A molecular mechanism for the generation of ligand-dependent differential outputs by the epidermal growth factor receptor

Huang, Yongjian; Ognjenović, Jana; Karandur, Deepti; Merk, Alan; Subramaniam, Sriram; Kuriyan, John

doi:10.1101/2020.12.08.417006

Cited by 4 publications

(2 citation statements)

References 82 publications

(100 reference statements)

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“…Next, we performed PIE-FCCS experiments in the presence of EGF ligand to evaluate whether or not ex19del variants differ in their response to extracellular stimulation. A recent study demonstrated that KD mutations can directly change the conformational preferences of the ECD, potentially modulating signaling responses to ligand (41). Here, we observed that WT forms multimers upon stimulation with EGF, consistent with prior studies (ƒ c = 0.31; D = 0.13 μm 2 /s) (32, 33, 40, 42).…”

Section: Resultsmentioning

confidence: 99%

Allele-specific activation, enzyme kinetics, and inhibitor sensitivities of EGFR exon 19 deletion mutations in lung cancer

Brown

Zhang

Kim

et al. 2022

Preprint

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Oncogenic mutations within the epidermal growth factor receptor (EGFR) are found in 15-30% of all non-small cell lung carcinomas. The term exon 19 deletion (ex19del) is collectively used to refer to more than 20 distinct genomic alterations within exon 19 that comprise the most common EGFR mutation subtype in lung cancer. Despite this heterogeneity, clinical treatment decisions are made irrespective of which EGFR ex19del variant is present within the tumor, and there is a paucity of information regarding how individual ex19del variants influence protein structure and function. Herein, we identify allele-specific functional differences among ex19del variants attributable to recurring sequence and structure motifs. We build all-atom structural models of 60 ex19del variants identified in patients and combine ~400 μs of molecular dynamics simulations with biochemical and biophysical experiments to analyze three ex19del mutations (E746_A750, E746_S752>V, and L747_A750>P). We demonstrate that sequence variation in ex19del alters oncogenic cell growth, dimerization propensity, and tyrosine kinase inhibitor (TKI) sensitivity. We show that in contrast to E746_A750 and E746_S752>V, the L747_A750>P variant forms highly active ligand-independent dimers. E746_S752>V displays the least TKI sensitivity among the variants tested, which enzyme kinetic analysis and TKI inhibition experiments suggest is due to increased ATP Km relative to the common E746_A750 variant. Through these analyses, we propose an expanded framework for interpreting ex19del variants and new considerations for therapeutic intervention.SignificanceEGFR mutations are detected in approximately 30% of all lung adenocarcinomas, and the most common EGFR mutation occurring in ~50% of patients is termed “exon 19 deletion” (ex19del). Despite the existence of dozens of different genomic variants comprising what is generically referred to clinically as ex19del, clinicians currently do not distinguish between ex19del variants in considering treatment options, and the differences between ex19del variants are largely unstudied in the broader scientific community. Herein, we describe functional differences between distinct EGFR ex19del variants attributable to the structural features of each variant. These findings suggest a possible explanation for observed differences in patient outcomes stratified by ex19del subtype and reinforce the need for allele-specific considerations in clinical treatment decision making.

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Section: Resultsmentioning

confidence: 99%

Allele-specific activation, enzyme kinetics, and inhibitor sensitivities of EGFR exon 19 deletion mutations in lung cancer

Brown

Zhang

Kim

et al. 2022

Preprint

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show abstract

“…Membrane-proximal domains IV are visualized at lower resolution for both receptors (Extended Data Fig. 2) indicating their flexibility, as observed in EGFR and HER4 [22][23][24]33 . Our structure features the previously uncharacterized extended state of the HER3 extracellular domain, with NRG1 clearly resolved in the density (Fig.…”

Section: Structure Of the Asymmetric Her2/her3 Dimer Reveals That The Dimerization Arm Of Her3 Is Disengagedmentioning

confidence: 90%

Structures of the active HER2/HER3 receptor complex reveal dynamics at the dimerization interface induced by binding of a single ligand

Diwanji

Trenker

et al. 2021

Preprint

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The Human Epidermal Growth Factor Receptor 2 (HER2) and HER3 form a potent pro-oncogenic heterocomplex upon binding of growth factor neuregulin-1β (NRG1β). The mechanism by which HER2 and HER3 interact remains unknown in the absence of any structures of the complex. We isolated the NRG1β-bound near full-length HER2/HER3 dimer and obtained a 2.9Å cryo-electron microscopy (cryo-EM) reconstruction of the extracellular domain module which reveals unexpected dynamics at the HER2/HER3 dimerization interface. We show that the dimerization arm of NRG1β-bound HER3 is unresolved likely because the apo HER2 monomer fails to undergo a ligand-induced conformational change needed to establish a HER3 dimerization arm binding pocket. In a second structure of an oncogenic extracellular domain mutant of HER2, S310F, we observe a compensatory interaction with the HER3 dimerization arm that stabilizes the dimerization interface. We show that both HER2/HER3 and HER2-S310F/HER3 retain the capacity to bind to the HER2-directed therapeutic antibody, trastuzumab, but the mutant complex does not bind to pertuzumab. Our 3.5Å structure of the HER2-S310F/HER3/NRG1β/trastuzumab Fragment antigen binding (Fab) complex shows that the receptor dimer undergoes a conformational change to accommodate trastuzumab. Thus, like oncogenic mutations, therapeutics exploit the intrinsic dynamics of the HER2/HER3 heterodimer. The unique features of a singly liganded HER2/HER3 heterodimer underscore the allosteric sensing of ligand occupancy by the dimerization interface and explain why extracellular domains of HER2 do not homo-associate via a canonical active dimer interface.

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Architecture of the Sema3A/PlexinA4/Neuropilin tripartite complex

Shang

et al. 2021

Nat Commun

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Secreted class 3 semaphorins (Sema3s) form tripartite complexes with the plexin receptor and neuropilin coreceptor, which are both transmembrane proteins that together mediate semaphorin signal for neuronal axon guidance and other processes. Despite extensive investigations, the overall architecture of and the molecular interactions in the Sema3/plexin/neuropilin complex are incompletely understood. Here we present the cryo-EM structure of a near intact extracellular region complex of Sema3A, PlexinA4 and Neuropilin 1 (Nrp1) at 3.7 Å resolution. The structure shows a large symmetric 2:2:2 assembly in which each subunit makes multiple interactions with others. The two PlexinA4 molecules in the complex do not interact directly, but their membrane proximal regions are close to each other and poised to promote the formation of the intracellular active dimer for signaling. The structure reveals a previously unknown interface between the a2b1b2 module in Nrp1 and the Sema domain of Sema3A. This interaction places the a2b1b2 module at the top of the complex, far away from the plasma membrane where the transmembrane regions of Nrp1 and PlexinA4 embed. As a result, the region following the a2b1b2 module in Nrp1 must span a large distance to allow the connection to the transmembrane region, suggesting an essential role for the long non-conserved linkers and the MAM domain in neuropilin in the semaphorin/plexin/neuropilin complex.

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A molecular mechanism for the generation of ligand-dependent differential outputs by the epidermal growth factor receptor

Cited by 4 publications

References 82 publications

Allele-specific activation, enzyme kinetics, and inhibitor sensitivities of EGFR exon 19 deletion mutations in lung cancer

Allele-specific activation, enzyme kinetics, and inhibitor sensitivities of EGFR exon 19 deletion mutations in lung cancer

Structures of the active HER2/HER3 receptor complex reveal dynamics at the dimerization interface induced by binding of a single ligand

Architecture of the Sema3A/PlexinA4/Neuropilin tripartite complex

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