. Activated cyclin D1-CDK4 kinase phosphorylates and activates GCN5, which then acetylates and inhibits PGC-1α activity on gluconeogenic genes. Loss of hepatic cyclin D1 results in increased gluconeogenesis and hyperglycemia. In diabetic models, cyclin D1-CDK4 is chronically elevated and refractory to fasting/feeding transitions; nevertheless further activation of this kinase normalizes glycemia. Our findings show that insulin uses components of the cell cycle machinery in post-mitotic cells to control glucose homeostasis independently of cell division

Application of kernel functions for accurate similarity search in large chemical databases

Background Similaritysearch in chemical structure databases is an important problem with many applications in chemical genomics, drug design, and efficient chemical probe screening among others. It is widely believed that structure based methods provide an efficient way to do the query. Recently various graph kernel functions have been designed to capture the intrinsic similarity of graphs. Though successful in constructing accurate predictive and classification models, graph kernel functions can not be applied to large chemical compound database due to the high computational complexity and the difficulties in indexing similarity search for large databases. Results To bridge graph kernel function and similarity search in chemical databases, we applied a novel kernel-based similarity measurement, developed in our team, to measure similarity of graph represented chemicals. In our method, we utilize a hash table to support new graph kernel function definition, efficient storage and fast search. We have applied our method, named G-hash, to large chemical databases. Our results show that the G-hash method achieves state-of-the-art performance for k-nearest neighbor (k-NN) classification. Moreover, the similarity measurement and the index structure is scalable to large chemical databases with smaller indexing size, and faster query processing time as compared to state-of-the-art indexing methods such as Daylight fingerprints, C-tree and GraphGrep. Conclusions Efficient similarity query processing method for large chemical databases is challenging since we need to balance running time efficiency and similarity search accuracy. Our previous similarity search method, G-hash, provides a new way to perform similarity search in chemical databases. Experimental study validates the utility of G-hash in chemical databases

Zds1/Zds2-PP2ACdc55 complex specifies signaling output from Rho1 GTPase

Acknowledgments We thank David Pellman, John Pringle, Daniel Lew, Masaki Mizunuma, Kenji Irie, and the Yeast Genome Resource Center for yeast strains and plasmids and members of Yoshida Laboratory and Keiko Kono for their support. Multicopy suppressor screening for gef∆ was initiated in the Pellman Laboratory with the help of Didem Ilter. This research was supported by Sprout grant from Brandeis University (E.M. Jonasson and S. Yoshida), an American-Italian Cancer Foundation Postdoctoral fellowship (V. Rossio), and a Massachusetts Life Sciences Center grant (S. Yoshida).Peer reviewedPublisher PD

A Chemical Screen Probing the Relationship between Mitochondrial Content and Cell Size

The cellular content of mitochondria changes dynamically during development and in response to external stimuli, but the underlying mechanisms remain obscure. To systematically identify molecular probes and pathways that control mitochondrial abundance, we developed a high-throughput imaging assay that tracks both the per cell mitochondrial content and the cell size in confluent human umbilical vein endothelial cells. We screened 28,786 small molecules and observed that hundreds of small molecules are capable of increasing or decreasing the cellular content of mitochondria in a manner proportionate to cell size, revealing stereotyped control of these parameters. However, only a handful of compounds dissociate this relationship. We focus on one such compound, BRD6897, and demonstrate through secondary assays that it increases the cellular content of mitochondria as evidenced by fluorescence microscopy, mitochondrial protein content, and respiration, even after rigorous correction for cell size, cell volume, or total protein content. BRD6897 increases uncoupled respiration 1.6-fold in two different, non-dividing cell types. Based on electron microscopy, BRD6897 does not alter the percent of cytoplasmic area occupied by mitochondria, but instead, induces a striking increase in the electron density of existing mitochondria. The mechanism is independent of known transcriptional programs and is likely to be related to a blockade in the turnover of mitochondrial proteins. At present the molecular target of BRD6897 remains to be elucidated, but if identified, could reveal an important additional mechanism that governs mitochondrial biogenesis and turnover