Oncogene-Induced Mitotic Stress: p53 and pRb Get Mad Too

Oncogenic signaling frequently results in unscheduled overexpression of cell cycle proteins and replicative stress. In this issue of Cancer Cell, Schvartzman et al. show that inactivation of p53 or pRb induces Mad2, and the overexpression of this mitotic checkpoint protein is a necessary event during oncogene-induced mitotic stress

E2F7 regulates transcription and maturation of multiple microRNAs to restrain cell proliferation

This work was supported by the Spanish Ministry [SAF2012-33551, co-funded by the European RegionalDevelopment fund to A.M.Z., SAF2012-38215 to M.M.,SAF2014-57791-REDC to A.M.Z. and to M.M.]; BasqueGovernment [IT634-13 to A.M.Z.]; University of theBasque Country UPV/EHU [UFI1120 to A.M.Z.]; Excellence Network CellSYS [BFU2014-52125-REDT to M.M.];Comunidad de Madrid [S2010/BMD-2470 to M.M.];Basque Government Fellowship for graduate studies (to J.M.). Funding for open access charge: Basque Government [IT634-13]. Conflict of interest statement. None declared.E2F transcription factors (E2F1-8) are known to coordinately regulate the expression of a plethora of target genes, including those coding for microRNAs (miRNAs), to control cell cycle progression. Recent work has described the atypical E2F factor E2F7 as a transcriptional repressor of cell cycle-related protein-coding genes. However, the contribution of E2F7 to miRNA gene expression during the cell cycle has not been defined. We have performed a genome-wide RNA sequencing analysis to identify E2F7-regulated miRNAs and show that E2F7 plays as a major role in the negative regulation of a set of miRNAs that promote cellular proliferation. We provide mechanistic evidence for an interplay between E2F7 and the canonical E2F factors E2F1-3 in the regulation of multiple miRNAs. We show that miR-25, -26a, -27b, -92a and -7 expression is controlled at the transcriptional level by the antagonistic activity of E2F7 and E2F1-3. By contrast, let-7 miRNA expression is controlled indirectly through a novel E2F/c-MYC/LIN28B axis, whereby E2F7 and E2F1-3 modulate c-MYC and LIN28B levels to impact let-7 miRNA processing and maturation. Taken together, our data uncover a new regulatory network involving transcriptional and post-transcriptional mechanisms controlled by E2F7 to restrain cell cycle progression through repression of proliferation-promoting miRNAs.S

The APC/C cofactor Cdh1 prevents replicative stress and p53-dependent cell death in neural progenitors

The E3-ubiquitin ligase APC/C-Cdh1 is essential for endoreduplication but its relevance in the mammalian mitotic cell cycle is still unclear. Here we show that genetic ablation of Cdh1 in the developing nervous system results in hypoplastic brain and hydrocephalus. These defects correlate with enhanced levels of Cdh1 substrates and increased entry into the S phase in neural progenitors. However, cell division is prevented in the absence of Cdh1 due to hyperactivation of cyclin-dependent kinases, replicative stress, induction of p53, G2 arrest and apoptotic death of these progenitor cells. Concomitant ablation of p53 rescues apoptosis but not replicative stress, resulting in the presence of damaged neurons throughout the adult brain. These data indicate that the inactivation of Cdh1 in vivo results in replicative stress, cell cycle arrest and cell death, supporting recent therapeutic proposals aimed to inhibit the APC/C in tumours.M.E. was supported by the Spanish Ministry of Economy and Competitiveness (MINECO). This work was funded by grants from the Foundation Ramón Areces and MINECO SAF2012-38215 to M.M.). The Cell Division and Cancer Group of the CNIO are supported by the OncoCycle Programme (S2010/BMD-2470) from the Comunidad de Madrid, the OncoBIO Consolider-Ingenio 2010 Programme (MINECO, CSD2007-00017) and the European Union Seventh Framework Programme (MitoSys project; HEALTH-F5-2010-241548).Peer Reviewe

PLK1 regulates centrosome migration and spindle dynamics in male mouse meiosis

Cell division requires the regulation of karyokinesis and cytokinesis, which includes an essential role of the achromatic spindle. Although the functions of centrosomes are well characterised in somatic cells, their role during vertebrate spermatogenesis remains elusive. We have studied the dynamics of the meiotic centrosomes in male mouse during both meiotic divisions. Results show that meiotic centrosomes duplicate twice: first duplication occurs in the leptotene/zygotene transition, while the second occurs in interkinesis. The maturation of duplicated centrosomes during the early stages of prophase I and II are followed by their separation and migration to opposite poles to form bipolar spindles I and II. The study of the genetic mouse model Plk1(Δ/Δ) indicates a central role of Polo-like kinase 1 in pericentriolar matrix assembly, in centrosome maturation and migration, and in the formation of the bipolar spindles during spermatogenesis. In addition, in vitro inhibition of Polo-like kinase 1 and Aurora A in organotypic cultures of seminiferous tubules points out to a prominent role of both kinases in the regulation of the formation of meiotic bipolar spindlesThis work was supported by funding from Ministerio de Economía, Industria y Competitividad, Gobierno de España (MINECO) (Spain) [grant number BFU2014-53681-P and MEIONET (BFU2015-71786-REDT) to J. A. S. and grant number RTI2018-095582-B-I00 to M. M.]

p27Kip1 V109G as a biomarker for CDK4/6 inhibitors indication in hormone receptor–positive breast cancer

Biomarker; Hormone receptor-positive; Breast cancerBiomarcador; Cáncer de mama; Receptor hormonal positivoBiomarcador; Càncer de mama; Receptor hormonal positiuCDK4/6 inhibitors benefit a minority of patients who receive them in the breast cancer adjuvant setting. p27Kip1 is a protein that inhibits CDK/Cyclin complexes. We hypothesized that single-nucleotide polymorphisms that impaired p27Kip1 function could render patients refractory to endocrine therapy but responsive to CDK4/6 inhibitors, narrowing the patient subpopulation that requires CDK4/6 inhibitors. We found that the p27Kip1 V109G single-nucleotide polymorphism is homozygous in approximately 15% of hormone-positive breast cancer patients. Polymorphic patients experience rapid failure in response to endocrine monotherapy compared with wild-type or heterozygous patients in the first-line metastatic setting (progression-free survival: 92 vs 485 days, P < .001); when CDK4/6 inhibitors are added, the differences disappear (progression-free survival: 658 vs 761 days, P = .92). As opposed to wild-type p27Kip1, p27Kip1 V109G is unable to suppress the kinase activity of CDK4 in the presence of endocrine inhibitors; however, palbociclib blocks CDK4 kinase activity regardless of the p27Kip1 status. p27Kip1 genotyping could constitute a tool for treatment selection.MM is supported by the Spanish Ministry of Science and Innovation (RTI2018-095582-B-100; PLEC2021-007892 and RED2018-102723-T), AES (DTS21/00132) and Comunidad de Madrid (B2017/BMD-3884 and Y2020/BIO-6519). MQF is a recipient of the following grants: AES—PI 19/00454 funded by the ISCIII and co-funded by the European Regional Development Fund (ERDF), and B2017/BMD3733 (Immunothercan-CM) – Call for Coordinated Research Groups from the Madrid Region—Madrid Regional Government—ERDF funds. This study was also funded by a donation from CRIS Contra El Cancer Foundation

Biochemical analyses reveal amino acid residues critical for cell cycle-dependent phosphorylation of human Cdc14A phosphatase by cyclin-dependent kinase 1

Cdc14 enzymes compose a family of highly conserved phosphatases that are present in a wide range of organisms, including yeast and humans, and that preferentially reverse the phosphorylation of Cyclin-Dependent Kinase (Cdk) substrates. The budding yeast Cdc14 orthologue has essential functions in the control of late mitosis and cytokinesis. In mammals, however, the two Cdc14 homologues, Cdc14A and Cdc14B, do not play a prominent role in controlling late mitotic events, suggesting that some Cdc14 functions are not conserved across species. Moreover, in yeast, Cdc14 is regulated by changes in its subcellular location and by phosphorylation events. In contrast, little is known about the regulation of human Cdc14 phosphatases. Here, we have studied how the human Cdc14A orthologue is regulated during the cell cycle. We found that Cdc14A is phosphorylated on Ser411, Ser453 and Ser549 by Cdk1 early in mitosis and becomes dephosphorylated during late mitotic stages. Interestingly, in vivo and in vitro experiments revealed that, unlike in yeast, Cdk1-mediated phosphorylation of human Cdc14A did not control its catalytic activity but likely modulated its interaction with other proteins in early mitosis. These findings point to differences in Cdk1-mediated mechanisms of regulation between human and yeast Cdc14 orthologues.We thank S. Andrés for technical assistance and other members of laboratory for helpful discussions. We are grateful to Dr. I. García-Higuera and S. Moreno (IBFG, Salamanca) for Cdh1 plasmids and anti-Plk1 antibodies, Dr. J. Dong (Univ. Nebraska Medical Center, Nebraska) for the KIBRA plasmids, Dr. C. Guerrero (IBMCC, Salamanca) for the anti-PP2A antibody and Dr. Jallepalli (Memorial Sloan Kettering Cancer Center, New York) for the RPE Cdc14A − / − cells. We are grateful to the proteomics facility of Centro de Investigación del Cáncer, Salamanca, Spain, where the proteomic analysis was performed, Grant PRB2 (IPT13/0001 - ISCIII-SGEFI / FEDER). This work was funded by grants from the Spanish Ministry of Economy, Industry and Competitiveness (MINECO; BFU2015-69709-P and SAF2015-69920-R). S.O. was supported by a FPU fellowship from the Spanish Ministry of Education and P.A. was supported by a JAE-Predoctoral fellowship from the Spanish National research Council (CSIC).S

Dependency of the Spindle Assembly Checkpoint on Cdk1 Renders the Anaphase Transition Irreversible

SummaryActivation of anaphase-promoting complex/cyclosome (APC/CCdc20) by Cdc20 is delayed by the spindle assembly checkpoint (SAC). When all kinetochores come under tension, the SAC is turned off and APC/CCdc20 degrades cyclin B and securin, which activates separase [1]. The latter then cleaves cohesin holding sister chromatids together [2]. Because cohesin cleavage also destroys the tension responsible for turning off the SAC, cells must possess a mechanism to prevent SAC reactivation during anaphase, which could be conferred by a dependence of the SAC on Cdk1 [3–5]. To test this, we analyzed mouse oocytes and embryos expressing nondegradable cyclin B together with a Cdk1-resistant form of separase. After biorientation and SAC inactivation, APC/CCdc20 activates separase but the resulting loss of (some) cohesion is accompanied by SAC reactivation and APC/CCdc20 inhibition, which aborts the process of further securin degradation. Cyclin B is therefore the only APC/CCdc20 substrate whose degradation at the onset of anaphase is necessary to prevent SAC reactivation. The mutual activation of tension sensitive SAC and Cdk1 creates a bistable system that ensures complete activation of separase and total downregulation of Cdk1 when all chromosomes have bioriented

Harnessing transcriptionally driven chromosomal instability adaptation to target therapy-refractory lethal prostate cancer

Chromosomal instability; Lethal prostate cancer; Therapy resistanceInestabilidad cromosómica; Cáncer de próstata letal; Resistencia a la terapiaInestabilitat cromosòmica; Càncer de pròstata letal; Resistència a la teràpiaMetastatic prostate cancer (PCa) inevitably acquires resistance to standard therapy preceding lethality. Here, we unveil a chromosomal instability (CIN) tolerance mechanism as a therapeutic vulnerability of therapy-refractory lethal PCa. Through genomic and transcriptomic analysis of patient datasets, we find that castration and chemotherapy-resistant tumors display the highest CIN and mitotic kinase levels. Functional genomics screening coupled with quantitative phosphoproteomics identify MASTL kinase as a survival vulnerability specific of chemotherapy-resistant PCa cells. Mechanistically, MASTL upregulation is driven by transcriptional rewiring mechanisms involving the non-canonical transcription factors androgen receptor splice variant 7 and E2F7 in a circuitry that restrains deleterious CIN and prevents cell death selectively in metastatic therapy-resistant PCa cells. Notably, MASTL pharmacological inhibition re-sensitizes tumors to standard therapy and improves survival of pre-clinical models. These results uncover a targetable mechanism promoting high CIN adaptation and survival of lethal PCa.Research was supported by NIH/NCI grants P30CA08748 (to R.C.H.); R01CA207311 and R01CA261925 (to J.D.-D.); K22CA207458 and R01CA237398 (to V.R.-B.); and funding to V.R.-B. from the Mayo Clinic Foundation, The Margaret Q. Landenberger Research Foundation, The W.W. Smith Charitable Trust, The AACR, and The Prostate Cancer Foundation (PCF)

A Gene Encoding Arginyl-tRNA Synthetase Is Located in the Upstream Region of the lysA Gene in Brevibacterium lactofermentum: Regulation of argS-lysA Cluster Expression by Arginine

International audienceThe Brevibacterium lactofermentum argS gene, which encodes an arginyl-tRNA synthetase, was identified in the upstream region of the lysA gene. The cloned gene was sequenced; it encodes a 550-amino-acid protein with an Mr of 59,797. The deduced amino acid sequence showed 28% identical and 49% similar residues when compared with the sequence of the Escherichia coli arginyl-tRNA synthetase. The B. lactofermentum enzyme showed the highly conserved motifs of class I aminoacyl-tRNA synthetases. Expression of the argS gene in B. lactofermentum and E. coli resulted in an increase in aminoacyl-tRNA synthetase activity, correlated with the presence in sodium dodecyl sulfate-polyacrylamide gels of a clear protein band that corresponds to this enzyme. One single transcript of about 3,000 nucleotides and corresponding to the B. lactofermentum argS-lysA operon was identified. The transcription of these genes is repressed by lysine and induced by arginine, showing an interesting pattern of biosynthetic interlock between the pathways of both amino acids in corynebacteria