scholarly journals The SH3-domain protein Bem1 coordinates mitogen-activated protein kinase cascade activation with cell cycle control in Saccharomyces cerevisiae.

1996 ◽  
Vol 16 (8) ◽  
pp. 4095-4106 ◽  
Author(s):  
D M Lyons ◽  
S K Mahanty ◽  
K Y Choi ◽  
M Manandhar ◽  
E A Elion
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Transcriptional Activation ◽  
Mapk Cascade ◽  
Mitogen Activated Protein Kinase ◽  
G1 Arrest ◽  
Polarized Growth ◽  
Cortical Actin ◽  
Mapk Kinase ◽  
Mitogen Activated Protein

The mating mitogen-activated protein kinase (MAPK) cascade has three major outputs prior to fusion: transcriptional activation of many genes, cell cycle arrest in the G1 phase, and polarized growth. Bem1 localizes near the cortical actin cytoskeleton and is essential for polarized growth during mating. Here we show that Bem1 is required for efficient signal transduction and coordinates MAPK cascade activation with G1 arrest and mating. bem1delta null mutants are defective in G1 arrest and transcriptional activation in response to mating pheromone. Bem1 protein stimulates Fus3 (MAPK) activity and associates with Ste5, the tethering protein essential for activation of the MAPK kinase kinase Ste11. Bem1-Ste5 complexes also contain Ste11, Ste7 (MAPK kinase), and Fus3, suggesting that Ste5 localizes the MAPK cascade to Bem1. Strikingly, Bem1 also copurifies with Far1, a Fus3 substrate required for G1 arrest and proper polarized growth during mating. These and other results suggest that Bem1 may cross-link the Ste5-MAPK cascade complex to upstream activators and specific downstream substrates at the shmoo tip, thus enabling efficient circuitry for G1 arrest and mating.

Phosphorylation of mitogen-activated protein kinase cascade during early embryo development in the mouse

2000 ◽  
Vol 12 (4) ◽  
pp. 209 ◽  
Author(s):  
Naoki Iwamori ◽  
Kunihiko Naito ◽  
Koji Sugiura ◽  
Hideyuki Kagii ◽  
Masakane Yamashita ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Embryo Development ◽  
Somatic Cells ◽  
Mouse Embryos ◽  
Mapk Cascade ◽  
Mitogen Activated Protein Kinase ◽  
Cell Stage ◽  
Mitogen Activated Protein ◽  
Early Mouse

The mitogen-activated protein kinase (MAPK) cascade is one of the most important signal transduction pathways that regulate the cell cycle in somatic cells. The present study examined the phosphorylation states of components in the MAPK cascade, Raf-1, MEK-1, and extracellular signal regulated kinases (ERKs), which are activated by mitogens, throughout early mouse embryo development and in cultured somatic cells generally. In somatic cells, Raf-1 and MEK-1 were phosphorylated at M-phase and dephosphorylated during interphase. ERKs were not phosphorylated at any stage during the cell cycle. These results were similar to previous findings for the first and second cell cycles of early mouse embryos. In contrast, after the four-cell stage, not only ERKs, but also Raf-1 and MEK-1, were not phosphorylated at any stage during the cell cycle in mouse early embryos. These results suggest that the MAPK cascade in mouse embryos is regulated by the same mechanism as in somatic cells before the two-cell stage, and that regulation is changed to an embryo-specific mechanism after the four-cell stage.

scholarly journals Cell Cycle Arrest and Reversion of Ras-Induced Transformation by a Conditionally Activated Form of Mitogen-Activated Protein Kinase Kinase Kinase 3

1999 ◽  
Vol 19 (5) ◽  
pp. 3857-3868 ◽  
Author(s):  
Heidrun Ellinger-Ziegelbauer ◽  
Kathleen Kelly ◽  
Ulrich Siebenlist
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Cyclin D1 ◽  
Stress Responses ◽  
Cell Cycle Progression ◽  
Cellular Transformation ◽  
Mitogen Activated Protein Kinase ◽  
Cycle Progression ◽  
Mapk Kinase ◽  
Mitogen Activated Protein

ABSTRACT Signal-induced proliferation, differentiation, or stress responses of cells depend on mitogen-activated protein kinase (MAPK) cascades, the core modules of which consist of members of three successively acting kinase families (MAPK kinase kinase [MAP3K], MAPK kinase, and MAPK). It is demonstrated here that the MEKK3 kinase inhibits cell proliferation, a biologic response not commonly associated with members of the MAP3K family of kinases. A conditionally activated form of MEKK3 stably expressed in fibroblasts arrests these cells in early G1. MEKK3 critically blocks mitogen-driven expression of cyclin D1, a cyclin which is essential for progression of fibroblasts through G1. The MEKK3-induced block of cyclin D1 expression and of cell cycle progression may be mediated via p38 MAPK, a downstream effector of MEKK3. The MEKK3-mediated block of proliferation also reverses Ras-induced cellular transformation, suggesting possible tumor-suppressing functions for this kinase. Together, these results suggest an involvement of the MEKK3 kinase in negative regulation of cell cycle progression, and they provide the first insights into biologic activities of this kinase.

scholarly journals Mutational Analysis Suggests That Activation of the Yeast Pheromone Response Mitogen-activated Protein Kinase Pathway Involves Conformational Changes in the Ste5 Scaffold Protein

2000 ◽  
Vol 11 (11) ◽  
pp. 4033-4049 ◽  
Author(s):  
Claudio Sette ◽  
Carla J. Inouye ◽  
Shannon L. Stroschein ◽  
Phillip J. Iaquinta ◽  
Jeremy Thorner
Keyword(s):  
Protein Kinase ◽  
Conformational Changes ◽  
Transcriptional Activation ◽  
Mutational Analysis ◽  
Morphological Changes ◽  
Mapk Cascade ◽  
Mitogen Activated Protein Kinase ◽  
Pheromone Response ◽  
Activated State ◽  
Mitogen Activated Protein

Ste5 is essential for pheromone response and binds components of a mitogen-activated protein kinase (MAPK) cascade: Ste11 (MEKK), Ste7 (MEK), and Fus3 (MAPK). Pheromone stimulation releases Gβγ (Ste4-Ste18), which recruits Ste5 and Ste20 (p21-activated kinase) to the plasma membrane, activating the MAPK cascade. A RING-H2 domain in Ste5 (residues 177–229) negatively regulates Ste5 function and mediates its interaction with Gβγ. Ste5(C177A C180A), carrying a mutated RING-H2 domain, cannot complement a ste5Δ mutation, yet supports mating even in ste4Δ ste5Δ cells when artificially dimerized by fusion to glutathioneS-transferase (GST). In contrast, wild-type Ste5 fused to GST permits mating of ste5Δ cells, but does not allow mating of ste4Δ ste5Δ cells. This differential behavior provided the basis of a genetic selection for STE5 gain-of-function mutations. MATaste4Δ ste5Δ cells expressing Ste5-GST were mutagenized chemically and plasmids conferring the capacity to mate were selected. Three independent single-substitution mutations were isolated. These constitutive STE5 alleles induce cell cycle arrest, transcriptional activation, and morphological changes normally triggered by pheromone, even when Gβγ is absent. The first, Ste5(C226Y), alters the seventh conserved position in the RING-H2 motif, confirming that perturbation of this domain constitutively activates Ste5 function. The second, Ste5(P44L), lies upstream of a basic segment, whereas the third, Ste5(S770K), is situated within an acidic segment in a region that contacts Ste7. None of the mutations increased the affinity of Ste5 for Ste11, Ste7, or Fus3. However, the positions of these novel-activating mutations suggested that, in normal Ste5, the N terminus may interact with the C terminus. Indeed, in vitro, GST-Ste5(1-518) was able to associate specifically with radiolabeled Ste5(520-917). Furthermore, both the P44L and S770K mutations enhanced binding of full-length Ste5 to GST-Ste5(1-518), whereas they did not affect Ste5 dimerization. Thus, binding of Gβγ to the RING-H2 domain may induce a conformational change that promotes association of the N- and C-terminal ends of Ste5, stimulating activation of the MAPK cascade by optimizing orientation of the bound kinases and/or by increasing their accessibility to Ste20-dependent phosphorylation (or both). In accord with this model, the novel Ste5 mutants copurified with Ste7 and Fus3 in their activated state and their activation required Ste20.

scholarly journals Cell-cycle arrest by PD184352 requires inhibition of extracellular signal-regulated kinases (ERK) 1/2 but not ERK5/BMK1

2002 ◽  
Vol 366 (2) ◽  
pp. 673-680 ◽  
Author(s):  
Matthew S. SQUIRES ◽  
Paula M. NIXON ◽  
Simon J. COOK
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Cyclin D1 ◽  
Chinese Hamster ◽  
Mitogen Activated Protein Kinase ◽  
G1 Arrest ◽  
Low Doses ◽  
Proliferative Effect ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

Serum and growth factors activate both the canonical extracellular signal-regulated kinase (ERK) 1/2 pathway and the ERK5/big mitogen-activated protein kinase 1 (BMK) 1 pathway. Pharmacological inhibition of the ERK1/2 pathway using PD98059 and U0126 prevents cyclin D1 expression and inhibits cell proliferation, arguing that the ERK1/2 pathway is rate limiting for cell-cycle re-entry. However, both PD98059 and U0126 also inhibit the ERK5/BMK1 pathway, raising the possibility that the anti-proliferative effect of such drugs may be due to inhibition of ERK5 or both pathways. Here we characterize the effect of the novel mitogen-activated protein kinase/ERK kinase (MEK) inhibitor, PD184352, on the ERK1/2 and ERK5 pathways in the Chinese hamster fibroblast cell line CCl39. In quiescent cells, serum-stimulated ERK1 activity was completely inhibited by PD184352 with an IC50 below 1μM, whereas ERK5 activation was unaffected even at 20μM. Serum-stimulated DNA synthesis and cyclin D1 expression was inhibited by low doses of PD184352, which abolished ERK1 activity but had no effect on ERK5. Similarly, in cycling cells PD184352 caused a dose-dependent G1 arrest and inhibition of cyclin D1 expression at low doses, which inhibited ERK1 but were without effect on ERK5. These results indicate that the anti-proliferative effect of PD184352 is due to inhibition of the classical ERK1/2 pathway and does not require inhibition of the ERK5 pathway.

Endothelins stimulate mitogen-activated protein kinase cascade through either ETA or ETB

1994 ◽  
Vol 267 (4) ◽  
pp. C1130-C1135 ◽  
Author(s):  
Y. Wang ◽  
P. M. Rose ◽  
M. L. Webb ◽  
M. J. Dunn
Keyword(s):  
Protein Kinase ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Mapk Cascade ◽  
Mitogen Activated Protein Kinase ◽  
Thymidine Uptake ◽  
Mapk Activation ◽  
Transfected Cells ◽  
Gel Retardation ◽  
Mitogen Activated Protein

Endothelin (ET) has been shown to activate mitogen-activated protein kinase (MAPK). However, it has been unclear which of the ET receptors is coupled to MAPK activation. In the present study, we conducted experiments to determine which ET receptor is linked to MAPK activation. We found that both human ETA and ETB were coupled to the MAPK cascade in ETA or ETB cDNA-transfected Chinese hamster ovary cells. ET-1 was more potent than ET-3 in the activation of p42 MAPK, induction of MAPK kinase (MAPKK) gel retardation and uptake of [3H]thymidine in ETA-transfected cells, whereas sarafotoxin (S6c) showed no stimulatory effect on the kinases and [3H]thymidine uptake. ET-1, ET-3, and S6c had approximately the same potency to activate p42 MAPK, MAPKK gel retardation, and [3H]thymidine uptake in ETB-transfected cells. These data suggest that 1) ET isopeptides, through either ETA or ETB receptors, induce the MAPK cascade as well as cell proliferation; and 2) the different potencies of ET isopeptides for activation of the MAPK cascade and induction of cell growth are mainly due to their different affinities toward ETA and ETB.

scholarly journals Mitogen-Activated Protein Kinase Hog1 Mediates Adaptation to G1 Checkpoint Arrest during Arsenite and Hyperosmotic Stress

2008 ◽  
Vol 7 (8) ◽  
pp. 1309-1317 ◽  
Author(s):  
Iwona Migdal ◽  
Yulia Ilina ◽  
Markus J. Tamás ◽  
Robert Wysocki
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Cell Cycle Arrest ◽  
Kinase Inhibitor ◽  
Hyperosmotic Stress ◽  
Mitogen Activated Protein Kinase ◽  
Hog Pathway ◽  
Cycle Arrest ◽  
Mitogen Activated Protein

ABSTRACT Cells slow down cell cycle progression in order to adapt to unfavorable stress conditions. Yeast (Saccharomyces cerevisiae) responds to osmotic stress by triggering G1 and G2 checkpoint delays that are dependent on the mitogen-activated protein kinase (MAPK) Hog1. The high-osmolarity glycerol (HOG) pathway is also activated by arsenite, and the hog1Δ mutant is highly sensitive to arsenite, partly due to increased arsenite influx into hog1Δ cells. Yeast cell cycle regulation in response to arsenite and the role of Hog1 in this process have not yet been analyzed. Here, we found that long-term exposure to arsenite led to transient G1 and G2 delays in wild-type cells, whereas cells that lack the HOG1 gene or are defective in Hog1 kinase activity displayed persistent G1 cell cycle arrest. Elevated levels of intracellular arsenite and “cross talk” between the HOG and pheromone response pathways, observed in arsenite-treated hog1Δ cells, prolonged the G1 delay but did not cause a persistent G1 arrest. In contrast, deletion of the SIC1 gene encoding a cyclin-dependent kinase inhibitor fully suppressed the observed block of G1 exit in hog1Δ cells. Moreover, the Sic1 protein was stabilized in arsenite-treated hog1Δ cells. Interestingly, Sic1-dependent persistent G1 arrest was also observed in hog1Δ cells during hyperosmotic stress. Taken together, our data point to an important role of the Hog1 kinase in adaptation to stress-induced G1 cell cycle arrest.

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