Mapping the KRAS Proteoform Landscape in Colorectal Cancer Identifies Truncated KRAS4B that Decreases MAPK Signaling

Author:

Adams, Lauren M

Dehart,Caroline

Drown, Bryon S

Anderson, Lissa C

Bocik,William

Boja, Emily S

Hiltke, Tara M

Hendrickson, Christopher L

Rodriguez, Henry

Caldwell, Michael

Vafabakhsh, Reza

Kelleher, Neil L
Author Address

Department of Molecular Biosciences, Northwestern University, Evanston, IL., NCI RAS Initiative, Frederick National Laboratory for Cancer Research, Frederick, MD., Department of Chemistry, Northwestern University, Evanston, IL., Ion Cyclotron Resonance Program, National High Magnetic Field Laboratory, Tallahassee, FL., Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Frederick, MD., Office of Cancer Clinical Proteomics Research, National Cancer Institute, Bethesda MD., Northwestern University Feinberg School of Medicine, Chicago, IL; Chemistry of Life Processes Institute, Northwestern University, Evanston, IL; Proteomics Center of Excellence, Northwestern University, Evanston, IL., Department of Molecular Biosciences, Northwestern University, Evanston, IL; Department of Chemistry, Northwestern University, Evanston, IL; Northwestern University Feinberg School of Medicine, Chicago, IL; Chemistry of Life Processes Institute, Northwestern University, Evanston, IL; Proteomics Center of Excellence, Northwestern University, Evanston, IL. Electronic address: n-kelleher@northwestern.edu.,

Abstract:

The KRAS gene is one of the most frequently mutated oncogenes in human cancer and gives rise to two isoforms, KRAS4A and KRAS4B. KRAS post-translational modifications (PTMs) have the potential to influence downstream signaling. However, the relationship between KRAS PTMs and oncogenic mutations remains unclear, and the extent of isoform-specific modification is unknown. Here, we present the first top-down proteomics study evaluating both KRAS4A and KRAS4B, resulting in 39 completely characterized proteoforms across colorectal cancer cell lines and primary tumor samples. We determined which KRAS PTMs are present, along with their relative abundance, and that proteoforms of KRAS4A versus KRAS4B are differentially modified. Moreover, we identified a subset of KRAS4B proteoforms lacking the C185 residue and associated C-terminal PTMs. By confocal microscopy, we confirmed that this truncated GFP-KRAS4BC185* proteoform is unable to associate with the plasma membrane, resulting in a decrease in MAPK signaling pathway activation. Collectively, our study provides a reference set of functionally distinct KRAS proteoforms and the colorectal cancer contexts in which they are present. Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.

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