Germline KRAS mutations cause aberrant biochemical and physical properties leading to developmental disorders

Abstract

The KRAS gene is the most common locus for somatic gain-of-function mutations in human cancer. Germline KRAS mutations were shown recently to be associated with developmental disorders, including Noonan syndrome (NS), cardio-facio-cutaneous syndrome (CFCS), and Costello syndrome (CS). The molecular basis of this broad phenotypic variability has in part remained elusive so far. Here, we comprehensively analyzed the biochemical and structural features of ten germline KRAS mutations using physical and cellular biochemistry. According to their distinct biochemical and structural alterations, the mutants can be grouped into five distinct classes, four of which markedly differ from RAS oncoproteins. Investigated functional alterations comprise the enhancement of intrinsic and guanine nucleotide exchange factor (GEF) catalyzed nucleotide exchange, which is alternatively accompanied by an impaired GTPase-activating protein (GAP) stimulated GTP hydrolysis, an overall loss of functional properties, and a deficiency in effector interaction. In conclusion, our data underscore the important role of RAS in the pathogenesis of the group of related disorders including NS, CFCS, and CS, and provide clues to the high phenotypic variability of patients with germline KRAS mutations.

Oncogenic and RASopathy-associated K-RAS mutations relieve membrane-dependent occlusion of the effector-binding site

“…Two germ-line KRAS mutations identified in Noonan and cardio-facio-cutaneous (CFC) syndrome patients, K5N (21) and D153V (22)(23)(24), were shown to induce phosphorylation of MEK1/2 more strongly than WT KRAS; however, the mechanism of their activation has remained elusive. Unlike oncogenic K-RAS mutations such as G12D, G13D, and Q61H, neither of these germ-line mutations appreciably altered intrinsic GTPase activity or nucleotide exchange or sensitivity to GAPs or guanine nucleotide exchange factors (GEFs) in solution; thus, they have been classified as mutants whose phenotypes are not explained by their biochemical properties (25,26). Intriguingly, we find that these mutations change the electrostatic surface of the two main membrane interfaces of K-RAS4B (Fig.…”

“…Isoleucine residues exhibiting peak broadening of >20% were defined as active, and the adjacent N-and C-terminal residues were defined as passive residues. For simulation of K-RAS4B-GDP, isoleucines 24, 46, 55, 100, 139, 142, and 163 were defined as active and residues 23,25,45,47,54,56,99,101,138,140,141,143,162, and 164 were defined as passive. For simulation of K-RAS4B-GMPPNP, isoleucines 24, 36, 46, 55, 100, 139, 142, and 163 were identified as active residues and 23, 25,35,37,45,47,54,56,99,101,138,140,141,143,162, and 164 as passive residues.…”

“…For simulation of K-RAS4B-GDP, isoleucines 24, 46, 55, 100, 139, 142, and 163 were defined as active and residues 23,25,45,47,54,56,99,101,138,140,141,143,162, and 164 were defined as passive. For simulation of K-RAS4B-GMPPNP, isoleucines 24, 36, 46, 55, 100, 139, 142, and 163 were identified as active residues and 23, 25,35,37,45,47,54,56,99,101,138,140,141,143,162, and 164 as passive residues. Surface electrostatics of K-RAS4B were generated by the Poisson-Boltzmann Solver (34)(35)(36).…”

C4ST-1/CHST11-controlled chondroitin sulfation interferes with oncogenic HRAS signaling in Costello syndrome

“…The MAPK pathways are critical downstream mediators of RAS signal transduction, and have been shown to be upregulated in response to oncogenic HRAS mutations in Costello syndrome. 8,18,38,39 Thus, C7669 fibroblasts were treated with inhibitors of the MAP-kinases Erk (PD98059), p38 (SB203580) and SAPK/JNK (SP600125) for 48 h, and subsequently analyzed for C4ST-1 mRNA expression by qRT-PCR (Figure 2n). Blocking all three MAPK cascades was able to significantly elevate the C4ST-1 expression by approximately 2-to 3.5-fold when compared with a DMSO-treated control.…”

The Function of Embryonic Stem Cell-expressed RAS (E-RAS), a Unique RAS Family Member, Correlates with Its Additional Motifs and Its Structural Properties

“…Therefore, they play a key role in various cellular processes, including gene expression, metabolism, cell cycle progression, proliferation, survival, and differentiation. Somatic or germ line mutations in genes related to members of the RAS family or their regulators are commonly associated with cancer progression or developmental disorders (3)(4)(5)(6)(7)(8)(9).…”