Noonan syndrome gain-of-function mutations in<i>NRAS</i>cause zebrafish gastrulation defects

Runtuwene, Vincent; Eekelen, Mark van; Overvoorde, John; Rehmann, Holger; Yntema, Helger G.; Nillesen, Willy M.; Haeringen, Arie van; Burgt, Ineke van der; Burgering, Boudewijn M.T.; Hertog, Jeroen den

doi:10.1242/dmm.007112

Cited by 61 publications

(91 citation statements)

References 33 publications

(48 reference statements)

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“…The morphological defects elicited by mutant A2ml1 resembled those induced by NS-associated Shp2-D61G and N-Ras-I24N. 12,18 Of note, A2ml1-mutation-induced morphological defects appeared later in development (apparent from 3 dpf onwards) and were less severe than those observed for Shp2-D61G-induced defects (from 6 hpf onwards).…”

Section: A2ml1 Mutations In a Noonan Syndrome-like Disorder Lelm Vissmentioning

confidence: 91%

Heterozygous germline mutations in A2ML1 are associated with a disorder clinically related to Noonan syndrome

et al. 2014

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Noonan syndrome (NS) is a developmental disorder characterized by short stature, facial dysmorphisms and congenital heart defects. To date, all mutations known to cause NS are dominant, activating mutations in signal transducers of the RAS/ mitogen-activated protein kinase (MAPK) pathway. In 25% of cases, however, the genetic cause of NS remains elusive, suggesting that factors other than those involved in the canonical RAS/MAPK pathway may also have a role. Here, we used family-based whole exome sequencing of a case-parent trio and identified a de novo mutation, p.(Arg802His), in A2ML1, which encodes the secreted protease inhibitor a-2-macroglobulin (A2M)-like-1. Subsequent resequencing of A2ML1 in 155 cases with a clinical diagnosis of NS led to the identification of additional mutations in two families, p.(Arg802Leu) and p.(Arg592Leu). Functional characterization of these human A2ML1 mutations in zebrafish showed NS-like developmental defects, including a broad head, blunted face and cardiac malformations. Using the crystal structure of A2M, which is highly homologous to A2ML1, we identified the intramolecular interaction partner of p.Arg802. Mutation of this residue, p.Glu906, induced similar developmental defects in zebrafish, strengthening our conclusion that mutations in A2ML1 cause a disorder clinically related to NS. This is the first report of the involvement of an extracellular factor in a disorder clinically related to RASopathies, providing potential new leads for better understanding of the molecular basis of this family of developmental diseases.

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Section: A2ml1 Mutations In a Noonan Syndrome-like Disorder Lelm Vissmentioning

confidence: 91%

Heterozygous germline mutations in A2ML1 are associated with a disorder clinically related to Noonan syndrome

et al. 2014

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“…However, as the pregnancy in the prior reports and two reported here were either Initial studies suggest that variants in NRAS have a similar spectrum as those in PTPN11, with mildly activating variants causing Noonan syndrome and strong gain-of-function activity variants acting as initiating drivers for isolated JMML. 10,11 Supporting this, most germline NRAS variants associated with Noonan syndrome (c.179G4A (p. (Gly60Glu)), c.71T4A (p. (Ile24Asn)), and c.149C4T (p.(Thr50Ile))) were found to be mildly activating when compared with the recurrent oncogenic variant, c.35G4T (p.(Gly12Val)). 10,11 In addition, embryonic expression of another oncogenic NRAS variant, p.(Gly12Asp), was embryonic lethal in mice.…”

Section: Discussionmentioning

confidence: 80%

“…9 Pathogenic variants in PTPN11 or NRAS causing isolated JMML rarely overlap with those causing Noonan syndrome. [10][11][12][13][14] This has been attributed to the JMML variants having a stronger gain-offunction activity than RASopathy-associated variants. There are rare observations of Noonan-associated PTPN11 variants identified in isolated JMML, and a few reported cases of germline inheritance of JMML variants.…”

Section: Introductionmentioning

confidence: 99%

Juvenile myelomonocytic leukemia-associated variants are associated with neo-natal lethal Noonan syndrome

et al. 2017

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Gain-of-function variants in some RAS-MAPK pathway genes, including PTPN11 and NRAS, are associated with RASopathies and/or acquired hematological malignancies, most notably juvenile myelomonocytic leukemia (JMML). With rare exceptions, the spectrum of germline variants causing RASopathies does not overlap with the somatic variants identified in isolated JMML. Studies comparing these variants suggest a stronger gain-of-function activity in the JMML variants. As JMML variants have not been identified as germline defects and have a greater impact on protein function, it has been speculated that they would be embryonic lethal. Here we identified three variants, which have previously only been identified in isolated somatic JMML and other sporadic cancers, in four cases with a severe pre-or neo-natal lethal presentation of Noonan syndrome. These cases support the hypothesis that these stronger gain-of-function variants are rarely compatible with life.

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“…This easily detectable oval shape phenotype in the zebrafish embryo was first noticed to be caused by RASopathy variants in Shp2, a protein tyrosine phosphatase (35), and later observed with four MEK1 RASopathy mutations (36). This assay has since been used in a more quantitative manner by using the major-to-minor axis ratio of the zebrafish embryo as a metric to denote the severity of the defect (29). Because previous studies measured embryo shapes at different times, we measured the major-tominor axis ratio (aspect ratio) over time and determined 11 hpf as the best time to maximize dynamic range (SI Appendix, Figs.…”

Section: Resultsmentioning

confidence: 99%

“…Although previous studies have indirectly observed mutationbased severity of effects using cultured cells (16,27,28) and model organisms (29)(30)(31) by comparing a few mutations, in no case were the studies comprehensive. To address this more systematically, we develop a framework for the quantitative comparison of MEK1 mutants found in human diseases.…”

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confidence: 99%

In vivo severity ranking of Ras pathway mutations associated with developmental disorders

Jindal

Goyal

Yamaya

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Germ-line mutations in components of the Ras/MAPK pathway result in developmental disorders called RASopathies, affecting about 1/1,000 human births. Rapid advances in genome sequencing make it possible to identify multiple disease-related mutations, but there is currently no systematic framework for translating this information into patient-specific predictions of disease progression. As a first step toward addressing this issue, we developed a quantitative, inexpensive, and rapid framework that relies on the early zebrafish embryo to assess mutational effects on a common scale. Using this assay, we assessed 16 mutations reported in MEK1, a MAPK kinase, and provide a robust ranking of these mutations. We find that mutations found in cancer are more severe than those found in both RASopathies and cancer, which, in turn, are generally more severe than those found only in RASopathies. Moreover, this rank is conserved in other zebrafish embryonic assays and Drosophila-specific embryonic and adult assays, suggesting that our ranking reflects the intrinsic property of the mutant molecule. Furthermore, this rank is predictive of the drug dose needed to correct the defects. This assay can be readily used to test the strengths of existing and newly found mutations in MEK1 and other pathway components, providing the first step in the development of rational guidelines for patient-specific diagnostics and treatment of RASopathies.T he Ras/MAPK (rat sarcoma/mitogen-activated protein kinase) signaling pathway is involved in essentially all aspects of organismal development, from the first cell divisions in the early embryo to postnatal development and growth (1-3). Given its critical function, it is not surprising that deregulated Ras/ MAPK signaling, resulting from either genetic or environmental perturbations, can lead to developmental abnormalities. A large class of such abnormalities, known as RASopathies, is associated with activating germ-line mutations in many components of the Ras pathway (4). Estimated to affect 1/1,000 human births, these abnormalities are characterized by a broad spectrum of phenotypes, including cardiac defects, craniofacial dysmorphisms, and neurocognitive delays (4). Although the origins of these phenotypes are still poorly understood, studies of model organisms show that many of the observed structural and functional defects can indeed be mimicked by targeted introduction of mutations found in RASopathies (5). Hundreds of such mutations have already been identified, and many more are likely to be discovered by the sequencing of affected individuals (6).Importantly, these studies identify new mutations in the very same components that are mutated in cancer and have been extensively studied both in vitro and in vivo (7-9). In particular, multiple new mutations were identified in MEK1, a MAPK kinase (10) and an important target for anticancer therapeutics (11)(12)(13)(14)(15)(16)(17)(18)(19). Because it is commonly believed that cancer mutations would be embryonic lethal when inherited through t...

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Noonan syndrome gain-of-function mutations inNRAScause zebrafish gastrulation defects

Cited by 61 publications

References 33 publications

Heterozygous germline mutations in A2ML1 are associated with a disorder clinically related to Noonan syndrome

Heterozygous germline mutations in A2ML1 are associated with a disorder clinically related to Noonan syndrome

Juvenile myelomonocytic leukemia-associated variants are associated with neo-natal lethal Noonan syndrome

In vivo severity ranking of Ras pathway mutations associated with developmental disorders

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