rs730881003

Positions:

• chr3-12585794-A-G

Variant summary

Our verdict is Pathogenic. Variant got 14 ACMG points: 14P and 0B. PM1 PM2 PP2 PP3 PP5_Very_Strong

The NM_002880.4(RAF1):​c.1423T>C​(p.Phe475Leu) 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 (★★).

• Frequency: Genomes: not found (cov: 32)
• Consequence: RAF1 NM_002880.4 missense
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:6
• 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]

RAF1 (HGNC:):

MKRN2 (HGNC:7113): (makorin ring finger protein 2) This gene encodes a probable E3 ubiquitin ligase containing several zinc finger domains, that is a member of the makorin RING zinc-finger protein family. This gene overlaps the v-raf-1 murine leukemia viral oncogene homolog 1 (RAF1) gene in an antisense orientation and may have a co-regulatory function with RAF1. Alternative splicing results in multiple transcript variants. [provided by RefSeq, Nov 2012]

ACMG classification

Verdict is Pathogenic. Variant got 14 ACMG points.

• PM1: In a domain Protein kinase (size 260) in uniprot entity RAF1_HUMAN there are 6 pathogenic changes around while only 1 benign (86%) in NM_002880.4
• PM2: Very rare variant in population databases, with high coverage
• PP2: Missense variant in gene, where missense usually causes diseases (based on misZ statistic), RAF1. . Gene score misZ 2.4628 (greater than the threshold 3.09). Trascript score misZ 3.4185 (greater than threshold 3.09). GenCC has associacion of gene with Noonan syndrome 5, dilated cardiomyopathy 1NN, Noonan syndrome with multiple lentigines, Noonan syndrome, cardiofaciocutaneous syndrome, familial isolated dilated cardiomyopathy, LEOPARD syndrome 2, Costello syndrome.
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.746
• PP5: Variant 3-12585794-A-G is Pathogenic according to our data. Variant chr3-12585794-A-G is described in ClinVar as [Likely_pathogenic]. Clinvar id is 40617.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
RAF1	NM_002880.4	c.1423T>C	p.Phe475Leu	missense_variant	14/17	ENST00000251849.9	NP_002871.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
RAF1	ENST00000251849.9	c.1423T>C	p.Phe475Leu	missense_variant	14/17	1	NM_002880.4	ENSP00000251849.4

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Cov.: 28

GnomAD4 genome Cov.: 32

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:6

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Noonan syndrome Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Jul 12, 2017	The p.Phe475Leu variant in RAF1 has been reported in 2 individuals with clinical features of Noonan syndrome, was de novo in 1 of those individuals, and segrega ted with disease in 2 affected relatives from 1 family (personal communication, GeneDx). It was absent from large population studies. Computational prediction t ools and conservation analysis suggest that the p.Phe475Leu variant may impact t he protein, though this information is not predictive enough to determine pathog enicity. In summary, although additional studies are required to fully establish its clinical significance, the p.Phe475Leu variant is likely pathogenic. -
Likely pathogenic, no assertion criteria provided	clinical testing	Service de Génétique Moléculaire, Hôpital Robert Debré	-	- -

Dilated cardiomyopathy 1NN Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Institute of Medical Genetics and Applied Genomics, University Hospital Tübingen

Nov 30, 2023

- -

not provided Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

GeneDx

Jun 17, 2013

p.Phe475Leu (TTT>CTT): c.1423 T>C in exon 14 of the RAF1 gene (NM_002880.3). An F475L missense mutation was identified in the RAF1 gene. It has not been published as a mutation, nor has it been reported as a benign polymorphism to our knowledge. However, this mutation has been seen several times previously at GeneDx in one proband and segregating with disease in another unrelated family; all were reported to have phenotypes consistent with the diagnosis of a disorder of the Noonan-CFC-Costello syndrome spectrum. F475L is a semi-conservative amino acid substitution with a non-polar aromatic residue (Phe) being replaced by a non-polar aliphatic residue (Leu). The position at which this mutation occurs is highly conserved across species and is located within 1 of 3 exons where all published mutations have been reported (Pandit et al., 2007 and Razzaque et al., 2007). In silico analysis (PolyPhen) predicts that this sequence variant is probably damaging to protein structure/function. Therefore, F475L is considered to be a disease-causing mutation consistent with the diagnosis. The variant is found in NOONAN panel(s). -

Noonan syndrome 5 Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Victorian Clinical Genetics Services, Murdoch Childrens Research Institute

Dec 21, 2023

Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as Pathogenic. Following criteria are met: 0103 - Loss of function and gain of function are known mechanisms of disease in this gene. Gain of function is associated with Noonan (MIM#611553) and LEOPARD syndromes (MIM#2611554), while loss of function is associated with non-HCM-associated variants (PMIDs: 17603483, 17603482). (I) 0107 - This gene is associated with autosomal dominant disease. (I) 0200 - Variant is predicted to result in a missense amino acid change from phenylalanine to leucine. (I) 0251 - This variant is heterozygous. (I) 0301 - Variant is absent from gnomAD (both v2 and v3). (SP) 0502 - Missense variant with conflicting in silico predictions and uninformative conservation. (I) 0600 - Variant is located in the annotated protein tyrosine and serine/threonine kinase (DECIPHER). (I) 0705 - No comparable missense variants have previous evidence for pathogenicity. (I) 0801 - This variant has strong previous evidence of pathogenicity in unrelated individuals. This variant has been classified as pathogenic or likely pathogenic by clinical laboratories in ClinVar; one of these laboratories reported the variant as de novo. This variant has also been observed in an individual with Noonan syndrome (PMID: 31560489). (SP) 1203 - This variant has been shown to be de novo in the proband (parental status confirmed) (by trio analysis). (SP) -

RASopathy Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Labcorp Genetics (formerly Invitae), Labcorp

Mar 30, 2023

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. 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. ClinVar contains an entry for this variant (Variation ID: 40617). This missense change has been observed in individuals with clinical features of Noonan syndrome (PMID: 31560489; Invitae). This variant is not present in population databases (gnomAD no frequency). This sequence change replaces phenylalanine, which is neutral and non-polar, with leucine, which is neutral and non-polar, at codon 475 of the RAF1 protein (p.Phe475Leu). -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	1.0
BayesDel_addAF	Pathogenic	0.30	D
BayesDel_noAF	Pathogenic	0.19
CADD	Pathogenic	33
DANN	Uncertain	1.0
DEOGEN2	Pathogenic	0.91	D;.;D
Eigen	Uncertain	0.47
Eigen_PC	Uncertain	0.57
FATHMM_MKL	Pathogenic	0.98	D
LIST_S2	Uncertain	0.95	D;D;D
M_CAP	Uncertain	0.13	D
MetaRNN	Pathogenic	0.75	D;D;D
MetaSVM	Benign	-0.34	T
MutationAssessor	Benign	-1.2	N;.;.
PrimateAI	Pathogenic	0.92	D
PROVEAN	Pathogenic	-5.4	D;D;D
REVEL	Pathogenic	0.67
Sift	Uncertain	0.0010	D;D;D
Sift4G	Benign	0.12	T;T;T
Polyphen		0.84	P;.;.
Vest4		0.86
MutPred		0.61		Gain of ubiquitination at K470 (P = 0.1204);.;.;
MVP		0.95
MPC		2.4
ClinPred		1.0	D
GERP RS		5.3
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.7
Varity_R		0.94
gMVP		0.81

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.16		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: rs730881003; hg19: chr3-12627293;