3-12604195-A-G

Position:

• chr3-12604195-A-G

Variant summary

Our verdict is Pathogenic. Variant got 15 ACMG points: 15P and 0B. PM1 PM2 PM5 PP2 PP5_Very_Strong

The NM_002880.4(RAF1):​c.775T>C​(p.Ser259Pro) variant causes a missense change involving the alteration of a 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. S259F) has been classified as 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
• 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:):

ACMG classification

Verdict is Pathogenic. Variant got 15 ACMG points.

• PM1: In a compositionally_biased_region Polar residues (size 47) in uniprot entity RAF1_HUMAN there are 14 pathogenic changes around while only 0 benign (100%) 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 chr3-12604194-G-A is described in Lovd as [Pathogenic].
• 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.
• PP5: Variant 3-12604195-A-G is Pathogenic according to our data. Variant chr3-12604195-A-G is described in ClinVar as [Likely_pathogenic]. Clinvar id is 228288.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars. Variant chr3-12604195-A-G is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
RAF1	NM_002880.4	c.775T>C	p.Ser259Pro	missense_variant	7/17	ENST00000251849.9	NP_002871.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
RAF1	ENST00000251849.9	c.775T>C	p.Ser259Pro	missense_variant	7/17	1	NM_002880.4	ENSP00000251849.4

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 33

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:4

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	Oct 08, 2015	The p.Ser259Pro variant in RAF1 has been previously identified as a de novo vari ant in 1 fetus with abnormal ultrasound findings, including cystic hygroma and C HD (Croonen 2013). It was absent from large population studies. In addition, two different amino acid changes at this position (p.Ser259Phe, p.Ser259Tyr) have b een identified in multiple individuals with Noonan spectrum disorders, suggestin g a change at this position may not be tolerated (Pandit 2007, Kobayashi 2009, L MM unpublished data). In vitro functional studies suggest that decreased phospho rylation of the wildtype Serine (Ser) at position 259 may impact protein functio n (Kobayashi 2009); however, these types of assays may not accurately represent biological function. Computational prediction tools and conservation analysis do not provide strong support for or against an impact to the protein. In summary, although additional studies are required to fully establish its clinical signif icance, the p.Ser259Pro variant is likely pathogenic. -
Likely pathogenic, no assertion criteria provided	clinical testing	Service de Génétique Moléculaire, Hôpital Robert Debré	-	- -

RASopathy Pathogenic:2

Pathogenic, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Mar 31, 2021	This variant is not present in population databases (ExAC no frequency). This sequence change replaces serine with proline at codon 259 of the RAF1 protein (p.Ser259Pro). The serine residue is highly conserved and there is a moderate physicochemical difference between serine and proline. This variant has been observed in individual(s) with Noonan syndrome (PMID: 23321623). In at least one individual the variant was observed to be de novo. ClinVar contains an entry for this variant (Variation ID: 228288). For these reasons, this variant has been classified as Pathogenic. This variant disrupts the p.Ser259 amino acid residue in RAF1. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 19020799, 17603483, 20052757, 22465605, Invitae). This suggests that this residue is clinically significant, and that variants that disrupt this residue are likely to be disease-causing. 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. -
Likely pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Jan 29, 2018	Variant summary: RAF1 c.775T>C (p.Ser259Pro) located in the conserved region 2 (CR2) domain causes an alteration of a conserved nucleotide that alters a phosphorylation site (Kobayashi_2010). Four of five in-silico tools predict a damaging effect of the variant on protein function. Functional studies, Kobayashi_2010 and Pandit_2007, indicate that Ser259 is a phosphorylation site that plays a key role in RAF1 regulation, although the variant of interest was not directly evaluated. Other variants affecting codon 259 have been reported as Pathogneic/Likely Pathogenic in ClinVar (p.Ser259Phe, Ser259Thr), further evidence of the importance of this codon. The variant was absent in 246244 control chromosomes (gnomAD). Multiple publications, Bhoj_202016 and Cronnen_2013, cite the variant in Noonan patients including one being indicated as a de novo event. A clinical diagnostic laboratory (evaluation after 2014) classified the variant as "likely pathogenic." Based on the evidence outlined above, the variant was classified as likely pathogenic. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.99
BayesDel_addAF	Pathogenic	0.38	D
BayesDel_noAF	Pathogenic	0.31
CADD	Pathogenic	26
DANN	Uncertain	0.99
DEOGEN2	Uncertain	0.77	D;.;T
Eigen	Uncertain	0.53
Eigen_PC	Uncertain	0.55
FATHMM_MKL	Uncertain	0.93	D
LIST_S2	Pathogenic	0.99	D;D;D
M_CAP	Uncertain	0.27	D
MetaRNN	Uncertain	0.71	D;D;D
MetaSVM	Uncertain	0.40	D
MutationAssessor	Uncertain	2.9	M;M;.
PrimateAI	Pathogenic	0.81	D
PROVEAN	Uncertain	-2.9	D;D;D
REVEL	Pathogenic	0.70
Sift	Uncertain	0.0010	D;D;D
Sift4G	Uncertain	0.0040	D;D;D
Polyphen		0.20	B;.;.
Vest4		0.81
MutPred		0.40		Gain of glycosylation at S257 (P = 0.0137);Gain of glycosylation at S257 (P = 0.0137);.;
MVP		0.91
MPC		0.83
ClinPred		0.96	D
GERP RS		5.7
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.7
Varity_R		0.72
gMVP		0.76

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: rs3730271; hg19: chr3-12645694; COSMIC: COSV52577994; COSMIC: COSV52577994;