rs730880382

chr3-12591708-C-Achr3-12591708-C-Gchr3-12591708-C-T

Variant summary

Our verdict is Pathogenic. Variant got 11 ACMG points: 11P and 0B. PM1 PM2 PP2 PP3_Moderate PP5_Strong

The NM_002880.4(RAF1):c.1193G>T(p.Arg398Leu) variant causes a missense, splice region 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. 13/23 in silico tools predict a damaging outcome for this variant. 2/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Uncertain significance (★★★). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. R398C) has been classified as Uncertain significance.

• Frequency: Genomes: not found (cov: 32)
• Consequence: RAF1 NM_002880.4 missense, splice_region
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Uncertain significance reviewed by expert panel P:2 U:4
• Conservation: PhyloP100: 7.90

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 11 ACMG points.

• PM1: In a domain Protein kinase (size 260) in uniprot entity RAF1_HUMAN there are 17 pathogenic changes around while only 4 benign (81%) in NM_002880.4
• PM2: Very rare variant in population databases, with high coverage
• PP2: Missense variant where missense usually causes diseases, RAF1
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.912
• PP5: Variant 3-12591708-C-A is Pathogenic according to our data. Variant chr3-12591708-C-A is described in ClinVar as [Uncertain_significance]. Clinvar id is 40614.Status of the report is reviewed_by_expert_panel, 3 stars. We mark this variant Likely_pathogenic, oryginal submissions are: {Uncertain_significance=3, Likely_pathogenic=2}.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
RAF1	NM_002880.4	c.1193G>T	p.Arg398Leu	missense_variant, splice_region_variant	11/17	ENST00000251849.9

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
RAF1	ENST00000251849.9	c.1193G>T	p.Arg398Leu	missense_variant, splice_region_variant	11/17	1	NM_002880.4	P3

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 32

ClinVar

Significance: Uncertain significance

Submissions summary: Pathogenic:2 Uncertain:4

Revision: reviewed by expert panel

Submissions by phenotype

RASopathy Pathogenic:1 Uncertain:1

Likely pathogenic, criteria provided, single submitter	clinical testing	Invitae	Jul 28, 2023	This sequence change replaces arginine, which is basic and polar, with leucine, which is neutral and non-polar, at codon 398 of the RAF1 protein (p.Arg398Leu). This variant also falls at the last nucleotide of exon 11, which is part of the consensus splice site for this exon. This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individuals with clinical features of RAF1-related RASopathy and/or clinical features of RASopathy spectrum disorders (Invitae). ClinVar contains an entry for this variant (Variation ID: 40614). An algorithm developed to predict the effect of missense changes on protein structure and function (PolyPhen-2) suggests that this variant is likely to be disruptive. Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). In summary, the currently available evidence indicates that the variant is pathogenic, but additional data are needed to prove that conclusively. Therefore, this variant has been classified as Likely Pathogenic. -
Uncertain significance, reviewed by expert panel	curation	ClinGen RASopathy Variant Curation Expert Panel	Dec 05, 2019	The c.1193G>T (p.Arg398Leu) variant has been identified in at least 4 probands with variable phenotypic features with two of them having a clinical suspicion of Noonan syndrome (PS4 not met; Rady Children's Institute for Genomic Medicine internal communications; SCV000209025.5; SCV000207671.1; SCV000552095.2). This variant was absent from large population studies (PM2). 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; however, it is in a region that generally lacks pathogenic variants (PP2; PMID: 29493581). Computational prediction tools and conservation analysis suggest that the p.Arg398Leu variant may impact the protein (PP3). In summary, the clinical significance of the p.Arg398Leu variant is uncertain. RASopathy-specific ACMG/AMP criteria applied (PMID:29493581): PM2, PP2, PP3. -

not provided Uncertain:2

Uncertain significance, criteria provided, single submitter	clinical testing	GeneDx	May 04, 2017	The c.1193 G>T variant in the RAF1 gene has not been reported previously as a pathogenic variant, nor as a benign variant, to our knowledge. The c.1193 G>T variant is not observed in large population cohorts (Lek et al., 2016; 1000 Genomes Consortium et al., 2015; Exome Variant Server). In-silico splice prediction models predict that c.1193 G>T damages the splice donor site in intron 11, which may cause abnormal gene splicing. However, in the absence of RNA/functional studies, the actual effect of the c.1193 G>T change in this individual is unknown. If c.1193 G>T does not alter splicing, it will result in the R398L missense change. The R398L variant is a non-conservative amino acid substitution, which is likely to impact secondary protein structure as these residues differ in polarity, charge, size and/or other properties. While this substitution occurs at a position that is not conserved, in silico analysis predicts this variant is probably damaging to the protein structure/function. In vitro functional studies indicate that another missense substitution (R398A) at this residue leads to reduction of RAF1 kinase activity (Baljuls et al., 2011); however, increased activity is expected for RASopathy disorders. We interpret c.1193 G>T as a variant of uncertain significance. -
Uncertain significance, no assertion criteria provided	clinical testing	Greenwood Genetic Center Diagnostic Laboratories, Greenwood Genetic Center	Jan 15, 2015	- -

Noonan syndrome 5 Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Rady Children's Institute for Genomic Medicine, Rady Children's Hospital San Diego

Nov 07, 2019

This variant has been reported in the literature (PMID: 30095857, 29493581) and is present in ClinVar (Variation ID: 49084). The residue p.Arg398 has been characterized in-vitro as functionally important (PMID: 21454547, 10801816, 12925535). The c.1193G>T, p.Arg398Leu is absent from the ExAC and gnomAD population databases and thus is presumed to be rare. This alteration affect the last nucleotide of exon 11 of RAF1 and in-silico splicing algorithms (SpliceSiteFinder-like, MaxEntScan, NNSplice, GeneSplicer) predict that the variant weakens the canonical splice site. In addition, in-silico analyses for the mssense change support a deleterious effect of this variant on protein function. Analysis of the parental samples was negative for the variant, indicating this variant likely occurred as a de novo event. Based on the available evidence, the c.1193G>T, p.Arg398Leu variant is classified as Likely Pathogenic. -

LEOPARD syndrome 2;C1969057:Noonan syndrome 5;C4014656:Dilated cardiomyopathy 1NN Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Center for Genomics, Ann and Robert H. Lurie Children's Hospital of Chicago

Dec 15, 2021

RAF1 NM_002880.3 exon 11 p.Arg398Leu (c.1193G>T): This variant has not been reported in the literature in association with disease and is not present in large control databases. This variant is present in ClinVar with classifications ranging from Uncertain significance to likely pathogenic, including as a Uncertain significance by the ClinGen RASopathy Variant Curation Expert Panel (Variation ID:40614). Evolutionary conservation and computational predictive tools suggest that this variant may impact the protein. RAF1 has a low rate of benign missense variation, and pathogenic missense variants are enriched in affected individuals; however, this variant occurs in a region in which pathogenic variants are uncommon (Gelb 2018 PMID: 29493581). Of note, this variant occurs in the splice region and computational splice prediction tools support a possible impact to splicing; however, further studies are needed to understand its impact. In summary, data on this variant is insufficient for disease classification. Therefore, the clinical significance of this variant is uncertain. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	1.0
BayesDel_addAF	Pathogenic	0.40	D
BayesDel_noAF	Pathogenic	0.34
Cadd	Pathogenic	35
Dann	Uncertain	1.0
DEOGEN2	Pathogenic	0.92	D;.;D
Eigen	Pathogenic	0.75
Eigen_PC	Pathogenic	0.76
FATHMM_MKL	Pathogenic	1.0	D
LIST_S2	Pathogenic	0.99	D;D;D
M_CAP	Pathogenic	0.29	D
MetaRNN	Pathogenic	0.91	D;D;D
MetaSVM	Uncertain	0.15	D
MutationAssessor	Benign	0.85	L;.;.
MutationTaster	Benign	1.0	D;D;D;D
PrimateAI	Pathogenic	0.85	D
PROVEAN	Pathogenic	-6.4	D;D;D
REVEL	Pathogenic	0.86
Sift	Uncertain	0.0010	D;D;D
Sift4G	Uncertain	0.0020	D;D;D
Polyphen		1.0	D;.;.
Vest4		0.92
MutPred		0.73		Loss of MoRF binding (P = 0.0144);.;.;
MVP		0.98
MPC		2.6
ClinPred		1.0	D
GERP RS		5.2
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.7
Varity_R		0.95
gMVP		0.94

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.97
SpliceAI score (max)		0.080		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: rs730880382; hg19: chr3-12633207; COSMIC: COSV52583001;