chr3-12604204-T-C

Variant names:

• chr3-12604204-T-C
• rs397516825
• NM_002880.4:c.766A>G

Variant summary

Our verdict is Pathogenic. The variant received 14 ACMG points: 14P and 0B. PM1 PM2 PM5 PP5_Very_Strong

The NM_002880.4(RAF1):​c.766A>G​(p.Arg256Gly) variant causes a missense change involving the alteration of a non-conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. 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. R256S) has been classified as Likely pathogenic.

• Frequency: Genomes: not found (cov: 33)
• Consequence: RAF1 NM_002880.4 missense
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:4 U:1
• Conservation: PhyloP100: 1.97
• Publications: 3 publications found

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]

RAF1 Gene-Disease associations (from GenCC):

• Noonan syndrome

  Inheritance: AD Classification: DEFINITIVE, SUPPORTIVE Submitted by: Orphanet, ClinGen
• Noonan syndrome 5

  Inheritance: AD Classification: DEFINITIVE, STRONG Submitted by: Genomics England PanelApp, Labcorp Genetics (formerly Invitae), Ambry Genetics, PanelApp Australia, G2P
• dilated cardiomyopathy 1NN

  Inheritance: AD Classification: STRONG, LIMITED Submitted by: Labcorp Genetics (formerly Invitae), Ambry Genetics
• LEOPARD syndrome 2

  Inheritance: AD Classification: STRONG Submitted by: Genomics England PanelApp, Labcorp Genetics (formerly Invitae)
• familial isolated dilated cardiomyopathy

  Inheritance: AD Classification: SUPPORTIVE Submitted by: Orphanet
• Noonan syndrome with multiple lentigines

  Inheritance: AD Classification: SUPPORTIVE, LIMITED Submitted by: Orphanet, ClinGen
• cardiofaciocutaneous syndrome

  Inheritance: AD Classification: NO_KNOWN Submitted by: ClinGen
• Costello syndrome

  Inheritance: AD Classification: NO_KNOWN Submitted by: ClinGen

ACMG classification

Our verdict: Pathogenic. The variant received 14 ACMG points.

• PM1: In a hotspot region, there are 8 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 chr3-12604202-C-A is described in ClinVar as [Likely_pathogenic]. Clinvar id is 40599.Status of the report is reviewed_by_expert_panel, 3 stars.
• PP5: Variant 3-12604204-T-C is Pathogenic according to our data. Variant chr3-12604204-T-C is described in ClinVar as [Likely_pathogenic]. Clinvar id is 44631.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	MANE	Protein	UniProt
RAF1	NM_002880.4	c.766A>G	p.Arg256Gly	missense_variant	Exon 7 of 17	ENST00000251849.9	NP_002871.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	Exon rank	TSL	MANE	Protein	Appris	UniProt
RAF1	ENST00000251849.9	c.766A>G	p.Arg256Gly	missense_variant	Exon 7 of 17	1	NM_002880.4	ENSP00000251849.4

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 33

Alfa AF: 0.00 Hom.: 0

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:4 Uncertain:1

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Noonan syndrome Pathogenic:1 Uncertain:1

May 10, 2012

Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The Arg256Gly variant in RAF1 has not been reported in the literature nor previo usly identified by our laboratory. However, a different amino acid change at thi s position (Arg256Ser) has been identified in individuals with clinical features of Noonan syndrome, suggesting that a change to this position may not be tolera ted (Pandit 2007). Computational analyses (biochemical amino acid properties, co nservation, AlignGVGD, PolyPhen2, and SIFT) suggest that the Arg256Gly variant m ay impact the protein, though this information is not predictive enough to deter mine pathogenicity. In summary, this variant is likely to be pathogenic, though segregation studies and functional analyses are required to fully establish the pathogenicity of this variant. Individuals with pathogenic variants in exon 7 or 17 in RAF1 have a high incidence of hypertrophic cardiomyopathy (80-95%, Razzaq ue 2007, Pandit 2007). The presence of a heterozygous pathogenic variant in RAF1 is consistent with a diagnosis of Noonan syndrome but this information should b e reconciled with the complete clinical history of this individual. -

-

Service de Génétique Moléculaire, Hôpital Robert Debré

Significance: Uncertain significance

Review Status: no assertion criteria provided

Collection Method: clinical testing

- -

Noonan syndrome 1 Pathogenic:1

-

Molecular Genetics, Centre for Human Genetics

Significance: Likely pathogenic

Review Status: no assertion criteria provided

Collection Method: clinical testing

- -

not provided Pathogenic:1

Jan 13, 2022

ARUP Laboratories, Molecular Genetics and Genomics, ARUP Laboratories

Significance: Likely pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

The RAF1 c.766A>G; p.Arg256Gly variant (rs397516825) is reported in the literature in an individual with hypertrophic cardiomyopathy (Burstein 2021) and is reported as occurring de novo in an individual described in the ClinVar database (Variation ID: 44631). This variant is absent from general population databases (Exome Variant Server, Genome Aggregation Database), indicating it is not a common polymorphism. The arginine at codon 256 is highly conserved, but computational analyses are uncertain whether this variant is neutral or deleterious (REVEL: 0.648). However, this amino acid occurs in the conserved CR2 domain, involved in 14-3-3 binding, and variants in this region cause altered phosphorylation of the RAF1 protein (Kobayashi 2010). Several variants in the CR2 domain have been described in affected individuals (Kobayashi 2010, Pandit 2007). Additionally, another variant in this codon, c.768G>T; p.Arg256Ser, is described in affected individuals (Pandit 2007) and is considered pathogenic by a ClinVar expert panel (Variation ID: 40599). Based on available information, this variant is considered to be likely pathogenic. References: Burstein DS et al. Genetic variant burden and adverse outcomes in pediatric cardiomyopathy. Pediatr Res. 2021 May;89(6):1470-1476. PMID: 32746448. Kobayashi T et al. Molecular and clinical analysis of RAF1 in Noonan syndrome and related disorders: dephosphorylation of serine 259 as the essential mechanism for mutant activation. Hum Mutat. 2010 Mar;31(3):284-94. PMID: 20052757. Pandit B et al. Gain-of-function RAF1 mutations cause Noonan and LEOPARD syndromes with hypertrophic cardiomyopathy. Nat Genet. 2007 Aug;39(8):1007-12. PMID: 17603483. -

RASopathy Pathogenic:1

Apr 27, 2023

Labcorp Genetics (formerly Invitae), Labcorp

Significance: Pathogenic

Review Status: criteria provided, single submitter

Collection Method: clinical testing

This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individual(s) with hypertrophic cardiomyopathy and Noonan syndrome (Invitae). In at least one individual the variant was observed to be de novo. ClinVar contains an entry for this variant (Variation ID: 44631). 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. For these reasons, this variant has been classified as Pathogenic. This sequence change replaces arginine, which is basic and polar, with glycine, which is neutral and non-polar, at codon 256 of the RAF1 protein (p.Arg256Gly). -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.97
BayesDel_addAF	Pathogenic	0.35	D
BayesDel_noAF	Pathogenic	0.26
CADD	Pathogenic	26
DANN	Uncertain	1.0
DEOGEN2	Pathogenic	0.88	D;.;T
Eigen	Uncertain	0.39
Eigen_PC	Uncertain	0.30
FATHMM_MKL	Benign	0.60	D
LIST_S2	Uncertain	0.96	D;D;D
M_CAP	Pathogenic	0.34	D
MetaRNN	Uncertain	0.64	D;D;D
MetaSVM	Uncertain	0.25	D
MutationAssessor	Pathogenic	3.0	M;M;.
PhyloP100		2.0
PrimateAI	Uncertain	0.76	T
PROVEAN	Uncertain	-3.9	D;D;D
REVEL	Pathogenic	0.65
Sift	Uncertain	0.0010	D;D;D
Sift4G	Uncertain	0.0050	D;D;D
Polyphen		1.0	D;.;.
Vest4		0.81
MutPred		0.27		Gain of glycosylation at S252 (P = 0.0093);Gain of glycosylation at S252 (P = 0.0093);.;
MVP		0.94
MPC		1.5
ClinPred		0.99	D
GERP RS		3.2
PromoterAI		-0.027	Neutral
Varity_R		0.86
gMVP		0.82
Mutation Taster		=3/97	disease causing (ClinVar)

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

Other links and lift over

dbSNP: rs397516825; hg19: chr3-12645703;