An Arp2/3 Nucleated F-Actin Shell Fragments Nuclear Membranes at Nuclear Envelope Breakdown in Starfish Oocytes

Animal cells disassemble and reassemble their nuclear envelopes (NEs) upon each division. Nuclear envelope breakdown (NEBD) serves as a major regulatory mechanism by which mixing of cytoplasmic and nuclear compartments drives the complete reorganization of cellular architecture, committing the cell for division. Breakdown is initiated by phosphorylation-driven partial disassembly of the nuclear pore complexes (NPCs), increasing their permeability but leaving the overall NE structure intact. Subsequently, the NE is rapidly broken into membrane fragments, defining the transition from prophase to prometaphase and resulting in complete mixing of cyto- and nucleoplasm. However, the mechanism underlying this rapid NE fragmentation remains largely unknown. Here, we show that NE fragmentation during NEBD in starfish oocytes is driven by an Arp2/3 complex-nucleated F-actin “shell” that transiently polymerizes on the inner surface of the NE. Blocking the formation of this F-actin shell prevents membrane fragmentation and delays entry of large cytoplasmic molecules into the nucleus. We observe spike-like protrusions extending from the F-actin shell that appear to “pierce” the NE during the fragmentation process. Finally, we show that NE fragmentation is essential for successful reproduction, because blocking this process in meiosis leads to formation of aneuploid eggs.MIT Faculty Start-up FundNational Science Foundation (U.S.). Graduate Research Fellowshi

Plk1 overexpression induces chromosomal instability and suppresses tumor development

Polo-like kinase 1 (Plk1) is overexpressed in a wide spectrum of human tumors, being frequently considered as an oncogene and an attractive cancer target. However, its contribution to tumor development is unclear. Using a new inducible knock-in mouse model we report here that Plk1 overexpression results in abnormal chromosome segregation and cytokinesis, generating polyploid cells with reduced proliferative potential. Mechanistically, these cytokinesis defects correlate with defective loading of Cep55 and ESCRT complexes to the abscission bridge, in a Plk1 kinase-dependent manner. In vivo, Plk1 overexpression prevents the development of Kras-induced and Her2-induced mammary gland tumors, in the presence of increased rates of chromosome instability. In patients, Plk1 overexpression correlates with improved survival in specific breast cancer subtypes. Therefore, despite the therapeutic benefits of inhibiting Plk1 due to its essential role in tumor cell cycles, Plk1 overexpression has tumor-suppressive properties by perturbing mitotic progression and cytokinesis.We are indebted to Stephen Taylor for the Sgo1 antibody. We thank Simone Kraut, Jessica Steiner, and the DKFZ light microscopy unit for excellent technical assistance. The results published here are in part based on data generated by TCGA pilot project (https://cancergenome.nih.gov/established by the NCI and the National Human Gen- ome Research Institute. The data were retrieved through dbGaP authorization (accession no. phs000178.v9.p8). S.V.V. was supported by the Marie Curie Network Ploidynet, funded by the European Union Seventh Framework Programme (FP7/2007–2013) under Grant Agreement #316964. L.S. is supported by a postdoctoral fellowship from Funda- cion Ramon Areces. Work in the R.S. laboratory was supported by an ERC starting grant (#281614), Marie Curie PCIG09-GA-2011 –293745 and the Howard Hughes Medical Institute. G.d.C. is funded by AECC Scientific Foundation (LABAE16017DECA). Work in the M.M. laboratory was supported by grants from the MINECO (SAF2015 –69920-R cofunded by ERDF-EU), Worldwide Cancer Research (WCR no. 150278), and Comunidad de Madrid (iLUNG-CM; B2017/BMD3884). The CNIO is a Severo Ochoa Center of Excellence (MINECO award SEV-2015-0510).S

Gain-of-function screen for genes that affect Drosophila muscle pattern formation.

This article reports the production of an EP-element insertion library with more than 3,700 unique target sites within the Drosophila melanogaster genome and its use to systematically identify genes that affect embryonic muscle pattern formation. We designed a UAS/GAL4 system to drive GAL4-responsive expression of the EP-targeted genes in developing apodeme cells to which migrating myotubes finally attach and in an intrasegmental pattern of cells that serve myotubes as a migration substrate on their way towards the apodemes. The results suggest that misexpression of more than 1.5% of the Drosophila genes can interfere with proper myotube guidance and/or muscle attachment. In addition to factors already known to participate in these processes, we identified a number of enzymes that participate in the synthesis or modification of protein carbohydrate side chains and in Ubiquitin modifications and/or the Ubiquitin-dependent degradation of proteins, suggesting that these processes are relevant for muscle pattern formation

Acquisition of chromosome instability is a mechanism to evade oncogene addiction

Abstract Chromosome instability (CIN) has been associated with therapeutic resistance in many cancers. However, whether tumours become genomically unstable as an evolutionary mechanism to overcome the bottleneck exerted by therapy is not clear. Using a CIN model of Kras‐driven breast cancer, we demonstrate that aneuploid tumours acquire genetic modifications that facilitate the development of resistance to targeted therapy faster than euploid tumours. We further show that the few initially chromosomally stable cancers that manage to persist during treatment do so concomitantly with the acquisition of CIN. Whole‐genome sequencing analysis revealed that the most predominant genetic alteration in resistant tumours, originated from either euploid or aneuploid primary tumours, was an amplification on chromosome 6 containing the cMet oncogene. We further show that these tumours are dependent on cMet since its pharmacological inhibition leads to reduced growth and increased cell death. Our results highlight that irrespective of the initial CIN levels, cancer genomes are dynamic and the acquisition of a certain level of CIN, either induced or spontaneous, is a mechanism to circumvent oncogene addiction

Bibliografi över arbeten som producerats av uppsalaslavister under året 1977

PLK1 is a mitotic regulator overexpressed in cancer; however, whether this overexpression causally contributes to tumor development is unclear. Here the authors produce an inducible mouse model to overexpress PLK1 and show that actually this can act as a tumor suppressor by perturbing mitotic progression and cytokinesis

Acute expression of human APOBEC3B in mice results in RNA editing and lethality

Abstract Background RNA editing has been described as promoting genetic heterogeneity, leading to the development of multiple disorders, including cancer. The cytosine deaminase APOBEC3B is implicated in tumor evolution through DNA mutation, but whether it also functions as an RNA editing enzyme has not been studied. Results Here, we engineer a novel doxycycline-inducible mouse model of human APOBEC3B-overexpression to understand the impact of this enzyme in tissue homeostasis and address a potential role in C-to-U RNA editing. Elevated and sustained levels of APOBEC3B lead to rapid alteration of cellular fitness, major organ dysfunction, and ultimately lethality in mice. Importantly, RNA-sequencing of mouse tissues expressing high levels of APOBEC3B identifies frequent UCC-to-UUC RNA editing events that are not evident in the corresponding genomic DNA. Conclusions This work identifies, for the first time, a new deaminase-dependent function for APOBEC3B in RNA editing and presents a preclinical tool to help understand the emerging role of APOBEC3B as a driver of carcinogenesis

Additional file 1 of Acute expression of human APOBEC3B in mice results in RNA editing and lethality

Additional file 1. Supplementary figures

Expression Patterns of Genes that Cause a Gain-of-Function Muscle Phenotype

<p>Lateral views of embryos at stage 11 (M), stage 13 (A, C, E, G, I, K, and N), and stage 16 (B, D, F, H, J, and L) that were stained with transcript-specific anti-sense RNA probes or with anti-Toll antibodies (A and B). Note the expression of Toll (A and B) in segment border cells, <i>sdc</i> (LD08230) (C and D) in trachea, segment border cells, and the differentiated apodemes, CG3563 (LD15689) (E and F) in the apodeme precusor cells at the segment border, CG13913 (RE53394) (G and H) and CG5008/<i>gnbp3</i> (SD21560) (I and J) in a subset of apodeme precursors and cells of the epidermis, CG14713/14714 transcripts (AT17253) (K and L) in the dorsal and ventral epidermis around the segment border, and <i>pxb</i> (SD26190) (M and N) in intrasegmental epidermal stripes.</p

Muscle Pattern Defects in <i>esg</i> and <i>sdc</i> Mutants

<p>Muscle pattern of three segments of <i>oreR</i> (A, D, and G), <i>esg ^L2</i> (B and H), and <i>sdc ²³</i> mutant embryos (C, E, F, and I) after staining with anti-MHC antibodies or using a <i>delilah</i> transcript-specific anti-sense RNA probe (G–H). Lateral (A–C and G–I) and ventral views (D–F) of embryos at stage 14 (E) and stage 16 (A–D and F–I). <i>esg</i> mutant embryos show variable muscle pattern defects with muscles absent ([B], arrowheads). In <i>sdc</i> mutant embryos few muscles cross the ventral midline in a position dorsal to the central nervous system ([E], arrowheads), and they show disruptions of the pattern in the ventral region ([C], arrowheads). The typical “finger-type pattern” of the ventral muscles of wild-type embryos (D) is unordered in <i>sdc ²³</i> mutant embryos, with ventral muscles aligning in parallel with the anterio-posterior axis, ignoring the segment border attachment ([F], arrowheads). Also shown is the pattern of epidermal muscle attachment sites (<i>delilah</i> marker gene expression) in wild-type (G), <i>esg</i> (H), and <i>sdc</i> (I) mutant embryos. Note that the pattern is unchanged in the mutants.</p