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Reduced HRAS G12V-Driven Tumorigenesis of Cell Lines Expressing KRAS C118S
Lu Huang1, Christopher M Counter2
1Department of Pharmacology & Cancer Biology, Duke University Medical Center, Durham, North Carolina, United States of America.
Abstract:
In many different human cancers, one of the HRAS, NRAS, or KRAS genes in the RAS family of small GTPases acquires an oncogenic mutation that renders the encoded protein constitutively GTP-bound and thereby active, which is well established to promote tumorigenesis. In addition to oncogenic mutations, accumulating evidence suggests that the wild-type isoforms may also be activated and contribute to oncogenic RAS-driven tumorigenesis. In this regard, redox-dependent reactions with cysteine 118 (C118) have been found to promote activation of wild-type HRAS and NRAS. We sought to determine if this residue is also important for the activation of wild-type KRAS and promotion of tumorigenesis. Thus, we mutated C118 to serine (C118S) in wild-type KRAS to block redox-dependent reactions at this site. We now report that this mutation reduced the level of GTP-bound KRAS and impaired RAS signaling stimulated by the growth factor EGF. With regards to tumorigenesis, we also report that oncogenic HRAS-transformed human cells in which endogenous KRAS was knocked down and replaced with KRASC118S exhibited reduced xenograft tumor growth, as did oncogenic HRAS-transformed KrasC118S/C118S murine cells in which the C118S mutation was knocked into the endogenous Kras gene. Taken together, these data suggest a role for redox-dependent activation of wild-type KRAS through C118 in oncogenic HRAS-driven tumorigenesis.
Insights
Wild-type KRAS, like HRAS and NRAS, can be activated by redox reactions at cysteine 118, promoting cancer. Blocking this cysteine 118 site in KRAS reduced tumor growth in preclinical models.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- RAS proteins (HRAS, NRAS, KRAS) are key regulators of cell signaling.
- Oncogenic mutations in RAS genes drive tumorigenesis by locking RAS proteins in an active state.
- Wild-type RAS isoforms may also contribute to cancer development through alternative activation mechanisms.
Purpose of the Study:
- To investigate the role of cysteine 118 (C118) in the activation of wild-type KRAS.
- To determine if redox-dependent activation of wild-type KRAS at C118 contributes to tumorigenesis.
Main Methods:
- Site-directed mutagenesis was used to create a KRAS C118S mutant to block redox reactions.
- The impact of the KRAS C118S mutation on GTP-bound KRAS levels and RAS signaling was assessed.
- Tumorigenesis was evaluated in human and murine cancer models with KRAS C118S expression.
Main Results:
- The KRAS C118S mutation reduced GTP-bound KRAS levels and impaired epidermal growth factor (EGF)-stimulated RAS signaling.
- Human cancer cells expressing KRAS C118S showed reduced xenograft tumor growth.
- Murine cancer cells with the endogenous Kras C118S mutation also exhibited decreased tumor growth.
Conclusions:
- Redox-dependent activation of wild-type KRAS through C118 plays a role in oncogenic HRAS-driven tumorigenesis.
- Cysteine 118 is a critical residue for KRAS activation and its contribution to cancer.
- Targeting C118-mediated KRAS activation may offer a therapeutic strategy for RAS-driven cancers.
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