KRAS4a and KRAS4b show distinct lipid-dependent regulation of RAS-RAF membrane dynamics

Konstantia Georgouli; Jeremy O.B. Tempkin; Liam G. Stanton; Tomas Oppelstrup; Rebika Shrestha; Timothy S. Carpenter; Fikret Aydin; Xiaohua Zhang; Harsh Bhatia; Yue Yang; Que N. Van; Pedro Andrade Bonilla; Gulcin Gulten; Debanjan Goswami; Francesco Di Natale; Joseph R. Chavez; Joseph Y. Moon; Gautham Dharuman; Nicolas W. Hengartner; Dhirendra K. Simanshu; Timothy H. Tran; Kien Nguyen; Christopher B. Stanley; Brian Van Essen; Peer Timo Bremer; Felice C. Lightstone; Andrew G. Stephen; James N. Glosli; Sandrasegaram Gnanakaran; Thomas J. Turbyville; Frank McCormick; Dwight V. Nissley; Frederick H. Streitz; Helgi I. Ingólfsson

doi:10.1016/j.jbc.2026.111237

KRAS4a and KRAS4b show distinct lipid-dependent regulation of RAS-RAF membrane dynamics

Konstantia Georgouli
, Jeremy O.B. Tempkin
, Liam G. Stanton
, Tomas Oppelstrup
, Rebika Shrestha
, Timothy S. Carpenter
, Fikret Aydin
, Xiaohua Zhang
, Harsh Bhatia
, Yue Yang
, Que N. Van
, Pedro Andrade Bonilla
, Gulcin Gulten
, Debanjan Goswami
, Francesco Di Natale
, Joseph R. Chavez
, Joseph Y. Moon
, Gautham Dharuman
, Nicolas W. Hengartner
, Dhirendra K. Simanshu

Timothy H. Tran, Kien Nguyen, Christopher B. Stanley, Brian Van Essen, Peer Timo Bremer, Felice C. Lightstone, Andrew G. Stephen, James N. Glosli, Sandrasegaram Gnanakaran, Thomas J. Turbyville, Frank McCormick, Dwight V. Nissley, Frederick H. Streitz, Helgi I. Ingólfsson

Scalable Biomedical Modeling

Research output: Contribution to journal › Article › peer-review

Abstract

KRAS4a and KRAS4b are important regulators of signaling, and their interactions with the plasma membrane are dynamic and influenced by lipid composition. KRAS 4a and 4b have nearly identical globular domains but differ in their membrane-associated hyper variable region (HVR). The functional distinctions between these isoforms remain unclear, particularly with regards to their dependence on specific lipids and the membrane environment. Previous work showed that the membrane orientation of KRAS4b affects its ability to bind to RAF kinase RBDCRD and that the KRAS–RBDCRD complex adopts different poses on the membrane as well as influences the size and composition of the lipid environment. To model differences between KRAS 4a and 4b protein–lipid interactions, we extended the Multiscale Machine-Learned Modeling Infrastructure (MuMMI) to incorporate continuum simulations in the grand canonical ensemble, enabling sampling across macroscopic, coarse-grained, and all-atom resolutions. Using this framework, we systematically altered PIP2 concentrations, KRAS 4a versus 4b, and RAF RBDCRD complexation to assess impacts on membrane–protein interactions and dynamics. Our results reveal that reducing PIP2 shifts and broadens the membrane orientational preference of both KRAS 4b and 4a, with stronger effects on 4b HVR localization versus 4a. We demonstrate that with depletion of the strong negatively charged PIP2 lipid, the less charged phosphatidylserine replaces PIP2. Our findings highlight similarities and distinctions in the dynamics and lipid dependency of KRAS isoforms and suggest that ordering of the local lipid composition by HVRs is a shared property and key modulator of RAS-mediated signaling at the plasma membrane.

Original language	English
Article number	111237
Journal	Journal of Biological Chemistry
Volume	302
Issue number	3
DOIs	https://doi.org/10.1016/j.jbc.2026.111237
State	Published - Mar 2026

Funding

This project has been funded in part with federal funds from the NCI , NIH , under contract no. 75N91019D00024 . This work was supported by the Joint Design of Advanced Computing Solutions for Cancer ( JDACS4C ) program established by the U.S. DOE and the NCI of the National Institutes of Health. This work was performed under the auspices of the U.S. Department of Energy (DOE) by Lawrence Livermore National Laboratory (LLNL) under Contract DE-AC52-07NA27344, Los Alamos National Laboratory (LANL) under Contract DE-AC5206NA25396, Oak Ridge National Laboratory under Contract DE-AC05-00OR22725, Argonne National Laboratory (ANL) under Contract DE-AC02-06-CH11357, and under the auspices of the National Cancer Institute (NCI) by Frederick National Laboratory for Cancer Research (FNLCR) under Contract 75N91019D00024. This research used resources of the Oak Ridge Leadership Computing Facility (OLCF), which is a DOE Office of Science User Facility supported under Contract DE-AC05-00OR22725. For computing time, we thank the Advanced Scientific Computing Research Leadership Computing Challenge (ALCC) for time onSummit, the Livermore Institutional Grand Challenge for time onLassenand LANL Institutional computing. The LANL Institutional Computing Program is supported by the U.S. DOE National Nuclear Security Administration under Contract No. DE-AC52-06NA25396. For computing support, we thank OLCF and LC staff. Release: LLNL-JRNL-2009802.Author contributionsK. G., J. O. B. T., L. G. S., T. O., R. S., T. S. C., F. A., H. B., A. G. S., T. J. T., D. V. N., and H. I. I. writing–original draft; K. G., J. O. B. T., L. G. S., T. O., R. S., T. S. C., F. A., X. Z., H. B., Y. Y., Q. N. V., P. A. B., G. G., D. G., F. D. N., J. R. C., J. Y. M., G. D., N. W. H., D. K. S., T. H. T., K. N., C. B. S., B. V. E., P.-T. B., F. C. L., A. G. S., J. N. G., S. G., T. J. T., F. M., D. V. N., F. H S., and H. I. I. methodology; K. G., J. O. B. T., L. G. S., T. O., R. S., T. S. C., F. A., and H. I. I. formal analysis; K. G., J. O. B. T., L. G. S., T. O., R. S., T. S. C., F. A., X. Z., H. B., Y. Y., Q. N. V., P. A. B., G. G., D. G., F. D. N., J. R. C., J. Y. M., G. D., N. W. H., D. K. S., T. H. T., K. N., C. B. S., B. V. E., P.-T. B., F. C. L., A. G. S., J. N. G., S. G., T. J. T., F. M., D. V. N., F. H S., and H. I. I. conceptualization; F. C. L., S. G., T. J. T., D. V. N., F. H S., and H. I. I. supervision.Funding and additional informationThis project has been funded in part with federal funds from theNCI,NIH, under contract no.75N91019D00024. This work was supported by theJoint Design of Advanced Computing Solutionsfor Cancer (JDACS4C) program established by the U.S. DOE and the NCI of the National Institutes of Health. This work was performed under the auspices of the U.S. Department of Energy (DOE) by Lawrence Livermore National Laboratory (LLNL) under Contract DE-AC52-07NA27344, Los Alamos National Laboratory (LANL) under Contract DE-AC5206NA25396, Oak Ridge National Laboratory under Contract DE-AC05-00OR22725, Argonne National Laboratory (ANL) under Contract DE-AC02-06-CH11357, and under the auspices of the National Cancer Institute (NCI) by Frederick National Laboratory for Cancer Research (FNLCR) under Contract 75N91019D00024. This research used resources of the Oak Ridge Leadership Computing Facility (OLCF), which is a DOE Office of Science User Facility supported under Contract DE-AC05-00OR22725. For computing time, we thank the Advanced Scientific Computing Research Leadership Computing Challenge (ALCC) for time on Summit, the Livermore Institutional Grand Challenge for time on Lassen and LANL Institutional computing. The LANL Institutional Computing Program is supported by the U.S. DOE National Nuclear Security Administration under Contract No. DE-AC52-06NA25396. For computing support, we thank OLCF and LC staff. Release: LLNL-JRNL-2009802.

Keywords

KRAS function
KRAS4a
KRAS4b
RAS-RBDCRD membrane dynamics
RAS-membrane biology
RBDCRD of RAF
massive parallel simulations
multiscale modeling

Access to Document

10.1016/j.jbc.2026.111237

Cite this

Georgouli, K., Tempkin, J. O. B., Stanton, L. G., Oppelstrup, T., Shrestha, R., Carpenter, T. S., Aydin, F., Zhang, X., Bhatia, H., Yang, Y., Van, Q. N., Bonilla, P. A., Gulten, G., Goswami, D., Di Natale, F., Chavez, J. R., Moon, J. Y., Dharuman, G., Hengartner, N. W., ... Ingólfsson, H. I. (2026). KRAS4a and KRAS4b show distinct lipid-dependent regulation of RAS-RAF membrane dynamics. Journal of Biological Chemistry, 302(3), Article 111237. https://doi.org/10.1016/j.jbc.2026.111237

@article{66005c3613174e839474940756411494,

title = "KRAS4a and KRAS4b show distinct lipid-dependent regulation of RAS-RAF membrane dynamics",

abstract = "KRAS4a and KRAS4b are important regulators of signaling, and their interactions with the plasma membrane are dynamic and influenced by lipid composition. KRAS 4a and 4b have nearly identical globular domains but differ in their membrane-associated hyper variable region (HVR). The functional distinctions between these isoforms remain unclear, particularly with regards to their dependence on specific lipids and the membrane environment. Previous work showed that the membrane orientation of KRAS4b affects its ability to bind to RAF kinase RBDCRD and that the KRAS–RBDCRD complex adopts different poses on the membrane as well as influences the size and composition of the lipid environment. To model differences between KRAS 4a and 4b protein–lipid interactions, we extended the Multiscale Machine-Learned Modeling Infrastructure (MuMMI) to incorporate continuum simulations in the grand canonical ensemble, enabling sampling across macroscopic, coarse-grained, and all-atom resolutions. Using this framework, we systematically altered PIP2 concentrations, KRAS 4a versus 4b, and RAF RBDCRD complexation to assess impacts on membrane–protein interactions and dynamics. Our results reveal that reducing PIP2 shifts and broadens the membrane orientational preference of both KRAS 4b and 4a, with stronger effects on 4b HVR localization versus 4a. We demonstrate that with depletion of the strong negatively charged PIP2 lipid, the less charged phosphatidylserine replaces PIP2. Our findings highlight similarities and distinctions in the dynamics and lipid dependency of KRAS isoforms and suggest that ordering of the local lipid composition by HVRs is a shared property and key modulator of RAS-mediated signaling at the plasma membrane.",

keywords = "KRAS function, KRAS4a, KRAS4b, RAS-RBDCRD membrane dynamics, RAS-membrane biology, RBDCRD of RAF, massive parallel simulations, multiscale modeling",

author = "Konstantia Georgouli and Tempkin, \{Jeremy O.B.\} and Stanton, \{Liam G.\} and Tomas Oppelstrup and Rebika Shrestha and Carpenter, \{Timothy S.\} and Fikret Aydin and Xiaohua Zhang and Harsh Bhatia and Yue Yang and Van, \{Que N.\} and Bonilla, \{Pedro Andrade\} and Gulcin Gulten and Debanjan Goswami and \{Di Natale\}, Francesco and Chavez, \{Joseph R.\} and Moon, \{Joseph Y.\} and Gautham Dharuman and Hengartner, \{Nicolas W.\} and Simanshu, \{Dhirendra K.\} and Tran, \{Timothy H.\} and Kien Nguyen and Stanley, \{Christopher B.\} and \{Van Essen\}, Brian and Bremer, \{Peer Timo\} and Lightstone, \{Felice C.\} and Stephen, \{Andrew G.\} and Glosli, \{James N.\} and Sandrasegaram Gnanakaran and Turbyville, \{Thomas J.\} and Frank McCormick and Nissley, \{Dwight V.\} and Streitz, \{Frederick H.\} and Ing{\'o}lfsson, \{Helgi I.\}",

note = "Publisher Copyright: {\textcopyright} 2026 The Authors. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY-NC-ND license. http://creativecommons.org/licenses/by-nc-nd/4.0/",

year = "2026",

month = mar,

doi = "10.1016/j.jbc.2026.111237",

language = "English",

volume = "302",

journal = "Journal of Biological Chemistry",

issn = "0021-9258",

publisher = "Elsevier",

number = "3",

}

Georgouli, K, Tempkin, JOB, Stanton, LG, Oppelstrup, T, Shrestha, R, Carpenter, TS, Aydin, F, Zhang, X, Bhatia, H, Yang, Y, Van, QN, Bonilla, PA, Gulten, G, Goswami, D, Di Natale, F, Chavez, JR, Moon, JY, Dharuman, G, Hengartner, NW, Simanshu, DK, Tran, TH, Nguyen, K, Stanley, CB, Van Essen, B, Bremer, PT, Lightstone, FC, Stephen, AG, Glosli, JN, Gnanakaran, S, Turbyville, TJ, McCormick, F, Nissley, DV, Streitz, FH & Ingólfsson, HI 2026, 'KRAS4a and KRAS4b show distinct lipid-dependent regulation of RAS-RAF membrane dynamics', Journal of Biological Chemistry, vol. 302, no. 3, 111237. https://doi.org/10.1016/j.jbc.2026.111237

TY - JOUR

T1 - KRAS4a and KRAS4b show distinct lipid-dependent regulation of RAS-RAF membrane dynamics

AU - Georgouli, Konstantia

AU - Tempkin, Jeremy O.B.

AU - Stanton, Liam G.

AU - Oppelstrup, Tomas

AU - Shrestha, Rebika

AU - Carpenter, Timothy S.

AU - Aydin, Fikret

AU - Zhang, Xiaohua

AU - Bhatia, Harsh

AU - Yang, Yue

AU - Van, Que N.

AU - Bonilla, Pedro Andrade

AU - Gulten, Gulcin

AU - Goswami, Debanjan

AU - Di Natale, Francesco

AU - Chavez, Joseph R.

AU - Moon, Joseph Y.

AU - Dharuman, Gautham

AU - Hengartner, Nicolas W.

AU - Simanshu, Dhirendra K.

AU - Tran, Timothy H.

AU - Nguyen, Kien

AU - Stanley, Christopher B.

AU - Van Essen, Brian

AU - Bremer, Peer Timo

AU - Lightstone, Felice C.

AU - Stephen, Andrew G.

AU - Glosli, James N.

AU - Gnanakaran, Sandrasegaram

AU - Turbyville, Thomas J.

AU - McCormick, Frank

AU - Nissley, Dwight V.

AU - Streitz, Frederick H.

AU - Ingólfsson, Helgi I.

N1 - Publisher Copyright: © 2026 The Authors. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY-NC-ND license. http://creativecommons.org/licenses/by-nc-nd/4.0/

PY - 2026/3

Y1 - 2026/3

N2 - KRAS4a and KRAS4b are important regulators of signaling, and their interactions with the plasma membrane are dynamic and influenced by lipid composition. KRAS 4a and 4b have nearly identical globular domains but differ in their membrane-associated hyper variable region (HVR). The functional distinctions between these isoforms remain unclear, particularly with regards to their dependence on specific lipids and the membrane environment. Previous work showed that the membrane orientation of KRAS4b affects its ability to bind to RAF kinase RBDCRD and that the KRAS–RBDCRD complex adopts different poses on the membrane as well as influences the size and composition of the lipid environment. To model differences between KRAS 4a and 4b protein–lipid interactions, we extended the Multiscale Machine-Learned Modeling Infrastructure (MuMMI) to incorporate continuum simulations in the grand canonical ensemble, enabling sampling across macroscopic, coarse-grained, and all-atom resolutions. Using this framework, we systematically altered PIP2 concentrations, KRAS 4a versus 4b, and RAF RBDCRD complexation to assess impacts on membrane–protein interactions and dynamics. Our results reveal that reducing PIP2 shifts and broadens the membrane orientational preference of both KRAS 4b and 4a, with stronger effects on 4b HVR localization versus 4a. We demonstrate that with depletion of the strong negatively charged PIP2 lipid, the less charged phosphatidylserine replaces PIP2. Our findings highlight similarities and distinctions in the dynamics and lipid dependency of KRAS isoforms and suggest that ordering of the local lipid composition by HVRs is a shared property and key modulator of RAS-mediated signaling at the plasma membrane.

AB - KRAS4a and KRAS4b are important regulators of signaling, and their interactions with the plasma membrane are dynamic and influenced by lipid composition. KRAS 4a and 4b have nearly identical globular domains but differ in their membrane-associated hyper variable region (HVR). The functional distinctions between these isoforms remain unclear, particularly with regards to their dependence on specific lipids and the membrane environment. Previous work showed that the membrane orientation of KRAS4b affects its ability to bind to RAF kinase RBDCRD and that the KRAS–RBDCRD complex adopts different poses on the membrane as well as influences the size and composition of the lipid environment. To model differences between KRAS 4a and 4b protein–lipid interactions, we extended the Multiscale Machine-Learned Modeling Infrastructure (MuMMI) to incorporate continuum simulations in the grand canonical ensemble, enabling sampling across macroscopic, coarse-grained, and all-atom resolutions. Using this framework, we systematically altered PIP2 concentrations, KRAS 4a versus 4b, and RAF RBDCRD complexation to assess impacts on membrane–protein interactions and dynamics. Our results reveal that reducing PIP2 shifts and broadens the membrane orientational preference of both KRAS 4b and 4a, with stronger effects on 4b HVR localization versus 4a. We demonstrate that with depletion of the strong negatively charged PIP2 lipid, the less charged phosphatidylserine replaces PIP2. Our findings highlight similarities and distinctions in the dynamics and lipid dependency of KRAS isoforms and suggest that ordering of the local lipid composition by HVRs is a shared property and key modulator of RAS-mediated signaling at the plasma membrane.

KW - KRAS function

KW - KRAS4a

KW - KRAS4b

KW - RAS-RBDCRD membrane dynamics

KW - RAS-membrane biology

KW - RBDCRD of RAF

KW - massive parallel simulations

KW - multiscale modeling

UR - https://www.scopus.com/pages/publications/105034107644

U2 - 10.1016/j.jbc.2026.111237

DO - 10.1016/j.jbc.2026.111237

M3 - Article

C2 - 41651414

AN - SCOPUS:105034107644

SN - 0021-9258

VL - 302

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 3

M1 - 111237

ER -

KRAS4a and KRAS4b show distinct lipid-dependent regulation of RAS-RAF membrane dynamics

Abstract

Funding

Keywords

Access to Document

Other files and links

Fingerprint

Cite this