ClinVar Genomic variation as it relates to human health

The c.256_257del (p.Lys86ValfsTer33) variant in RAG1 is a frameshift variant that may cause a premature stop codon that is predicted to escape nonsense-mediated decay. The variant causes a truncation of a functionally important region (removes amino acids 86-1011) of the RAG1 protein. The variants also cause the usage of an alternative in-frame start codon (Met183), but the alternatively transcribed protein has decreased recombination activity and mislocalization in cells (PMIDs 11121059, 27301863). So the variant may cause protein loss-of-function (PVS1; PMIDs 11121059, 27301863). The filtering allele frequency based on the European (non-Finnish) population (upper bound of 95% CI of 6/128846 observed alleles) is 0.000007030 in gnomAD v2.1.1 which is below the SCID-VCEP threshold (<0.000102) and therefore meets this criterion (PM2_Supporting). The in vitro recombination assay shows that the p.Lys86ValfsTer33 variant in RAG1 causes a decrease of recombination activity to below 25%, indicating that the variant impacts the protein function. (PS3_Moderate; PMID 24290284). One homozygous individual with OS. One homozygous individual with SCID. 1pt for PM3. (PMID 11121059, patient OS8; PMID 32655540, patient #32). PM3 met. One patient with this variant was diagnosed with SCID (0.5pt). The patient showed T-B-NK+ lymphocyte profile (0.5pt). 1pt in total, PP4 met. (PMID 32655540, proband #32) In summary, this variant meets the criteria to be classified as Pathogenic for autosomal recessive SCID based on the ACMG/AMP criteria applied, as specified by the ClinGen SCID VCEP. Criteria applied: PVS1, PM2_supporting, PM3_moderate, PS3_moderate, and PP4_supporting. (VCEP specifications version 1).

The RAG1 c.256_257delAA (p.Lys86ValfsTer33) variant is a frameshift variant that is predicted to result in premature termination of the protein. The p.Lys86ValfsTer33 variant has been reported in five studies in which it is found in a total of 14 individuals with RAG1-related disorders, including in eight in a homozygous state, five in a compound heterozygous state, and one in a heterozygous state (Abraham et al. 2013; Lee et al. 2014; Buchbinder et al. 2015; Walter et al. 2015; Brauer et al. 2016). Homozygous individuals included six with severe combined immune deficiency (SCID) and two with Omenn syndrome (OS); compound heterozygous individuals included two with OS, one with common variable immunodeficiency disorder (CVID), two with combined immune deficiency with granuloma and/or autoimmunity (CID-G/A); the heterozygote had an unspecified immunodeficiency. Control data are unavailable for this variant, which is reported at a frequency of 0.00008 in the European (non-Finnish) population of the Exome Aggregation Consortium. The p.Lys86ValfsTer33 variant protein was found to have approximately three percent of wild type RAG1 recombinase activity when analyzed in pro-B cells that were deficient for wild type RAG1 (Abraham et al. 2013; Buchbinder et al. 2015; Brauer et al. 2016). Based on the evidence and potential impact of frameshift variants, the p.Lys86ValfsTer33 variant is classified as pathogenic for RAG1-related disorders. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population.

This sequence change creates a premature translational stop signal (p.Lys86Valfs*33) in the RAG1 gene. While this is not anticipated to result in nonsense mediated decay, it is expected to disrupt the last 958 amino acid(s) of the RAG1 protein. This variant is present in population databases (rs772962160, gnomAD 0.006%). This premature translational stop signal has been observed in individuals with Omenn syndrome or severe combined immunodeficiency (PMID: 9630231, 22424479, 23085344, 24290284, 25516070). ClinVar contains an entry for this variant (Variation ID: 285045). Algorithms developed to predict the effect of variants on protein structure and function are not available or were not evaluated for this variant. Experimental studies have shown that this premature translational stop signal affects RAG1 function (PMID: 10891452, 24418478, 25516070). For these reasons, this variant has been classified as Pathogenic.

Variant summary: RAG1 c.256_257delAA (p.Lys86ValfsX33) results in a premature termination codon, predicted to cause a truncation of the encoded protein or absence of the protein due to nonsense mediated decay, which are commonly known mechanisms for disease. Truncations downstream of this position have been classified as pathogenic by our laboratory. The variant allele was found at a frequency of 1.2e-05 in 251098 control chromosomes. c.256_257delAA has been reported in the literature in multiple individuals affected with Severe Combined Immunodeficiency Syndrome/Omenn Syndrome. Four clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. All laboratories classified the variant as pathogenic. Based on the evidence outlined above, the variant was classified as pathogenic.

The RAG1 c.256_257delAA variant is predicted to result in a frameshift and premature protein termination (p.Lys86Valfs*33). This variant has been reported as causative for autosomal recessive Omenn syndrome, as well as sever combined immunodeficiency syndrome (Table 1, referred to as deletion 368-369, Villa et al. 1998. PubMed ID: 9630231; Table 2, Kutukculer et al. 2012. PubMed ID: 22424479; Sharapova et al. 2013. PubMed ID: 23085344; IJspeert et al. 2014. PubMed ID: 24418478; Buchbinder et al. 2015. PubMed ID: 25516070). It is reported to be a founder mutation in Slavic populations that originated in Poland (Sharapova et al 2020. PubMed ID: 32655540). Experimental studies indicated this variant reduces RAG1 activity compared to wildtype (referred to as c.368_369delAA, Figure 1, IJspeert et al. 2014. PubMed ID: 24418478). This variant is reported in 0.0047% of alleles in individuals of European (Non-Finnish) descent in gnomAD (http://gnomad.broadinstitute.org/variant/11-36595109-TAA-T). Frameshift variants in RAG1 are expected to be pathogenic. This variant is interpreted as pathogenic.