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Related Concept Videos

Small GTPases - Ras and Rho01:24

Small GTPases - Ras and Rho

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Ras and Rho are small monomeric GTPases that act downstream of receptor tyrosine kinase (RTK) and regulate various cellular processes. These GTPases switch between active and inactive states by binding to guanine nucleotides.
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GTPases and their Regulation02:14

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Guanine nucleotide-binding proteins (G-proteins), also known as GTPases, are a superfamily of proteins that regulate many cellular processes, such as cell signaling, vesicular transport, and the regulation of cell shape and motility. Mutation or dysfunction of these proteins can lead to disease. There are around 40,000 known G-proteins that can broadly be classified into two groups ‒  small G-proteins consisting of a single domain and large multi-domain G-proteins.
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The Ras Gene02:38

The Ras Gene

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The Ras-gene-encoded proteins are regulators of signaling pathways controlling cell proliferation, differentiation, or cell survival. The Ras-gene family in humans constitutes three primary members—the HRas, NRas, and KRas. These genes code for four functionally distinct yet closely related proteins—the HRas, NRas, KRas4A, and KRas4B. The involvement of mutant Ras genes in human cancer was first discovered in 1982 and is among the most common causes of human tumorigenesis.
Ras is a...
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Rab Proteins01:14

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Rab proteins constitute the largest family of monomeric GTPases, of which 70 members are present in humans. Rab proteins and their effectors regulate consecutive stages of vesicle transport such as vesicle transport, docking, and fusion to the correct recipient membrane.
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Cell Polarization by Rho Proteins01:21

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Cell polarity is the asymmetric distribution of cellular and membrane components, making one side of the cell different from the other. This polarity is essential to many processes such as embryogenesis, axon migration, glucose transport across epithelial cells, and directional cell migration. A migrating cell responds to intracellular or extracellular signals via molecular cascades that reorganize the actin cytoskeleton to establish this polarity. In these cells, the Rho family proteins Cdc42,...
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Activation and Inactivation of G Proteins01:22

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Heterotrimeric G proteins are guanine nucleotide-binding proteins. As the name suggests, heterotrimeric G proteins are composed of three subunits: alpha, beta, and gamma. They remain GDP-bound or GTP-bound inside the cells and switch between inactive/active states. The Gα subunit possesses the nucleotide-binding pocket that binds guanine nucleotides and switches between GDP or GTP-bound states. In contrast, the Gꞵ and Gγ subunits are always bound together with high...
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Affinity Precipitation of Active Rho-GEFs Using a GST-tagged Mutant Rho Protein GST-RhoAG17A from Epithelial Cell Lysates
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Tumor-derived RHOA mutants interact with effectors in the GDP-bound state.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Structural Biology

Background:

  • Ras homolog family member A (RHOA) mutations are implicated in various cancers.
  • Specific RHOA mutations, A161P and A161V, are found in adult T-cell leukemia/lymphoma.

Purpose of the Study:

  • To investigate the molecular mechanisms of two gain-of-function RHOA mutations, A161P and A161V.
  • To understand how these mutations contribute to tumorigenesis in adult T-cell leukemia/lymphoma.

Main Methods:

  • Biochemical assays to measure guanine nucleotide exchange and GTP hydrolysis rates.
  • X-ray crystallography to determine the structures of RHOA mutants.
  • Nuclear Magnetic Resonance (NMR) spectroscopy (31P and 1H-15N HSQC) to study protein dynamics.
  • Molecular dynamics simulations to analyze conformational changes.

Main Results:

  • RHOA(A161P) and RHOA(A161V) exhibit fast cycling rates and reduced GTPase activity compared to wild-type RHOA.
  • Crystal structures reveal altered nucleotide binding in RHOA(A161P) and an open nucleotide pocket in RHOA(A161V).
  • Mutations destabilize RHOA switch regions, favoring an active conformation and enabling effector interaction in the GDP-bound state.

Conclusions:

  • The A161P and A161V mutations confer gain-of-function properties to RHOA by altering its nucleotide binding and dynamics.
  • These mutations promote constitutive RHOA activity, potentially driving tumorigenesis through aberrant effector interactions.