Aneuploidy facilitates dysplastic and tumorigenic phenotypes in the Drosophila gut

Aneuploidy has been strongly linked to cancer development, and published evidence has suggested that aneuploidy can have an oncogenic or a tumor suppressor role depending on the tissue context. Using the Drosophila midgut as a model, we have recently described that adult intestinal stem cells (ISCs), do not activate programmed cell death upon aneuploidy induction, leading to an increase in ISC proliferation rate, and tissue dysplasia. How aneuploidy impacts ISCs in intestinal tumorigenic models remains to be investigated, and it represents a very important biological question to address since data from multiple in vivo models suggests that the cellular impact of aneuploidy is highly dependent on the cellular and tissue context. Using manipulation of different genetic pathways such as EGFR, JAK-STAT and Notch that cause dysplastic phenotypes in the Drosophila gut, we found that concomitant aneuploidy induction by impairment of the Spindle Assembly Checkpoint (SAC) consistently leads to a more severe progression of intestinal dysplasia or tumorigenesis. This is characterized by an accumulation of progenitor cells, high tissue cell density and higher stem cell proliferation rates, revealing an additive or synergistic effect depending on the misregulated pathway in which aneuploidy was induced. Thus, our data suggests that in the Drosophila gut, both dysplasia and tumorigenic phenotypes can be fueled by inducing genomic instability of resident stem cells.This work was funded by National Funds through FCT – Fundação para a Ciência e a Tecnologia, I.P., under the project UIDB/04293/2020

Aurora A triggers Lgl cortical release during symmetric division to control planar spindle orientation

Mitotic spindle orientation is essential to control cell-fate specification and epithelial architecture. The tumor suppressor Lgl localizes to the basolateral cortex of epithelial cells, where it acts together with Dlg and Scrib to organize apicobasal polarity. Dlg and Scrib also control planar spindle orientation, but how the organization of polarity complexes is adjusted to control symmetric division is largely unknown. Here, we show that the Dlg complex is remodeled during Drosophila follicular epithelium cell division, when Lgl is released to the cytoplasm. Lgl redistribution during epithelial mitosis is reminiscent of asymmetric cell division, where it is proposed that Aurora A promotes aPKC activation to control the localization of Lgl and cell-fate determinants. We show that Aurora A controls Lgl localization directly, triggering its cortical release at early prophase in both epithelial and S2 cells. This relies on double phosphorylation within the putative aPKC phosphorylation site, which is required and sufficient for Lgl cortical release during mitosis and can be achieved by a combination of aPKC and Aurora A activities. Cortical retention of Lgl disrupts planar spindle orientation, but only when Lgl mutants that can bind Dlg are expressed. Hence, our work reveals that Lgl mitotic cortical release is not specifically linked to the asymmetric segregation of fate determinants, and we propose that Aurora A activation breaks the Dlg/Lgl interaction to allow planar spindle orientation during symmetric division via the Pins (LGN)/Dlg pathway.We thank J. Knoblich, D. St Johnston, D. Bilder, D. Glover, S. Brogna, R. Martinho, H. Maiato, D. Bergstralh, and the Bloomington Stock Center for fly stocks and reagents. This work was funded by FEDER funds through the Operational Competitiveness Programme COMPETE and by National Funds through FCT (Fundação para a Ciência e a Tecnologia) under the project FCOMP-01-0124-FEDER-019738 (PTDC/BIA-BCM/120132/2010), which also supported fellowships to C.C. and S.M. E.M. was funded by a Marie Curie-IEF and currently holds a FCT Investigator position

PP1-Mediated Dephosphorylation of Lgl Controls Apical-basal Polarity

Apical-basal polarity is a common trait that underlies epithelial function. Although the asymmetric distribution of cortical polarity proteins works in a functioning equilibrium, it also retains plasticity to accommodate cell division, during which the basolateral determinant Lgl is released from the cortex. Here, we investigated how Lgl restores its cortical localization to maintain the integrity of dividing epithelia. We show that cytoplasmic Lgl is reloaded to the cortex at mitotic exit in Drosophila epithelia. Lgl cortical localization depends on protein phosphatase 1, which dephosphorylates Lgl on the serines phosphorylated by aPKC and Aurora A kinases through a mechanism that relies on the regulatory subunit Sds22 and a PP1-interacting RVxF motif of Lgl. This mechanism maintains epithelial polarity and is of particular importance at mitotic exit to couple Lgl cortical reloading with the polarization of the apical domain. Hence, PP1-mediated dephosphorylation of Lgl preserves the apicalbasal organization of proliferative epithelia.We thank Daniel St Johnston, François Schweisguth, Guilles Hickson, Jürgen Knoblich, Torcato Martins, Yang Hong, and the Bloomington Drosophila Stock Center for providing plasmids and fly stocks. This work was funded by national funds through Fundação para a Ciência e a Tecnologia (FCT) under project PTDC/BEX-BCM/0432/2014 . This work has also received funding from the project Norte-01-0145-FEDER-000029 , supported by Norte Portugal Regional Operational Program (NORTE 2020). E.M. holds an FCT Investigator position. S.M. and M.G. are supported by FCT PhD grants. M.O. is supported by a fellowship from FCT and the GABBA PhD program from the University of Porto

Genome-wide RNAi screen for synthetic lethal interactions with the C. elegans kinesin-5 homolog BMK-1

Kinesins are a superfamily of microtubule-based molecular motors that perform various transport needs and have essential roles in cell division. Among these, the kinesin-5 family has been shown to play a major role in the formation and maintenance of the bipolar mitotic spindle. Moreover, recent work suggests that kinesin-5 motors may have additional roles. In contrast to most model organisms, the sole kinesin-5 gene in Caenorhabditis elegans, bmk-1, is not required for successful mitosis and animals lacking bmk-1 are viable and fertile. To gain insight into factors that may act redundantly with BMK-1 in spindle assembly and to identify possible additional cellular pathways involving BMK-1, we performed a synthetic lethal screen using the bmk-1 deletion allele ok391. We successfully knocked down 82% of the C. elegans genome using RNAi and assayed viability in bmk-1(ok391) and wild type strains using an automated high-throughput approach based on fluorescence microscopy. The dataset includes a final list of 37 synthetic lethal interactions whose further study is likely to provide insight into kinesin-5 function.We thank the members of the Medema, Kops, Lens, Boxem, The, van den Heuvel, Carvalho and Gassmann laboratories for helpful discussion. To Belen Fernandez-Garcia for helping on hit picking from the genome-wide library. To Oliver Pelz for help with web cellHTS2 application during data analysis. A.F. Maia is a FCT-Fundacao para a Ciencia e a Tecnologia postdoctoral fellow (SFRH/BPD/71364/2010). The R.H. Medema laboratory was supported by the Netherlands Organization for Scientific Research (ZonMw 918.46.616) and the Netherlands Genomics Initiative. S. van den Heuvel received funding from the Netherlands Organisation for Scientific Research (NWO), Chemical Sciences (CW ECHO project 711.011.010). Work on C.E. Sunkel laboratory was funded by FEDER funds through the Operational Competitiveness Programme-COMPETE and by National Funds through FCT-Fundacao para a Ciencia e a Tecnologia under the project FCOMP-01-0124-FEDER-019740 (PTDC/BIA-BCM/120366/2010). R. Gassmann laboratory is supported by an ERC Starting Grant (338410), an EMBO Installation Grant, and funding from the Fundacao para a Ciencia e a Tecnologia (IF/01015/2013/CP1157/CT0006). R.H. Medema had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis

PLK1 facilitates chromosome biorientation by suppressing centromere disintegration driven by BLM-mediated unwinding and spindle pulling

Centromeres provide a pivotal function for faithful chromosome segregation. They serve as a foundation for the assembly of the kinetochore complex and spindle connection, which is essential for chromosome biorientation. Cells lacking Polo-like kinase 1 (PLK1) activity suffer severe chromosome alignment defects, which is believed primarily due to unstable kinetochore-microtubule attachment. Here, we reveal a previously undescribed mechanism named ‘centromere disintegration’ that drives chromosome misalignment in PLK1-inactivated cells. We find that PLK1 inhibition does not necessarily compromise metaphase establishment, but instead its maintenance. We demonstrate that this is caused by unlawful unwinding of DNA by BLM helicase at a specific centromere domain underneath kinetochores. Under bipolar spindle pulling, the distorted centromeres are promptly decompacted into DNA threadlike molecules, leading to centromere rupture and whole-chromosome arm splitting. Consequently, chromosome alignment collapses. Our study unveils an unexpected role of PLK1 as a chromosome guardian to maintain centromere integrity for chromosome biorientation

The Transcription Factor YY1 Is a Substrate for Polo-Like Kinase 1 at the G2/M Transition of the Cell Cycle

Yin-Yang 1 (YY1) is an essential multifunctional zinc-finger protein. It has been shown over the past two decades to be a critical regulator of a vast array of biological processes, including development, cell proliferation and differentiation, DNA repair, and apoptosis. YY1 exerts its functions primarily as a transcription factor that can activate or repress gene expression, dependent on its spatial and temporal context. YY1 regulates a large number of genes involved in cell cycle transitions, many of which are oncogenes and tumor-suppressor genes. YY1 itself has been classified as an oncogene and was found to be upregulated in many cancer types. Unfortunately, our knowledge of what regulates YY1 is very minimal. Although YY1 has been shown to be a phosphoprotein, no kinase has ever been identified for the phosphorylation of YY1. Polo-like kinase 1 (Plk1) has emerged in the past few years as a major cell cycle regulator, particularly for cell division. Plk1 has been shown to play important roles in the G/M transition into mitosis and for the proper execution of cytokinesis, processes that YY1 has been shown to regulate also. Here, we present evidence that Plk1 directly phosphorylates YY1 in vitro and in vivo at threonine 39 in the activation domain. We show that this phosphorylation is cell cycle regulated and peaks at G2/M. This is the first report identifying a kinase for which YY1 is a substrate

The cellular geography of Aurora kinases

Aurora is the name given to a family of highly conserved protein kinases with essential roles in many aspects of cell division. Yeasts have a single Aurora kinase, whereas mammals have three: Aurora A, B and C. During mitosis, Aurora kinases regulate the structure and function of the cytoskeleton and chromosomes and the interactions between these two at the kinetochore. They also regulate signalling by the spindle-assembly checkpoint pathway and cytokinesis. Perturbation of Aurora kinase expression or function might lead to cancer