rs774119942

chr19-38510658-A-G

Variant summary

Our verdict is Likely pathogenic. Variant got 9 ACMG points: 9P and 0B. PVS1_Moderate PM2 PP3_Strong PP5

The NM_000540.3(RYR1):c.9001-2A>G variant causes a splice acceptor change involving the alteration of a conserved nucleotide. The variant allele was found at a frequency of 0.00000274 in 1,461,890 control chromosomes in the GnomAD database, with no homozygous occurrence. In-silico tool predicts a pathogenic outcome for this variant. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Conflicting classifications of pathogenicity (no stars).

• Frequency:
  • Genomes: not found (cov: 32)
  • Exomes 𝑓: 0.0000027 (0 hom.)
• Consequence: RYR1 NM_000540.3 splice_acceptor
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:4 U:1
• Conservation: PhyloP100: 9.29

Genes affected

RYR1 (HGNC:10483): (ryanodine receptor 1) This gene encodes a ryanodine receptor found in skeletal muscle. The encoded protein functions as a calcium release channel in the sarcoplasmic reticulum but also serves to connect the sarcoplasmic reticulum and transverse tubule. Mutations in this gene are associated with malignant hyperthermia susceptibility, central core disease, and minicore myopathy with external ophthalmoplegia. Alternatively spliced transcripts encoding different isoforms have been described. [provided by RefSeq, Jul 2008]

ACMG classification

Verdict is Likely_pathogenic. Variant got 9 ACMG points.

• PVS1: Splicing variant, NOT destroyed by nmd, known LOF gene, truncates exone, which is 0.008004233 fraction of the gene. Cryptic splice site detected, with MaxEntScore 4.7, offset of 24, new splice context is: tgctccctttgatcaaccAGtac. Cryptic site results in inframe change. If cryptic site found is not functional and variant results in exon loss, it results in frameshift change.
• PM2: Very rare variant in population databases, with high coverage
• PP3: Splicing scoreres supports a deletorius effect: Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF, max_spliceai. No scorers claiming Uncertain. No scorers claiming Benign.
• PP5: Variant 19-38510658-A-G is Pathogenic according to our data. Variant chr19-38510658-A-G is described in ClinVar as [Conflicting_classifications_of_pathogenicity]. Clinvar id is 590624.We mark this variant Likely_pathogenic, oryginal submissions are: {Pathogenic=2, Likely_pathogenic=2, Uncertain_significance=1}.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
RYR1	NM_000540.3	c.9001-2A>G		splice_acceptor_variant		ENST00000359596.8

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
RYR1	ENST00000359596.8	c.9001-2A>G		splice_acceptor_variant		5	NM_000540.3	A2
RYR1	ENST00000355481.8	c.9001-2A>G		splice_acceptor_variant		1		P4
RYR1	ENST00000594335.5	c.2453-2A>G		splice_acceptor_variant, NMD_transcript_variant		1
RYR1	ENST00000599547.6	c.9001-2A>G		splice_acceptor_variant, NMD_transcript_variant		2

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD3 exomes AF: 0.00000398 AC: 1 AN: 251484 Hom.: 0 AF XY: 0.00000736 AC XY: 1 AN XY: 135914

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.00

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.00

Gnomad SAS exome AF: 0.00

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.00000879

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.00000274 AC: 4 AN: 1461890 Hom.: 0 Cov.: 32 AF XY: 0.00000550 AC XY: 4 AN XY: 727248

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.00

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.00

Gnomad4 SAS exome AF: 0.00

Gnomad4 FIN exome AF: 0.00

Gnomad4 NFE exome AF: 0.00000360

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome Cov.: 32

ExAC AF: 0.00000824 AC: 1

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Pathogenic:4 Uncertain:1

Revision: criteria provided, conflicting classifications

Submissions by phenotype

RYR1-related disorder Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Jul 14, 2023	Variant summary: RYR1 c.9001-2A>G is located in a canonical splice-site and is predicted to affect mRNA splicing resulting in a significantly altered protein due to either exon skipping, shortening, or inclusion of intronic material. Several computational tools predict a significant impact on normal splicing: Four predict the variant abolishes a 3' acceptor site. However, these predictions have yet to be confirmed by functional studies. The variant allele was found at a frequency of 4e-06 in 251484 control chromosomes (i.e., 1 heterozygote; gnomAD v2.1, Exomes dataset). c.9001-2A>G has been reported in the literature in at least one compound heterozygous individual affected with autosomal recessive Central Core Disease and malignant hyperthermia during anesthesia (e.g., Snoeck_2015, Todd_2018, Lawal_2021). These data suggest the variant is likely to be associated with disease. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. The following publications have been ascertained in the context of this evaluation (PMID: 25960145, 30155738, 33646171). Three submitters have reported clinical-significance assessments for this variant to ClinVar after 2014. All submitters classified the variant as pathogenic/likely pathogenic. Based on the evidence outlined above, the variant was classified as likely pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Invitae	Jul 11, 2023	Algorithms developed to predict the effect of sequence changes on RNA splicing suggest that this variant may disrupt the consensus splice site. This sequence change affects an acceptor splice site in intron 59 of the RYR1 gene. It is expected to disrupt RNA splicing. Variants that disrupt the donor or acceptor splice site typically lead to a loss of protein function (PMID: 16199547), and loss-of-function variants in RYR1 are known to be pathogenic (PMID: 23919265, 25960145, 28818389, 30611313). This variant is present in population databases (rs774119942, gnomAD 0.0009%). Disruption of this splice site has been observed in individual(s) with RYR1-associated conditions (PMID: 25960145). In at least one individual the data is consistent with being in trans (on the opposite chromosome) from a pathogenic variant. ClinVar contains an entry for this variant (Variation ID: 590624). For these reasons, this variant has been classified as Pathogenic. -

not provided Pathogenic:2

Pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Nov 30, 2022	Canonical splice site variant predicted to result in a null allele in a gene for which loss of function is a known mechanism of disease; Not observed at significant frequency in large population cohorts (gnomAD); This variant is associated with the following publications: (PMID: 33646171, 25960145) -
Likely pathogenic, criteria provided, single submitter	clinical testing	PreventionGenetics, part of Exact Sciences	Aug 25, 2016	- -

Malignant hyperthermia, susceptibility to, 1 Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

All of Us Research Program, National Institutes of Health

Dec 13, 2023

This variant causes a A>G nucleotide substitution at the -2 position of intron 59 of the RYR1 gene. Splicing prediction tools indicate that this variant may disrupt RNA splicing. To our knowledge, functional studies have not been reported for this variant. This variant has been reported in at least two individuals affected with an RYR1-related myopathy who had a personal or family history of a malignant hyperthermia event (PMID: 25960145, 30155738, 32236737). All of these individuals were reported to carry a different pathogenic variant in the RYR1 gene in the compound heterozygous state or in unknown phase that could explain the observed malignant hyperthermia susceptibility phenotype (PMID: 25960145, 30155738, 32236737). This variant has been identified in 1/251484 chromosomes in the general population by the Genome Aggregation Database (gnomAD). Loss of RYR1 function due to truncation and splice variants is not an established disease mechanism for autosomal dominant malignant hyperthermia, although it is associated with other phenotype(s) (ClinVar variation ID: 590624). Due to insufficient evidence, this variant is classified as a Variant of Uncertain Significance for autosomal dominant malignant hyperthermia. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Pathogenic	0.35	D
BayesDel_noAF	Pathogenic	0.27
Cadd	Pathogenic	34
Dann	Uncertain	1.0
Eigen	Pathogenic	1.1
Eigen_PC	Pathogenic	0.90
FATHMM_MKL	Pathogenic	1.0	D
MutationTaster	Benign	1.0	D;D;D
GERP RS		4.7

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.94
SpliceAI score (max)		0.99		Details are displayed if max score is > 0.2
DS_AG_spliceai		0.66		Position offset: 26
DS_AL_spliceai		0.99		Position offset: 2

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

Publications

LitVar

Other links and lift over

dbSNP: rs774119942; hg19: chr19-39001298;