RESUMO
Proliferating cells must cross a point of no return before they replicate their DNA and divide. This commitment decision plays a fundamental role in cancer and degenerative diseases and has been proposed to be mediated by phosphorylation of retinoblastoma (Rb) protein. Here, we show that inactivation of the anaphase-promoting complex/cyclosome (APC(Cdh1)) has the necessary characteristics to be the point of no return for cell-cycle entry. Our study shows that APC(Cdh1) inactivation is a rapid, bistable switch initiated shortly before the start of DNA replication by cyclin E/Cdk2 and made irreversible by Emi1. Exposure to stress between Rb phosphorylation and APC(Cdh1) inactivation, but not after APC(Cdh1) inactivation, reverted cells to a mitogen-sensitive quiescent state, from which they can later re-enter the cell cycle. Thus, APC(Cdh1) inactivation is the commitment point when cells lose the ability to return to quiescence and decide to progress through the cell cycle.
Assuntos
Ciclossomo-Complexo Promotor de Anáfase/metabolismo , Proteínas Cdh1/metabolismo , Ciclo Celular , Ciclo Celular/efeitos dos fármacos , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Proteínas F-Box/metabolismo , Humanos , Mitógenos/toxicidade , Fosforilação , Proteína do Retinoblastoma/metabolismoRESUMO
Human cells license tens of thousands of origins of replication in G1 and then must stop all licensing before DNA synthesis in S phase to prevent re-replication and genome instability that ensue when an origin is licensed on replicated DNA. However, the E3 ubiquitin ligase CRL4Cdt2 only starts to degrade the licensing factor CDT1 after origin firing, raising the question of how cells prevent re-replication before CDT1 is fully degraded. Here, using quantitative microscopy and in-vitro-reconstituted human DNA replication, we show that CDT1 inhibits DNA synthesis during an overlap period when CDT1 is still present after origin firing. CDT1 inhibits DNA synthesis by suppressing CMG helicase at replication forks, and DNA synthesis commences once CDT1 is degraded. Thus, in contrast to the prevailing model that human cells prevent re-replication by strictly separating licensing from firing, licensing and firing overlap, and cells instead separate licensing from DNA synthesis.
Assuntos
Proteínas de Ciclo Celular , Replicação do DNA , Humanos , Fase S , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , DNA/genética , DNA Helicases/genética , DNA Helicases/metabolismoRESUMO
Tissue repair, immune defence and cancer progression rely on a vital cellular decision between quiescence and proliferation1,2. Mammalian cells proliferate by triggering a positive feedback mechanism3,4. The transcription factor E2F activates cyclin-dependent kinase 2 (CDK2), which in turn phosphorylates and inactivates the E2F inhibitor protein retinoblastoma (Rb). This action further increases E2F activity to express genes needed for proliferation. Given that positive feedback can inadvertently amplify small signals, understanding how cells keep this positive feedback in check remains a puzzle. Here we measured E2F and CDK2 signal changes in single cells and found that the positive feedback mechanism engages only late in G1 phase. Cells spend variable and often extended times in a reversible state of intermediate E2F activity before committing to proliferate. This intermediate E2F activity is proportional to the amount of phosphorylation of a conserved T373 residue in Rb that is mediated by CDK2 or CDK4/CDK6. Such T373-phosphorylated Rb remains bound on chromatin but dissociates from it once Rb is hyperphosphorylated at many sites, which fully activates E2F. The preferential initial phosphorylation of T373 can be explained by its relatively slower rate of dephosphorylation. Together, our study identifies a primed state of intermediate E2F activation whereby cells sense external and internal signals and decide whether to reverse and exit to quiescence or trigger the positive feedback mechanism that initiates cell proliferation.
Assuntos
Proliferação de Células , Quinase 2 Dependente de Ciclina , Fatores de Transcrição E2F , Proteína do Retinoblastoma , Fosforilação , Quinase 2 Dependente de Ciclina/metabolismo , Proteína do Retinoblastoma/metabolismo , Fatores de Transcrição E2F/metabolismo , Humanos , Animais , Camundongos , Análise de Célula Única , Cromatina/metabolismo , Fase G1 , Retroalimentação Fisiológica , Quinase 6 Dependente de Ciclina/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Linhagem CelularRESUMO
Directed cell migration is driven by the front-back polarization of intracellular signalling1-3. Receptor tyrosine kinases and other inputs activate local signals that trigger membrane protrusions at the front2,4-6. Equally important is a long-range inhibitory mechanism that suppresses signalling at the back to prevent the formation of multiple fronts7-9. However, the identity of this mechanism is unknown. Here we report that endoplasmic reticulum-plasma membrane (ER-PM) contact sites are polarized in single and collectively migrating cells. The increased density of these ER-PM contacts at the back provides the ER-resident PTP1B phosphatase more access to PM substrates, which confines receptor signalling to the front and directs cell migration. Polarization of the ER-PM contacts is due to microtubule-regulated polarization of the ER, with more RTN4-rich curved ER at the front and more CLIMP63-rich flattened ER at the back. The resulting ER curvature gradient leads to small and unstable ER-PM contacts only at the front. These contacts flow backwards and grow to large and stable contacts at the back to form the front-back ER-PM contact gradient. Together, our study suggests that the structural polarity mediated by ER-PM contact gradients polarizes cell signalling, directs cell migration and prolongs cell migration.
Assuntos
Membrana Celular , Movimento Celular , Retículo Endoplasmático , Humanos , Linhagem Celular , Membrana Celular/metabolismo , Polaridade Celular , Retículo Endoplasmático/metabolismo , Microtúbulos/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 1/metabolismo , Transdução de SinaisRESUMO
Junctions that connect the endoplasmic reticulum (ER) and the plasma membrane (PM) are unique yet ubiquitous subcellular compartments. Giordano et al. now report that extended synaptotagmins (E-Syts) promote their formation, providing fundamental insight into the molecular machinery controlling ER and plasma membrane crosstalk.
Assuntos
Cálcio/metabolismo , Membrana Celular/metabolismo , Retículo Endoplasmático/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Sinaptotagminas/metabolismo , HumanosRESUMO
Tissue homeostasis in metazoans is regulated by transitions of cells between quiescence and proliferation. The hallmark of proliferating populations is progression through the cell cycle, which is driven by cyclin-dependent kinase (CDK) activity. Here, we introduce a live-cell sensor for CDK2 activity and unexpectedly found that proliferating cells bifurcate into two populations as they exit mitosis. Many cells immediately commit to the next cell cycle by building up CDK2 activity from an intermediate level, while other cells lack CDK2 activity and enter a transient state of quiescence. This bifurcation is directly controlled by the CDK inhibitor p21 and is regulated by mitogens during a restriction window at the end of the previous cell cycle. Thus, cells decide at the end of mitosis to either start the next cell cycle by immediately building up CDK2 activity or to enter a transient G0-like state by suppressing CDK2 activity.
Assuntos
Quinase 2 Dependente de Ciclina/metabolismo , Mitose , Células 3T3 , Animais , Proliferação de Células , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Humanos , Camundongos , Proteína do Retinoblastoma/metabolismoRESUMO
The entry of mammalian cells into the DNA synthesis phase (S phase) represents a key event in cell division1. According to current models of the cell cycle, the kinase CDC7 constitutes an essential and rate-limiting trigger of DNA replication, acting together with the cyclin-dependent kinase CDK2. Here we show that CDC7 is dispensable for cell division of many different cell types, as determined using chemical genetic systems that enable acute shutdown of CDC7 in cultured cells and in live mice. We demonstrate that another cell cycle kinase, CDK1, is also active during G1/S transition both in cycling cells and in cells exiting quiescence. We show that CDC7 and CDK1 perform functionally redundant roles during G1/S transition, and at least one of these kinases must be present to allow S-phase entry. These observations revise our understanding of cell cycle progression by demonstrating that CDK1 physiologically regulates two distinct transitions during cell division cycle, whereas CDC7 has a redundant function in DNA replication.
Assuntos
Proteínas de Ciclo Celular , Fase G1 , Proteínas Serina-Treonina Quinases , Proteólise , Fase S , Animais , Proteínas de Ciclo Celular/metabolismo , Replicação do DNA , Camundongos , Proteínas Serina-Treonina Quinases/metabolismoRESUMO
Mitosis is triggered by the activation of Cdk1-cyclin B1 and its translocation from the cytoplasm to the nucleus. Positive feedback loops regulate the activation of Cdk1-cyclin B1 and help make the process irreversible and all-or-none in character. Here we examine whether an analogous process, spatial positive feedback, regulates Cdk1-cyclin B1 redistribution. We used chemical biology approaches and live-cell microscopy to show that nuclear Cdk1-cyclin B1 promotes the translocation of Cdk1-cyclin B1 to the nucleus. Mechanistic studies suggest that cyclin B1 phosphorylation promotes nuclear translocation and, conversely, nuclear translocation promotes cyclin B1 phosphorylation, accounting for the feedback. Interfering with the abruptness of Cdk1-cyclin B1 translocation affects the timing and synchronicity of subsequent mitotic events, underscoring the functional importance of this feedback. We propose that spatial positive feedback ensures a rapid, complete, robust, and irreversible transition from interphase to mitosis and suggest that bistable spatiotemporal switches may be widespread in biological regulation.
Assuntos
Proteína Quinase CDC2/metabolismo , Núcleo Celular/metabolismo , Ciclina B1/metabolismo , Retroalimentação , Mitose , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Ciclina B1/análise , Células HeLa , Humanos , Modelos Estatísticos , Fosforilação , Sirolimo/análogos & derivadosRESUMO
Cells escape the need for mitogens at a restriction point several hours before entering S phase. The restriction point has been proposed to result from CDK4/6 initiating partial Rb phosphorylation to trigger a bistable switch whereby cyclin E-CDK2 and Rb mutually reinforce each other to induce Rb hyperphosphorylation. Here, using single-cell analysis, we unexpectedly found that cyclin E/A-CDK activity can only maintain Rb hyperphosphorylation starting at the onset of S phase and that CDK4/6 activity, but not cyclin E/A-CDK activity, is required to hyperphosphorylate Rb throughout G1 phase. Mitogen removal in G1 results in a gradual loss of CDK4/6 activity with a high likelihood of cells sustaining Rb hyperphosphorylation until S phase, at which point cyclin E/A-CDK activity takes over. Thus, it is short-term memory, or transient hysteresis, in CDK4/6 activity following mitogen removal that sustains Rb hyperphosphorylation, demonstrating a probabilistic rather than an irreversible molecular mechanism underlying the restriction point.
Assuntos
Proliferação de Células , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/metabolismo , Células Epiteliais/efeitos dos fármacos , Pontos de Checagem da Fase G1 do Ciclo Celular , Mitógenos/farmacologia , Animais , Linhagem Celular , Relação Dose-Resposta a Droga , Células Epiteliais/enzimologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/enzimologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/enzimologia , Humanos , Camundongos , Modelos Biológicos , Fosforilação , Proteínas de Ligação a Retinoblastoma/metabolismo , Transdução de Sinais , Fatores de Tempo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Eukaryotic organelles can interact with each other through stable junctions where the two membranes are kept in close apposition. The junction that connects the endoplasmic reticulum to the plasma membrane (ER-PM junction) is unique in providing a direct communication link between the ER and the PM. In a recently discovered signaling process, STIM (stromal-interacting molecule) proteins sense a drop in ER Ca(2+) levels and directly activate Orai PM Ca(2+) channels across the junction space. In an inverse process, a voltage-gated PM Ca(2+) channel can directly open ER ryanodine-receptor Ca(2+) channels in striated-muscle cells. Although ER-PM junctions were first described 50 years ago, their broad importance in Ca(2+) signaling, as well as in the regulation of cholesterol and phosphatidylinositol lipid transfer, has only recently been realized. Here, we discuss research from different fields to provide a broad perspective on the structures and unique roles of ER-PM junctions in controlling signaling and metabolic processes.
Assuntos
Membrana Celular/química , Retículo Endoplasmático/química , Proteínas de Membrana/química , Animais , Comunicação Celular/fisiologia , Membrana Celular/metabolismo , Retículo Endoplasmático/metabolismo , Humanos , Membranas Intracelulares/química , Membranas Intracelulares/metabolismo , Proteínas de Membrana/metabolismo , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Transdução de Sinais/fisiologiaRESUMO
To orchestrate cell mechanics, trafficking, and motility, cytoskeletal filaments must assemble into higher-order networks whose local subcellular architecture and composition specify their functions. Cross-linking proteins bridge filaments at the nanoscale to control a network's µm-scale geometry, thereby conferring its mechanical properties and functional dynamics. While these interfilament linkages are key determinants of cytoskeletal function, their structural mechanisms remain poorly understood. Plastins/fimbrins are an evolutionarily ancient family of tandem calponin-homology domain (CHD) proteins required to construct multiple classes of actin networks, which feature diverse geometries specialized to power cytokinesis, microvilli and stereocilia biogenesis, and persistent cell migration. Here, we focus on the structural basis of actin network assembly by human T-plastin, a ubiquitously expressed isoform necessary for the maintenance of stable cellular protrusions generated by actin polymerization forces. By implementing a machine-learning-enabled cryo-electron microscopy pipeline for visualizing cross-linkers bridging multiple filaments, we uncover a sequential bundling mechanism enabling T-plastin to bridge pairs of actin filaments in both parallel and antiparallel orientations. T-plastin populates distinct structural landscapes in these two bridging orientations that are selectively compatible with actin networks featuring divergent architectures and functions. Our structural, biochemical, and cell biological data highlight inter-CHD linkers as key structural elements underlying flexible but stable cross-linking that are likely to be disrupted by T-plastin mutations that cause hereditary bone diseases.
Assuntos
Actinas , Glicoproteínas de Membrana , Proteínas dos Microfilamentos , Citoesqueleto de Actina/metabolismo , Actinas/química , Microscopia Crioeletrônica , Humanos , Glicoproteínas de Membrana/química , Proteínas dos Microfilamentos/química , PolimerizaçãoRESUMO
Mammalian cells integrate mitogen and stress signalling before the end of G1 phase to determine whether or not they enter the cell cycle1-4. Before cells can replicate their DNA in S phase, they have to activate cyclin-dependent kinases (CDKs), induce an E2F transcription program and inactivate the anaphase-promoting complex (APC/CCDH1, also known as the cyclosome), which is an E3 ubiquitin ligase that contains the co-activator CDH1 (also known as FZR, encoded by FZR1). It was recently shown that stress can return cells to quiescence after CDK2 activation and E2F induction but not after inactivation of APC/CCDH1, which suggests that APC/CCDH1 inactivation is the point of no return for cell-cycle entry 3 . Rapid inactivation of APC/CCDH1 requires early mitotic inhibitor 1 (EMI1)3,5, but the molecular mechanism that controls this cell-cycle commitment step is unknown. Here we show using human cell models that cell-cycle commitment is mediated by an EMI1-APC/CCDH1 dual-negative feedback switch, in which EMI1 is both a substrate and an inhibitor of APC/CCDH1. The inactivation switch triggers a transition between a state with low EMI1 levels and high APC/CCDH1 activity during G1 and a state with high EMI1 levels and low APC/CCDH1 activity during S and G2. Cell-based analysis, in vitro reconstitution and modelling data show that the underlying dual-negative feedback is bistable and represents a robust irreversible switch. Our study suggests that mammalian cells commit to the cell cycle by increasing CDK2 activity and EMI1 mRNA expression to trigger a one-way APC/CCDH1 inactivation switch that is mediated by EMI1 transitioning from acting as a substrate of APC/CCDH1 to being an inhibitor of APC/CCDH1.
Assuntos
Proteínas Cdh1/antagonistas & inibidores , Proteínas Cdh1/metabolismo , Proteínas de Ciclo Celular/metabolismo , Ciclo Celular/fisiologia , Proteínas F-Box/metabolismo , Proteínas de Ciclo Celular/genética , Ciclina E/metabolismo , Quinase 2 Dependente de Ciclina/metabolismo , Proteínas F-Box/genética , Retroalimentação Fisiológica , Fase G1 , Células HeLa , Humanos , Fase SRESUMO
BACKGROUND: The growing prominence of Artificial Intelligence (AI) in medicine introduces both transformative possibilities and potential challenges. Our study focuses on the current status and perceptions of AI in Head and Neck Surgery (HNS), examining its utilization, benefits, ethical concerns, and protective measures. OBJECTIVES: The study aims to illuminate the existing landscape of AI in HNS in Germany. MATERIALS AND METHODS: Conducted through a questionnaire, key aspects include its current usage, potential applications (e.g., diagnosis, surgical planning), anticipated benefits (e.g., improved patient care ), and significant ethical concerns (e.g., miscalculations by AI, data privacy). RESULTS: The survey reveals limited AI adoption in HNS, with substantial potential for improvement. Ethical considerations, especially miscalculations by AI and data privacy, emerge as central issues. The survey emphasizes the crucial role of physician control and the need for legal oversight to address concerns related to AI integration. While AI's presence in HNS is modest, the study identifies opportunities for enhancement. Ethical guidelines and practitioner-centric control are vital for discussions surrounding AI integration. CONCLUSIONS: This research underscores the significance of ethical considerations and practitioner control in the context of AI in surgical practices. It highlights the demand for targeted training to empower practitioners in navigating the complexities of AI technologies in healthcare, ensuring responsible and patient-centric implementation.
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Regulation of cell proliferation is necessary for immune responses, tissue repair, and upkeep of organ function to maintain human health. When proliferating cells complete mitosis, a fraction of newly born daughter cells immediately enter the next cell cycle, while the remaining cells in the same population exit to a transient or persistent quiescent state. Whether this choice between two cell-cycle pathways is due to natural variability in mitogen signalling or other underlying causes is unknown. Here we show that human cells make this fundamental cell-cycle entry or exit decision based on competing memories of variable mitogen and stress signals. Rather than erasing their signalling history at cell-cycle checkpoints before mitosis, mother cells transmit DNA damage-induced p53 protein and mitogen-induced cyclin D1 (CCND1) mRNA to newly born daughter cells. After mitosis, the transferred CCND1 mRNA and p53 protein induce variable expression of cyclin D1 and the CDK inhibitor p21 that almost exclusively determines cell-cycle commitment in daughter cells. We find that stoichiometric inhibition of cyclin D1-CDK4 activity by p21 controls the retinoblastoma (Rb) and E2F transcription program in an ultrasensitive manner. Thus, daughter cells control the proliferation-quiescence decision by converting the memories of variable mitogen and stress signals into a competition between cyclin D1 and p21 expression. We propose a cell-cycle control principle based on natural variation, memory and competition that maximizes the health of growing cell populations.
Assuntos
Ciclo Celular/fisiologia , Mitógenos/metabolismo , Transdução de Sinais , Estresse Fisiológico , Proteína Supressora de Tumor p53/metabolismo , Pontos de Checagem do Ciclo Celular , Proliferação de Células , Ciclina D1/antagonistas & inibidores , Ciclina D1/genética , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Dano ao DNA , Fatores de Transcrição E2F/metabolismo , Humanos , Mitose , Retinoblastoma/metabolismo , Retinoblastoma/patologiaRESUMO
The stability of perovskite oxide catalysts for the oxygen evolution reaction (OER) plays a critical role in their applicability in water splitting concepts. Decomposition of perovskite oxides under applied potential is typically linked to cation leaching and amorphization of the material. However, structural changes and phase transformations at the catalyst surface were also shown to govern the activity of several perovskite electrocatalysts under applied potential. Hence, it is crucial for the rational design of durable perovskite catalysts to understand the interplay between the formation of active surface phases and stability limitations under OER conditions. In the present study, we reveal a surface-dominated activation and deactivation mechanism of the prominent electrocatalyst La0.6Sr0.4CoO3-δ under steady-state OER conditions. Using a multiscale microscopy and spectroscopy approach, we identify the evolving Co-oxyhydroxide as catalytically active surface species and La-hydroxide as inactive species involved in the transient degradation behavior of the catalyst. While the leaching of Sr results in the formation of mixed surface phases, which can be considered as a part of the active surface, the gradual depletion of Co from a self-assembled active CoO(OH) phase and the relative enrichment of passivating La(OH)3 at the electrode surface result in the failure of the perovskite catalyst under applied potential.
RESUMO
Cell migration on two-dimensional substrates is typically characterized by lamellipodia at the leading edge, mature focal adhesions and spread morphologies. These observations result from adherent cell migration studies on stiff, elastic substrates, because most cells do not migrate on soft, elastic substrates. However, many biological tissues are soft and viscoelastic, exhibiting stress relaxation over time in response to a deformation. Here, we have systematically investigated the impact of substrate stress relaxation on cell migration on soft substrates. We observed that cells migrate minimally on substrates with an elastic modulus of 2 kPa that are elastic or exhibit slow stress relaxation, but migrate robustly on 2-kPa substrates that exhibit fast stress relaxation. Strikingly, migrating cells were not spread out and did not extend lamellipodial protrusions, but were instead rounded, with filopodia protrusions extending at the leading edge, and exhibited small nascent adhesions. Computational models of cell migration based on a motor-clutch framework predict the observed impact of substrate stress relaxation on cell migration and filopodia dynamics. Our findings establish substrate stress relaxation as a key requirement for robust cell migration on soft substrates and uncover a mode of two-dimensional cell migration marked by round morphologies, filopodia protrusions and weak adhesions.
Assuntos
Movimento Celular , Pseudópodes/metabolismo , Membrana Basal/metabolismo , Fenômenos Biomecânicos , Adesão Celular , Linhagem Celular , Linhagem Celular Tumoral , Elasticidade , HumanosRESUMO
The laser-induced ablation threshold of c-plane GaN films upon exposure to ultrashort laser pulses was investigated for different wavelengths from the IR to the UV range and pulse widths between 0.34 and 10 ps. The one-pulse ablation threshold ranges between 0.15 and 3â J/cm2 and shows an increase with the wavelength and the pulse width, except for deep UV pulses. Based on a rate equation model, we attribute this behavior to the efficiency of seed carrier generation by interband absorption. In addition, the multi-pulse ablation threshold was analyzed. Accumulation effects are more prominent in case of IR than with UV pulses and are closely linked to damage precursors. By a thorough structural investigation, we demonstrate that threading dislocations, especially those with a screw component, significantly contribute to laser damage, since they provide a variety of dispersed states within the band gap.
RESUMO
We provide a correction due to an erroneous repetition rate of one of the laser systems (90 fs pulse duration) in our previously published paper [Opt. Express28, 25037 (2020)10.1364/OE.399771].
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We demonstrate supercontinuum generation in a liquid-core microstructured optical fiber using carbon disulfide as the core material. The fiber provides a specific dispersion landscape with a zero-dispersion wavelength approaching the telecommunication domain where the corresponding capillary-type counterpart shows unsuitable dispersion properties for soliton fission. The experiments were conducted using two pump lasers with different pulse duration (30 fs and 90 fs) giving rise to different non-instantaneous contributions of carbon disulfide in each case. The presented results demonstrate an extraordinary high conversion efficiency from pump to soliton and to dispersive wave, overall defining a platform that enables studying the impact of non-instantaneous responses on ultrafast soliton dynamics and coherence using straightforward pump lasers and diagnostics.
RESUMO
Mammalian cells have a remarkable capacity to compensate for heterozygous gene loss or extra gene copies. One exception is Down syndrome (DS), where a third copy of chromosome 21 mediates neurogenesis defects and lowers the frequency of solid tumors. Here we combine live-cell imaging and single-cell analysis to show that increased dosage of chromosome 21-localized Dyrk1a steeply increases G1 cell cycle duration through direct phosphorylation and degradation of cyclin D1 (CycD1). DS-derived fibroblasts showed analogous cell cycle changes that were reversed by Dyrk1a inhibition. Furthermore, reducing Dyrk1a activity increased CycD1 expression to force a bifurcation, with one subpopulation of cells accelerating proliferation and the other arresting proliferation by costabilizing CycD1 and the CDK inhibitor p21. Thus, dosage of Dyrk1a repositions cells within a p21-CycD1 signaling map, directing each cell to either proliferate or to follow two distinct cell cycle exit pathways characterized by high or low CycD1 and p21 levels.