Roles of palmitoylation and the KIKK membrane-targeting motif in leukemogenesis by oncogenic KRAS4A

Abstract

BackgroundWe have previously shown that palmitoylation is essential for NRAS leukemogenesis, suggesting that targeting RAS palmitoylation may be an effective therapy for NRAS-related cancers. For KRAS-driven cancer, although much research has been focused on the KRAS4B splice variant, which does not undergo palmitoylation, KRAS4A has recently been shown to play an essential role in the development of carcinogen-induced lung cancer in mice and to be widely expressed in human cancers. However, the role of palmitoylation in KRAS4A tumorigenesis is not clear.MethodsThe expression of KRAS4A in KRAS-mutated leukemia cell lines and acute myeloid leukemia (AML) cells were checked using western blotting and reverse transcriptions-quantitative polymerase chain reaction (RT-qPCR) analysis, respectively. The leukemogenic potentials of oncogenic KRAS4A and its palmitoylation-defective mutants were examined by a mouse bone marrow transduction and transplantation model and the in vitro transformation assays. The activation of the RAS downstream signaling pathways and the membrane localizations of the KRAS4A and its mutants were analyzed via western blot analysis and confocal microscopy, respectively.ResultsWe show here that KRAS4A is expressed in human leukemia cell lines and in AML cells harboring KRAS mutations and that mutation at the palmitoylation site of oncogenic KRAS4A significantly abrogates its leukemogenic potential. However, unlike NRAS, palmitoylation-defective KRAS4A still induces leukemia in mice, albeit with a much longer latency. Using NRAS/KRAS4A chimeric constructs, we found that the KIKK motif of KRAS4A contributes to the transforming activity of KRAS4A. Mutations at both palmitoylation site and the KIKK motif abolish the ability of oncogenic KRAS4A to induce leukemia in mice.ConclusionsOur studies suggest that therapies targeting RAS palmitoylation may also be effective in treating KRAS4A associated malignancies and that interfering the KIKK membrane-targeting motif would enhance the therapeutic effectiveness.

Posttranslational Modifications of RAS Proteins

“…A similar approach to investigate palmitoylation of KRAS4A G12D in leukemogenesis revealed that even in the absence of palmitoylation, the polybasic region cooperates sufficiently with prenylation to promote disease, albeit with a notable delay in disease onset (82). …”

SIRT2 and lysine fatty acylation regulate the transforming activity of K-Ras4a

“…K-Ras4b has attracted most of the attention because it was assumed to be the more abundant and thus the more important K-Ras isoform mutated in human cancers. However, recent studies have revealed that K-Ras4a is widely expressed in many cancer cell lines and its level is similar to that of K-Ras4b in human colorectal tumors ( Tsai et al, 2015 ; Zhao et al, 2015 ). A requirement for oncogenic K-Ras4a in lung carcinogenesis has also been demonstrated in mice ( To et al, 2008 ).…”

N-Arachidonoyl Dopamine Inhibits NRAS Neoplastic Transformation by Suppressing Its Plasma Membrane Translocation

“…NRAS, HRAS, and KRAS4A are further palmitoylated on the Golgi, increasing their affinity to bind the plasma membrane (16,17). We have shown previously that interrupting the plasma membrane localization of NRAS and KRAS4A could significantly abrogate their leukemogenic potential (18)(19)(20). However, the exact mechanism of RAS proteins translocating from endomembranes to the plasma membrane is still elusive (17).…”