scholarly journals A Cdk4/6-dependent phosphorylation gradient regulates the early to late G1 phase transition

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Manuel Kaulich ◽  
Verena M. Link ◽  
John D. Lapek ◽  
Yeon J. Lee ◽  
Christopher K. Glass ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Single Cell ◽  
Specific Binding ◽  
Cellular Protein ◽  
G1 Phase ◽  
Protein Targets ◽  
Cell Cycle Entry ◽  
Phase Progression ◽  
Phase Functions ◽  
Cdk4 Inhibition

AbstractDuring early G1 phase, Rb is exclusively mono-phosphorylated by cyclin D:Cdk4/6, generating 14 different isoforms with specific binding patterns to E2Fs and other cellular protein targets. While mono-phosphorylated Rb is dispensable for early G1 phase progression, interfering with cyclin D:Cdk4/6 kinase activity prevents G1 phase progression, questioning the role of cyclin D:Cdk4/6 in Rb inactivation. To dissect the molecular functions of cyclin D:Cdk4/6 during cell cycle entry, we generated a single cell reporter for Cdk2 activation, RB inactivation and cell cycle entry by CRISPR/Cas9 tagging endogenous p27 with mCherry. Through single cell tracing of Cdk4i cells, we identified a time-sensitive early G1 phase specific Cdk4/6-dependent phosphorylation gradient that regulates cell cycle entry timing and resides between serum-sensing and cyclin E:Cdk2 activation. To reveal the substrate identity of the Cdk4/6 phosphorylation gradient, we performed whole proteomic and phospho-proteomic mass spectrometry, and identified 147 proteins and 82 phospho-peptides that significantly changed due to Cdk4 inhibition in early G1 phase. In summary, we identified novel (non-Rb) cyclin D:Cdk4/6 substrates that connects early G1 phase functions with cyclin E:Cdk2 activation and Rb inactivation by hyper-phosphorylation.

scholarly journals CLN3 expression is sufficient to restore G1-to-S-phase progression in Saccharomyces cerevisiae mutants defective in translation initiation factor eIF4E

1999 ◽  
Vol 340 (1) ◽  
pp. 135-141 ◽  
Author(s):  
Parisa DANAIE ◽  
Michael ALTMANN ◽  
Michael N. HALL ◽  
Hans TRACHSEL ◽  
Stephen B. HELLIWELL
Keyword(s):  
Cell Cycle ◽  
S Phase ◽  
Translation Initiation Factor ◽  
Yeast Cells ◽  
Initiation Factor ◽  
G1 Phase ◽  
Mrna Levels ◽  
Wild Type ◽  
Phase Progression ◽  
Type Gene

The essential cap-binding protein (eIF4E) of Saccharomycescerevisiae is encoded by the CDC33 (wild-type) gene, originally isolated as a mutant, cdc33-1, which arrests growth in the G1 phase of the cell cycle at 37 °C. We show that other cdc33 mutants also arrest in G1. One of the first events required for G1-to-S-phase progression is the increased expression of cyclin 3. Constructs carrying the 5ʹ-untranslated region of CLN3 fused to lacZ exhibit weak reporter activity, which is significantly decreased in a cdc33-1 mutant, implying that CLN3 mRNA is an inefficiently translated mRNA that is sensitive to perturbations in the translation machinery. A cdc33-1 strain expressing either stable Cln3p (Cln3-1p) or a hybrid UBI4 5ʹ-CLN3 mRNA, whose translation displays decreased dependence on eIF4E, arrested randomly in the cell cycle. In these cells CLN2 mRNA levels remained high, indicating that Cln3p activity is maintained. Induction of a hybrid UBI4 5ʹ-CLN3 message in a cdc33-1 mutant previously arrested in G1 also caused entry into a new cell cycle. We conclude that eIF4E activity in the G1-phase is critical in allowing sufficient Cln3p activity to enable yeast cells to enter a new cell cycle.

Hydroquinone and catechol interfere with T cell cycle entry and progression through the G1 phase

2003 ◽  
Vol 39 (16) ◽  
pp. 995-1001 ◽  
Author(s):  
Jesica M McCue ◽  
Sabine Lazis ◽  
J John Cohen ◽  
Jaime F Modiano ◽  
Brian M Freed
Keyword(s):  
Cell Cycle ◽  
T Cell ◽  
G1 Phase ◽  
Cell Cycle Entry

A single cell approach to understanding cell cycle entry in cancer

2016 ◽  
Vol 61 ◽  
pp. S172
Author(s):  
A. Barr ◽  
F.S. Heldt ◽  
S. Cooper ◽  
B. Novak ◽  
C. Bakal
Keyword(s):  
Cell Cycle ◽  
Single Cell ◽  
Cell Cycle Entry

Abstract 238: Single-Cell Transcriptomic Analysis and Patient-Specific IPSCs Reveal Dysregulated Cell Cycle in Coronary Endothelial Cell in Hypoplastic Left Heart Syndrome

2020 ◽  
Vol 127 (Suppl_1) ◽  
Author(s):  
Mingxia Gu ◽  
Yifei Miao ◽  
Xin Zhou ◽  
Lei Tian ◽  
Marcy Martin ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Endothelial Cell ◽  
Left Ventricle ◽  
Single Cell ◽  
Hypoplastic Left Heart Syndrome ◽  
Human Heart ◽  
G1 Phase ◽  
Left Heart ◽  
Hypoplastic Left Heart ◽  
Left Heart Syndrome

Hypoplastic left heart syndrome (HLHS) is a single ventricle congenital heart disease that results in severe underdevelopment of the left ventricle, mitral valve, aortic valve, and ascending aorta. Early serial postmortem examinations also revealed a high rate of coronary anomalies in HLHS, which included multiple ventriculo-coronary arterial connections as well as thick-walled and kinked coronary arteries. A previous study showed that fetal hypoplastic left hearts had a reduced endothelial cell (EC) population and lower capillary density compared with normal hearts. However, the mechanism underlying coronary abnormalities associated with HLHS remains unknown. Thus, we generated induced pluripotent stem cells derived ECs (iPSC-ECs) from three HLHS patients and three age-matched controls. Single Cell RNA-Seq (scRNA-seq) profiling identified both endocardial (NPR3 + /CDH5 + ) and coronary endothelial populations (APLN + /CDH5 + ) from the heterogeneous iPSC-ECs. Intriguingly, a subcluster of the coronary endothelial cells (CECs) with cell cycle arrest was specifically enriched in HLHS patients. Further cell cycle analysis showed that 30.6% of the HLHS cells were trapped in the G1 phase, while the majority of the control CECs entered cell cycle normally. Additionally, the cell cycle differences between control and HLHS was only seen in CECs, not in the endocardial population. To verify our transcriptomic analysis, we applied negative cell sorting (NPR3 - /CDH5 + ) on iPSC-ECs to purify CECs (iCECs) and confirmed that HLHS iCECs showed profound reduction of cell cycle/proliferative genes ( KI67, PCNA, CCNA2, CCNB1 ) and abnormal induction of CCND2 , which is the hallmark of G1 phase. BrdU assays also indicated suppressed proliferation in HLHS iCECs. Furthermore, we profiled the transcriptome from a human heart with an underdevelopment left ventricle (ULV) at single cell resolution. When compared to the normal human heart, pathway enrichment analysis of differentially expressed genes in ULV hearts demonstrated reduced cell proliferation in the CEC subpopulation. Here, we identified that CECs from HLHS patients exerted proliferative defects that can potentially impede the development of vascular/capillary structure and cause related functional deficiencies. Reformation of the coronary defect provides a promising therapeutic strategy to prevent HLHS deterioration.

scholarly journals The duration of mitosis and daughter cell size are modulated by nutrients in budding yeast

2017 ◽  
Vol 216 (11) ◽  
pp. 3463-3470 ◽  
Author(s):  
Ricardo M. Leitao ◽  
Douglas R. Kellogg
Keyword(s):  
Cell Cycle ◽  
Cell Size ◽  
Daughter Cell ◽  
Slow Growth ◽  
Budding Yeast ◽  
Size Control ◽  
G1 Phase ◽  
Large Reduction ◽  
Duration Of Mitosis ◽  
Cell Cycle Entry

The size of nearly all cells is modulated by nutrients. Thus, cells growing in poor nutrients can be nearly half the size of cells in rich nutrients. In budding yeast, cell size is thought to be controlled almost entirely by a mechanism that delays cell cycle entry until sufficient growth has occurred in G1 phase. Here, we show that most growth of a new daughter cell occurs in mitosis. When the rate of growth is slowed by poor nutrients, the duration of mitosis is increased, which suggests that cells compensate for slow growth in mitosis by increasing the duration of growth. The amount of growth required to complete mitosis is reduced in poor nutrients, leading to a large reduction in cell size. Together, these observations suggest that mechanisms that control the extent of growth in mitosis play a major role in cell size control in budding yeast.

scholarly journals Functional overlap among distinct G1/S inhibitory pathways allows robust G1 arrest by yeast mating pheromones

2013 ◽  
Vol 24 (23) ◽  
pp. 3675-3688 ◽  
Author(s):  
Patricia A. Pope ◽  
Peter M. Pryciak
Keyword(s):  
Cell Cycle ◽  
G1 Phase ◽  
G1 Arrest ◽  
Cdk Inhibitor ◽  
Transcriptional Repressors ◽  
Mating Pheromones ◽  
Cell Cycle Entry ◽  
Inhibitory Pathways ◽  
Yeast Mating ◽  
S Genes

In budding yeast, mating pheromones arrest the cell cycle in G1 phase via a pheromone-activated Cdk-inhibitor (CKI) protein, Far1. Alternate pathways must also exist, however, because deleting the cyclin CLN2 restores pheromone arrest to far1∆ cells. Here we probe whether these alternate pathways require the G1/S transcriptional repressors Whi5 and Stb1 or the CKI protein Sic1, whose metazoan analogues (Rb or p27) antagonize cell cycle entry. Removing Whi5 and Stb1 allows partial escape from G1 arrest in far1∆ cln2∆ cells, along with partial derepression of G1/S genes, which implies a repressor-independent route for inhibiting G1/S transcription. This route likely involves pheromone-induced degradation of Tec1, a transcriptional activator of the cyclin CLN1, because Tec1 stabilization also causes partial G1 escape in far1∆ cln2∆ cells, and this is additive with Whi5/Stb1 removal. Deleting SIC1 alone strongly disrupts Far1-independent G1 arrest, revealing that inhibition of B-type cyclin-Cdk activity can empower weak arrest pathways. Of interest, although far1∆ cln2∆ sic1∆ cells escaped G1 arrest, they lost viability during pheromone exposure, indicating that G1 exit is deleterious if the arrest signal remains active. Overall our findings illustrate how multiple distinct G1/S-braking mechanisms help to prevent premature cell cycle commitment and ensure a robust signal-induced G1 arrest.

scholarly journals A single-cell–resolution fate map of endoderm reveals demarcation of pancreatic progenitors by cell cycle

2021 ◽  
Vol 118 (25) ◽  
pp. e2025793118
Author(s):  
Yun Yang ◽  
Hao Wang ◽  
Jia He ◽  
Wenchao Shi ◽  
Zhanmei Jiang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Single Cell ◽  
Cell Fate ◽  
Progenitor Cell ◽  
Developmental Trajectories ◽  
Cell Labeling ◽  
G1 Phase ◽  
Fate Map ◽  
Pancreatic Progenitor ◽  
Foregut Endoderm

A progenitor cell could generate a certain type or multiple types of descendant cells during embryonic development. To make all the descendant cell types and developmental trajectories of every single progenitor cell clear remains an ultimate goal in developmental biology. Characterizations of descendant cells produced by each uncommitted progenitor for a full germ layer represent a big step toward the goal. Here, we focus on early foregut endoderm, which generates foregut digestive organs, including the pancreas, liver, foregut, and ductal system, through distinct lineages. Using unbiased single-cell labeling techniques, we label every individual zebrafish foregut endodermal progenitor cell out of 216 cells to visibly trace the distribution and number of their descendant cells. Hence, single-cell–resolution fate and proliferation maps of early foregut endoderm are established, in which progenitor regions of each foregut digestive organ are precisely demarcated. The maps indicate that the pancreatic endocrine progenitors are featured by a cell cycle state with a long G1 phase. Manipulating durations of the G1 phase modulates pancreatic progenitor populations. This study illustrates foregut endodermal progenitor cell fate at single-cell resolution, precisely demarcates different progenitor populations, and sheds light on mechanistic insights into pancreatic fate determination.

scholarly journals The duration of mitosis and daughter cell size are modulated by nutrients in budding yeast

2016 ◽  
Author(s):  
Ricardo M. Leitao ◽  
Douglas R. Kellogg
Keyword(s):  
Cell Cycle ◽  
Cell Size ◽  
Daughter Cell ◽  
Slow Growth ◽  
Budding Yeast ◽  
Size Control ◽  
G1 Phase ◽  
Large Reduction ◽  
Duration Of Mitosis ◽  
Cell Cycle Entry

AbstractThe size of nearly all cells is modulated by nutrients. Thus, cells growing in poor nutrients can be nearly half the size of cells in rich nutrients. In budding yeast, cell size is thought to be controlled almost entirely by a mechanism that delays cell cycle entry until sufficient growth has occurred in G1 phase. Here, we show that most growth of a new daughter cell occurs in mitosis. When the rate of growth is slowed by poor nutrients, the duration of mitosis is increased, which suggests that cells compensate for slow growth in mitosis by increasing the duration of growth. The amount of growth required to complete mitosis is reduced in poor nutrients, leading to a large reduction in cell size. Together, these observations suggest that mechanisms that control the extent of growth in mitosis play a major role in cell size control in budding yeast.

scholarly journals EGR1 dysregulation defines an inflammatory and leukemic program in cell trajectory of human-aged hematopoietic stem cells (HSC)

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Christophe Desterke ◽  
Annelise Bennaceur-Griscelli ◽  
Ali G. Turhan
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Single Cell ◽  
Cell Cycle Phase ◽  
Cycle Phase ◽  
Cell Compartment ◽  
Hematopoietic Stem ◽  
Phase Progression ◽  
Cell Trajectory

Abstract Background During aging, hematopoietic stem cells (HSC) lose progressively both their self-renewal and differentiation potential. The precise molecular mechanisms of this phenomenon are not well established. To uncover the molecular events underlying this event, we have performed a bioinformatics analysis of 650 single-cell transcriptomes. Methods Single-cell transcriptome analyses of expression heterogeneity, cell cycle, and cell trajectory in human cell compartment enriched in hematopoietic stem cell compartment were investigated in the bone marrow according to the age of the donors. Identification of aging-related nodules was identified by weighted correlation network analysis in this primitive compartment. Results The analysis of single-cell transcriptomes allowed to uncover a major upregulation of EGR1 in human-aged lineage−CD34+CD38− cells which present cell cycle dysregulation with reduction of G2/M phase according to less expression of CCND2 during S phase. EGR1 upregulation in aging hematopoietic stem cells was found to be independent of cell cycle phases and gender. EGR1 expression trajectory in aged HSC highlighted a signature enriched in hematopoietic and immune disorders with the best induction of AP-1 complex and quiescence regulators such as EGR1, BTG2, JUNB, and NR41A. Sonic Hedgehog-related TMEM107 transmembrane molecule followed also EGR1 cell trajectory. EGR1-dependent gene weighted network analysis in human HSC-associated IER2 target protein-specific regulators of PP2A activity, IL1B, TNFSF10 ligands, and CD69, SELP membrane molecules in old HSC module with immune and leukemogenic signature. In contrast, for young HSC which were found with different cell cycle phase progression, its specific module highlighted upregulation of HIF1A hypoxic factor, PDE4B immune marker, DRAK2 (STK17B) T cell apoptosis regulator, and MYADM myeloid-associated marker. Conclusion EGR1 was found to be connected to the aging of human HSC and highlighted a specific cell trajectory contributing to the dysregulation of an inflammatory and leukemia-related transcriptional program in aged human HSCs. EGR1 and its program were found to be connected to the aging of human HSC with dissociation of quiescence property and cell cycle phase progression in this primitive hematopoietic compartment.

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