Tyrosyl phosphorylation of KRAS stalls GTPase cycle via alteration of switch I and II conformation

The DNA replication checkpoint couples the onset of mitosis with the completion of S phase. It is clear that in the fission yeast Schizosaccharomyces pombe, operation of this checkpoint requires maintenance of the inhibitory tyrosyl phosphorylation of Cdc2. Cdc25 phosphatase induces mitosis by dephosphorylating tyrosine 15 of Cdc2. In this report, Cdc25 is shown to accumulate to a very high level in cells arrested in S. This shows that mechanisms which modulate the abundance of Cdc25 are unconnected to the DNA replication checkpoint. Using a Cdc2/cyclin B activation assay, we found that Cdc25 activity increased approximately 10-fold during transit through M phase. Cdc25 was activated by phosphorylations that were dependent on Cdc2 activity in vivo. Cdc25 activation was suppressed in cells arrested in G1 and S. However, Cdc25 was more highly modified and appeared to be somewhat more active in S than in G1. This finding might be connected to the fact that progression from G1 to S increases the likelihood that constitutive Cdc25 overproduction will cause inappropriate mitosis.

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Septin ring assembly involves cycles of GTP loading and hydrolysis by Cdc42p

The Journal of Cell Biology ◽

10.1083/jcb.200109062 ◽

2002 ◽

Vol 156 (2) ◽

pp. 315-326 ◽

Cited By ~ 131

Author(s):

Amy S. Gladfelter ◽

Indrani Bose ◽

Trevin R. Zyla ◽

Elaine S.G. Bardes ◽

Daniel J. Lew

Keyword(s):

Transcriptional Activation ◽

Gtpase Activating Protein ◽

Gtp Hydrolysis ◽

Septin Ring ◽

Turn On ◽

Ring Assembly ◽

Bud Emergence ◽

Assembly Factor ◽

Gtpase Cycle ◽

Budding Yeast Cell Cycle

At the beginning of the budding yeast cell cycle, the GTPase Cdc42p promotes the assembly of a ring of septins at the site of future bud emergence. Here, we present an analysis of cdc42 mutants that display specific defects in septin organization, which identifies an important role for GTP hydrolysis by Cdc42p in the assembly of the septin ring. The mutants show defects in basal or stimulated GTP hydrolysis, and the septin misorganization is suppressed by overexpression of a Cdc42p GTPase-activating protein (GAP). Other mutants known to affect GTP hydrolysis by Cdc42p also caused septin misorganization, as did deletion of Cdc42p GAPs. In performing its roles in actin polarization and transcriptional activation, GTP-Cdc42p is thought to function by activating and/or recruiting effectors to the site of polarization. Excess accumulation of GTP-Cdc42p due to a defect in GTP hydrolysis by the septin-specific alleles might cause unphysiological activation of effectors, interfering with septin assembly. However, the recessive and dose-sensitive genetic behavior of the septin-specific cdc42 mutants is inconsistent with the septin defect stemming from a dominant interference of this type. Instead, we suggest that assembly of the septin ring involves repeated cycles of GTP loading and GTP hydrolysis by Cdc42p. These results suggest that a single GTPase, Cdc42p, can act either as a ras-like GTP-dependent “switch” to turn on effectors or as an EF-Tu–like “assembly factor” using the GTPase cycle to assemble a macromolecular structure.

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MO14-2 KRAS Q61H mutation evades the regulation of tyrosyl phosphorylation and renders cancer cells resistant to SHP2 inhibitor

Annals of Oncology ◽

10.1016/j.annonc.2021.05.600 ◽

2021 ◽

Vol 32 ◽

pp. S306

Author(s):

Yoshihito Kano ◽

Teklab Gebregiworgis ◽

Christopher Marshall ◽

Mitsuhiko Ikura ◽

Satoshi Miyake ◽

...

Keyword(s):

Cancer Cells ◽

Tyrosyl Phosphorylation

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Receptor compaction and GTPase rearrangement drive SRP-mediated cotranslational protein translocation into the ER

Science Advances ◽

10.1126/sciadv.abg0942 ◽

2021 ◽

Vol 7 (21) ◽

pp. eabg0942

Author(s):

Jae Ho Lee ◽

Ahmad Jomaa ◽

SangYoon Chung ◽

Yu-Hsien Hwang Fu ◽

Ruilin Qian ◽

...

Keyword(s):

Single Molecule ◽

Protein Translocation ◽

Molecular Model ◽

Genetic Diseases ◽

Signal Recognition Particle ◽

Biological Regulation ◽

Single Molecule Studies ◽

Guanosine Triphosphatase ◽

Gtpase Cycle

The conserved signal recognition particle (SRP) cotranslationally delivers ~30% of the proteome to the eukaryotic endoplasmic reticulum (ER). The molecular mechanism by which eukaryotic SRP transitions from cargo recognition in the cytosol to protein translocation at the ER is not understood. Here, structural, biochemical, and single-molecule studies show that this transition requires multiple sequential conformational rearrangements in the targeting complex initiated by guanosine triphosphatase (GTPase)–driven compaction of the SRP receptor (SR). Disruption of these rearrangements, particularly in mutant SRP54G226E linked to severe congenital neutropenia, uncouples the SRP/SR GTPase cycle from protein translocation. Structures of targeting intermediates reveal the molecular basis of early SRP-SR recognition and emphasize the role of eukaryote-specific elements in regulating targeting. Our results provide a molecular model for the structural and functional transitions of SRP throughout the targeting cycle and show that these transitions provide important points for biological regulation that can be perturbed in genetic diseases.

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Stochastic simulation of the transducin GTPase cycle

Biophysical Journal ◽

10.1016/s0006-3495(96)79499-7 ◽

1996 ◽

Vol 71 (6) ◽

pp. 3051-3063 ◽

Cited By ~ 26

Author(s):

S. Felber ◽

H.P. Breuer ◽

F. Petruccione ◽

J. Honerkamp ◽

K.P. Hofmann

Keyword(s):

Stochastic Simulation ◽

Gtpase Cycle

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Deregulated expression of the RanBP1 gene alters cell cycle progression in murine fibroblasts

Journal of Cell Science ◽

10.1242/jcs.110.19.2345 ◽

1997 ◽

Vol 110 (19) ◽

pp. 2345-2357 ◽

Cited By ~ 1

Author(s):

A. Battistoni ◽

G. Guarguaglini ◽

F. Degrassi ◽

C. Pittoggi ◽

A. Palena ◽

...

Keyword(s):

Cell Cycle ◽

Dna Replication ◽

Cell Cycle Progression ◽

Protein Import ◽

Proliferating Cells ◽

Cycle Progression ◽

Ran Gtpase ◽

Genes Encoding ◽

Active Transcription ◽

Gtpase Cycle

RanBP1 is a molecular partner of the Ran GTPase, which is implicated in the control of several processes, including DNA replication, mitotic entry and exit, cell cycle progression, nuclear structure, protein import and RNA export. While most genes encoding Ran-interacting partners are constitutively active, transcription of the RanBP1 mRNA is repressed in non proliferating cells, is activated at the G1/S transition in cycling cells and peaks during S phase. We report here that forced expression of the RanBP1 gene disrupts the orderly execution of the cell division cycle at several stages, causing inhibition of DNA replication, defective mitotic exit and failure of chromatin decondensation during the telophase-to-interphase transition in cells that achieve nuclear duplication and chromosome segregation. These results suggest that deregulated RanBP1 activity interferes with the Ran GTPase cycle and prevents the functioning of the Ran signalling system during the cell cycle.

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Ran GTPase Cycle and Importins α and β Are Essential for Spindle Formation and Nuclear Envelope Assembly in LivingCaenorhabditis elegans Embryos

Molecular Biology of the Cell ◽

10.1091/mbc.e02-06-0346 ◽

2002 ◽

Vol 13 (12) ◽

pp. 4355-4370 ◽

Cited By ~ 127

Author(s):

Peter Askjaer ◽

Vincent Galy ◽

Eva Hannak ◽

Iain W. Mattaj

Keyword(s):

Nuclear Envelope ◽

Cell Function ◽

Small Gtpase ◽

Spindle Formation ◽

Ran Gtpase ◽

Early Embryos ◽

Nuclear Envelope Assembly ◽

Importin Α ◽

Gtpase Cycle

The small GTPase Ran has been found to play pivotal roles in several aspects of cell function. We have investigated the role of the Ran GTPase cycle in spindle formation and nuclear envelope assembly in dividing Caenorhabditis elegans embryos in real time. We found that Ran and its cofactors RanBP2, RanGAP, and RCC1 are all essential for reformation of the nuclear envelope after cell division. Reducing the expression of any of these components of the Ran GTPase cycle by RNAi leads to strong extranuclear clustering of integral nuclear envelope proteins and nucleoporins. Ran, RanBP2, and RanGAP are also required for building a mitotic spindle, whereas astral microtubules are normal in the absence of these proteins. RCC1(RNAi) embryos have similar abnormalities in the initial phase of spindle formation but eventually recover to form a bipolar spindle. Irregular chromatin structures and chromatin bridges due to spindle failure were frequently observed in embryos where the Ran cycle was perturbed. In addition, connection between the centrosomes and the male pronucleus, and thus centrosome positioning, depends upon the Ran cycle components. Finally, we have demonstrated that both IMA-2 and IMB-1, the homologues of vertebrate importin α and β, are essential for both spindle assembly and nuclear formation in early embryos.

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Membrane curvature and the control of GTP hydrolysis in Arf1 during COPI vesicle formation

Biochemical Society Transactions ◽

10.1042/bst0330619 ◽

2005 ◽

Vol 33 (4) ◽

pp. 619-622 ◽

Cited By ~ 24

Author(s):

B. Antonny ◽

J. Bigay ◽

J.-F. Casella ◽

G. Drin ◽

B. Mesmin ◽

...

Keyword(s):

Lipid Membranes ◽

Membrane Curvature ◽

Gtp Hydrolysis ◽

Membrane Fission ◽

Transport Vesicles ◽

Highly Sensitive ◽

Copi Vesicle ◽

Membrane Budding ◽

Gtpase Cycle ◽

Adp Ribosylation Factor

The GTP switch of the small G-protein Arf1 (ADP-ribosylation factor 1) on lipid membranes promotes the polymerization of the COPI (coat protein complex I) coat, which acts as a membrane deforming shell to form transport vesicles. Real-time measurements for coat assembly on liposomes gives insights into how the GTPase cycle of Arf1 is coupled in time with the polymerization of the COPI coat and the resulting membrane deformation. One key parameter seems to be the membrane curvature. Arf-GAP1 (where GAP stands for GTPase-activating protein), which promotes GTP hydrolysis in the Arf1–COPI complex is highly sensitive to lipid packing. Its activity on Arf1-GTP increases by two orders of magnitude as the diameter of the liposomes approaches that of authentic transport vesicles (60 nm). This suggests that during membrane budding, Arf1-GTP molecules are progressively eliminated from the coated area where the membrane curvature is positive, but are protected from Arf-GAP1 at the bud neck due to the negative curvature of this region. As a result, the coat should be stable as long as the bud remains attached and should disassemble as soon as membrane fission occurs.

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