Related Experiment Video

Updated: Jul 5, 2026

Characterize Disease-related Mutants of RAF Family Kinases by Using a Set of Practical and Feasible Methods
07:49

Characterize Disease-related Mutants of RAF Family Kinases by Using a Set of Practical and Feasible Methods

Published on: July 17, 2019

A novel role for Cdk1/cyclin B in regulating B-raf activation at mitosis

Sergiy I Borysov1, Thomas M Guadagno

  • 1Molecular Oncology Program, H. Lee Moffitt Cancer and Research Institute, Tampa, FL 33612, USA.

Molecular Biology of the Cell
|April 25, 2008
PubMed

Insights

Mitotic activation of B-Raf, a key MAPK cascade activator, is mediated by Cdk1/cyclin B phosphorylation at Serine-144 in Xenopus egg extracts. This phosphorylation is crucial for B-Raf activation during mitosis.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Mitosis requires Mitogen-Activated Protein Kinase (MAPK) activity for spindle assembly and checkpoint control.
  • B-Raf is a critical activator of the MAPK cascade during mitosis, with its activation regulated in an M-phase-dependent manner.
  • The precise mechanism for B-Raf activation during mitosis remains unclear, particularly how it differs from Ras-dependent activation.

Purpose of the Study:

  • To elucidate the mechanism of B-Raf activation during mitosis.
  • To investigate the role of Cdk1/cyclin B in mitotic B-Raf activation.
  • To identify key phosphorylation sites regulating B-Raf activity in mitosis.

Main Methods:

  • Utilized Xenopus egg extracts to study mitotic processes.
  • Employed co-immunoprecipitation to detect protein-protein interactions.
  • Performed mutagenesis and in vitro kinase assays to assess phosphorylation events and their functional consequences.

Main Results:

  • Demonstrated that Cdk1/cyclin B complexes associate with B-Raf during mitosis in Xenopus egg extracts.
  • Showed that Cdk1/cyclin B directly phosphorylates B-Raf at Serine-144 (S144), a novel site distinct from Ras-dependent phosphorylation sites.
  • Confirmed that phosphorylation of S144 is essential for B-Raf activation and subsequent MAPK cascade activation during mitosis.
  • Observed that a phospho-mimicking mutation at S144 did not lead to constitutive B-Raf activity, suggesting additional regulatory mechanisms.

Conclusions:

  • Revealed a novel cell cycle-dependent mechanism for activating the B-Raf/MEK/MAPK cascade.
  • Identified Cdk1/cyclin B as a direct upstream regulator of B-Raf during mitosis via phosphorylation at S144.
  • Highlighted that while S144 phosphorylation is necessary, other regulatory events likely contribute to full B-Raf activation in mitosis.

More Related Videos

Light-mediated Reversible Modulation of the Mitogen-activated Protein Kinase Pathway during Cell Differentiation and Xenopus Embryonic Development09:32

Light-mediated Reversible Modulation of the Mitogen-activated Protein Kinase Pathway during Cell Differentiation and Xenopus Embryonic Development

Published on: June 15, 2017

Experimental Approaches to Study Mitochondrial Localization and Function of a Nuclear Cell Cycle Kinase, Cdk113:15

Experimental Approaches to Study Mitochondrial Localization and Function of a Nuclear Cell Cycle Kinase, Cdk1

Published on: February 25, 2016

Related Concept Videos

Positive Regulator Molecules02:39

Positive Regulator Molecules

Mitotic cell division results in daughter cells that exactly resemble the parent cell. However, errors in the DNA replication or distribution of genetic material may lead to genetic mutations that may be passed down to every new cell formed from the resulting abnormal cell. Propagation of such mutant cells is restricted through checkpoint mechanisms present at different stages of the cell cycle. These checkpoints involve regulator molecules that either promote or demote cell cycle events.
Positive Regulator Molecules01:45

Positive Regulator Molecules

To consistently produce healthy cells, the cell cycle—the process that generates daughter cells—must be precisely regulated.
M-Cdk Drives Transition Into Mitosis02:15

M-Cdk Drives Transition Into Mitosis

Checkpoints throughout the cell cycle serve as safeguards and gatekeepers, allowing the cell cycle to progress in favorable conditions and slow or halt it in problematic ones. This regulation is known as the cell cycle control system.
Cyclin-dependent kinases, or Cdks, work in concert with cyclins to control cell cycle transitions. M-Cdk, a complex of Cdk1 bound to M cyclin, is a well-known example of this coordinated control that drives the transition from the G2 to the M phase.
M cyclin...
M-Cdk Drives Transition Into Mitosis02:15

M-Cdk Drives Transition Into Mitosis

Checkpoints throughout the cell cycle serve as safeguards and gatekeepers, allowing the cell cycle to progress in favorable conditions and slow or halt it in problematic ones. This regulation is known as the cell cycle control system.
Cyclin-dependent kinases, or Cdks, work in concert with cyclins to control cell cycle transitions. M-Cdk, a complex of Cdk1 bound to M cyclin, is a well-known example of this coordinated control that drives the transition from the G2 to the M phase.
M cyclin...
Inhibition of Cdk Activity02:34

Inhibition of Cdk Activity

The orderly progression of the cell cycle depends on the activation of Cdk protein by binding to its cyclin partner. However, the cell cycle must be restricted when undergoing abnormal changes. Most cancers correlate to the deregulated cell cycle, and since Cdks are a central component of the cell cycle, Cdk inhibitors are extensively studied to develop anticancer agents. For instance, cyclin D associates with several Cdks, such as Cdk 4/6, to form an active complex. The cyclin D-Cdk4/6 complex...
Inhibition of CDK Activity02:34

Inhibition of CDK Activity

The orderly progression of the cell cycle depends on the activation of Cdk protein by binding to its cyclin partner. However, the cell cycle must be restricted when undergoing abnormal changes. Most cancers correlate to the deregulated cell cycle, and since Cdks are a central component of the cell cycle, Cdk inhibitors are extensively studied to develop anticancer agents. For instance, cyclin D associates with several Cdks, such as Cdk 4/6, to form an active complex. The cyclin D-Cdk4/6 complex...