Abstract
Oncogenic mutations in KRAS result in a constitutively active, GTP-bound form that in turn activates many proliferative pathways. However, because of its compact and simple architecture, directly targeting KRAS with small molecule drugs has been challenging. Another approach is to identify targetable proteins that interact with KRAS. Argonaute 2 (AGO2) was recently identified as a protein that facilitates RAS-driven oncogenesis. Whereas previous studies described the in vivo effect of AGO2 on cancer progression in cells harboring mutated KRAS, here we sought to examine their direct interaction using purified proteins. We show that full length AGO2 co-immunoprecipitates with KRAS using purified components, however, a complex between FL AGO2 and KRAS could not be isolated. We also generated a smaller N-terminal fragment of AGO2 (NtAGO2) which is believed to represent the primary binding site of KRAS. A complex with NtAGO2 could be detected via ion-mobility mass spectrometry and size exclusion chromatography. However, the data suggest that the interaction of KRAS with purified AGO2 (NtAGO2 or FL AGO2) is weak and likely requires additional cellular components or proteo-forms of AGO2 that are not readily available in our purified assay systems. Future studies are needed to determine what conformation or modifications of AGO2 are necessary to enrich KRAS association and regulate its activities.
Original language | English (US) |
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Article number | 101191 |
Journal | Biochemistry and Biophysics Reports |
Volume | 29 |
DOIs | |
State | Published - Mar 2022 |
Externally published | Yes |
Bibliographical note
Funding Information:This study was supported by National Institutes of Health (NIH) grants CA046592 , GM007863 , and GM007315 and a University of Michigan Rackham Merit Fellowship to JJW, a NIH Pharmacological Sciences Training Program fellowship GM007767 and U.S. Department of Education GAANN fellowship P200A150164 to TSB, NIH grants HL071818 , HL122416 , and CA221289 to JJGT, NIH grant CA221011 to RFS, and the Howard Hughes Medical Institute (HHMI) to AMC. Research in the Ruotolo group was supported by the Agilent Technologies Thought Leader Award and University Relations grant programs.
Publisher Copyright:
© 2021 The Authors
PubMed: MeSH publication types
- Journal Article