rs397516830

chr3-12604182-A-Cchr3-12604182-A-Gchr3-12604182-A-T

Variant summary

Our verdict is Pathogenic. Variant got 16 ACMG points: 16P and 0B. PM1 PM2 PM5 PP2 PP3 PP5_Very_Strong

The NM_002880.4(RAF1):c.788T>G(p.Val263Gly) variant causes a missense change involving the alteration of a conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. In-silico tool predicts a pathogenic outcome for this variant. Variant has been reported in ClinVar as Likely pathogenic (★★★). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. V263A) has been classified as Likely pathogenic.

• Frequency: Genomes: not found (cov: 33)
• Consequence: RAF1 NM_002880.4 missense
• Clinical Significance: Likely pathogenic reviewed by expert panel P:7
• Conservation: PhyloP100: 9.32

Genes affected

RAF1 (HGNC:9829): (Raf-1 proto-oncogene, serine/threonine kinase) This gene is the cellular homolog of viral raf gene (v-raf). The encoded protein is a MAP kinase kinase kinase (MAP3K), which functions downstream of the Ras family of membrane associated GTPases to which it binds directly. Once activated, the cellular RAF1 protein can phosphorylate to activate the dual specificity protein kinases MEK1 and MEK2, which in turn phosphorylate to activate the serine/threonine specific protein kinases, ERK1 and ERK2. Activated ERKs are pleiotropic effectors of cell physiology and play an important role in the control of gene expression involved in the cell division cycle, apoptosis, cell differentiation and cell migration. Mutations in this gene are associated with Noonan syndrome 5 and LEOPARD syndrome 2. [provided by RefSeq, Jul 2008]

ACMG classification

Verdict is Pathogenic. Variant got 16 ACMG points.

• PM1: In a hotspot region, there are 9 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 0 benign, 7 uncertain in NM_002880.4
• PM2: Very rare variant in population databases, with high coverage
• PM5: Other missense variant is known to change same aminoacid residue: Variant chr3-12604182-A-G is described in ClinVar as [Likely_pathogenic]. Clinvar id is 40608.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars.
• PP2: Missense variant where missense usually causes diseases, RAF1
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.813
• PP5: Variant 3-12604182-A-C is Pathogenic according to our data. Variant chr3-12604182-A-C is described in ClinVar as [Likely_pathogenic]. Clinvar id is 40607.Status of the report is reviewed_by_expert_panel, 3 stars. Variant chr3-12604182-A-C is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
RAF1	NM_002880.4	c.788T>G	p.Val263Gly	missense_variant	7/17	ENST00000251849.9

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
RAF1	ENST00000251849.9	c.788T>G	p.Val263Gly	missense_variant	7/17	1	NM_002880.4	P3

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 33

ClinVar

Significance: Likely pathogenic

Submissions summary: Pathogenic:7

Revision: reviewed by expert panel

Submissions by phenotype

RASopathy Pathogenic:3

Likely pathogenic, reviewed by expert panel	curation	ClinGen RASopathy Variant Curation Expert Panel	Apr 03, 2017	The c.788T>G (p.Val263Gly) variant in RAF1 has been reported in the literature as an unconfirmed de novo occurrence in a patient with clinical features of a RASopathy (PM6; GeneDx internal data; GTR ID: 26957; ClinVar SCV000061370.9). The p.Val263Gly variant has been identified in 6 other independent occurrences in patients with a RASopathy (PS4 not met; Partners LMM, Blueprint genetics, Institute of Human Genetics, Otto von Guericke University Magdeburg, APHP-Robert Debré Hospital internal data; GTR ID's: 21766, 500188, 506381, 28338; ClinVar SCV000209021.9, SCV000207170.1). This variant was absent from large population studies (PM2; ExAC, http://exac.broadinstitute.org). A different pathogenic missense variant has been previously identified at this codon of RAF1 which may indicate that this residue is critical to the function of the protein (PM5; ClinVar 496189, 40608). Furthermore, the variant is in a location that has been defined by the ClinGen RASopathy Expert Panel to be a mutational hotspot or domain of RAF1 (PM1; PMID 29493581). The variant is located in the RAF1 gene, which has been defined by the ClinGen RASopathy Expert Panel as a gene with a low rate of benign missense variants and pathogenic missense variants are common (PP2; PMID: 29493581). Computational prediction tools and conservation analysis suggest that the p.Val263Gly variant may impact the protein (PP3). In summary, this variant meets criteria to be classified as likely pathogenic for RASopathies in an autosomal dominant manner. RASopathy-specific ACMG/AMP criteria applied (PMID:29493581): PM6, PM5, PM2, PM1, PP3, PP2. -
Pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Apr 01, 2020	RAF1 c.788T>G (p.Val263Gly) results in a non-conservative amino acid change located in a region of the CR2 functional domain of the encoded protein sequence supporting pathogenicity, as defined by the ClinGen RASopathy Expert Panel (PM1; PMID 29493581). Five of five in-silico tools predict a damaging effect of the variant on protein function (PP3). The variant was absent in 251466 control chromosomes in the gnomAD database (PM2). c.788T>G has been reported in the literature in at least 5 individuals affected with Noonan Syndrome and Related Conditions (e.g. Chen_2019, Clinton_2019, Ghedira_2018, Rodriguez_2019, Schulz_2012) (PS4). The variant was reported as a de novo occurrence (confirmed via parental testing) in at least 3 of the documented patients (e.g. Chen_2019, Ghedira_2018, Schulz_2012) (PS2). Additional patients with clinical features of a RASopathy or specifically Noonan syndrome have been reported in the ClinVar database (SCV000616423.3, SCV000061370.6). These data indicate that the variant is very likely to be associated with disease. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. However, a different variant affecting the same amino acid, p.Val263Ala, has been functionally assessed and showed increased in vitro kinase and ERK activation, behaving as a gain-of-function mutant (PMID 17603482). Kobayashi et al (2010) (PMID 20052757) report in their study that mutations in RAF1 were clustered in the conserved region 2 (CR2) domain, which carries an inhibitory phosphorylation site (S259). Functional studies suggest that dephosphorylation of S259 is the primary pathogenic mechanism in the activation of RAF1 mutants located in the CR2 domain. The ClinGen RASopathy Variant Curation Expert Panel cites the variant in ClinVar (evaluation after 2014) as likely pathogenic. Based on the evidence outlined above, the variant was re-classified as pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Invitae	Oct 22, 2023	This sequence change replaces valine, which is neutral and non-polar, with glycine, which is neutral and non-polar, at codon 263 of the RAF1 protein (p.Val263Gly). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individual(s) with Noonan syndrome (PMID: 30157809, 30732632). In at least one individual the variant was observed to be de novo. ClinVar contains an entry for this variant (Variation ID: 40607). Advanced modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) performed at Invitae indicates that this missense variant is expected to disrupt RAF1 protein function. This variant disrupts the p.Val263 amino acid residue in RAF1. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 17603482; Invitae). This suggests that this residue is clinically significant, and that variants that disrupt this residue are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic. -

Noonan syndrome Pathogenic:2

Pathogenic, no assertion criteria provided	clinical testing	Blueprint Genetics	Jul 07, 2014	- -
Likely pathogenic, criteria provided, single submitter	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Nov 19, 2013	The Val263Gly variant has now been identified by our laboratory in three individ uals with clinical features of Noonan syndrome. In addition, a different amino a cid change at the same position (Val263Ala) has also been associated with the cl inical features of Noonan syndrome (Razzaque 2007). This variant has not been i dentified in large population studies. Computational analyses (biochemical amino acid properties, conservation, AlignGVGD, PolyPhen2, and SIFT) suggest that thi s variant may impact the protein. Of note, individuals with pathogenic variants in exon 7 or 17 in RAF1 have a high incidence of hypertrophic cardiomyopathy (80 -95%, Razzaque 2007). In summary, the Val263Gly variant is likely to be pathogen ic, although additional information is needed to fully establish its clinical si gnificance. -

Noonan syndrome 1 Pathogenic:1

Pathogenic, no assertion criteria provided

clinical testing

Molecular Genetics, Centre for Human Genetics

-

- -

not provided Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

GeneDx

Mar 18, 2012

The V263G missense mutation has not been published as a mutation, nor has it been reported as a benign polymorphism to our knowledge. This mutation has been seen before at GeneDx and another missense mutation at this codon (V263A) was previously reported in a patient diagnosed with Noonan syndrome and hypertrophic cardiomyopathy (Razzaque et al., 2007). Furthermore, this mutation is located in a highly conserved portion of the RAF1 gene, where many other gain-of function mutations (P261A, P261S and P261L) have also been reported in association with Noonan syndrome (Pandit et al., 2007 and Razzaque et al., 2007). Therefore, V263G is considered to be a disease-causing mutation. The variant is found in NOONAN panel(s). -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.98
BayesDel_addAF	Pathogenic	0.41	D
BayesDel_noAF	Pathogenic	0.35
Cadd	Pathogenic	29
Dann	Uncertain	1.0
DEOGEN2	Pathogenic	0.86	D;.;T
Eigen	Pathogenic	0.73
Eigen_PC	Pathogenic	0.67
FATHMM_MKL	Uncertain	0.93	D
LIST_S2	Benign	0.85	D;T;T
M_CAP	Pathogenic	0.42	D
MetaRNN	Pathogenic	0.81	D;D;D
MetaSVM	Uncertain	0.37	D
MutationAssessor	Uncertain	2.9	M;M;.
MutationTaster	Benign	1.0	D;D;D;D
PrimateAI	Uncertain	0.70	T
PROVEAN	Uncertain	-3.3	D;D;D
REVEL	Pathogenic	0.74
Sift	Pathogenic	0.0	D;D;D
Sift4G	Uncertain	0.044	D;D;D
Polyphen		0.99	D;.;.
Vest4		0.74
MutPred		0.45		Loss of stability (P = 0.0023);Loss of stability (P = 0.0023);.;
MVP		0.91
MPC		1.5
ClinPred		0.98	D
GERP RS		5.7
Varity_R		0.88
gMVP		0.82

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.0		Details are displayed if max score is > 0.2

Find out detailed SpliceAI scores and Pangolin per-transcript scores at spliceailookup.broadinstitute.org

Publications

LitVar

Other links and lift over

dbSNP: rs397516830; hg19: chr3-12645681;