Overview

Clinical Phenotype: Alzheimer's Disease, Multiple Conditions
Position: (GRCh38/hg38):Chr19:44909101 C>G
Position: (GRCh37/hg19):Chr19:45412358 C>G
Transcript: NM_000041; ENSG00000130203
dbSNP ID: rs267606661
Coding/Non-Coding: Coding
DNA Change: Substitution
Expected RNA Consequence: Substitution
Expected Protein Consequence: Missense
Codon Change: CGC to GGC
Reference Isoform: APOE Isoform 1
Genomic Region: Exon 4

Findings

This rare variant reduces the risk of Alzheimer’s disease (AD) as assessed by a large association study including more than 500,000 individuals of European and admixed European ancestry (June 2022 news, Le Guen et al., 2022). Risk was decreased twofold (odds ratio, 0.44; 95% CI, 0.33-0.59; P = 4.7 × 10^-8) and appears to involve mitigation of the damaging effects of the common C130R (APOE4) allele, as the two variants have been found to be inherited together. As a comparison with the protection afforded by the R176C (APOE2) allele, the odds ratio of R269G on an APOE3/E4 background was similar to that of APOE2/E3, indicating a protective effect similar or even better than that of APOE2. Also of note, the cumulative incidence of AD in carriers trended toward a six-year delay in age at onset, and the population incidence in carriers grew more slowly with age than in non-carriers.

Paradoxically, in a study of more than 100,000 individuals from Denmark, the carriers of this variant had low levels of ApoE in plasma (mean levels in approximately the 20th percentile; p = 9x10^-24), a phenotype that was associated with increased risk of dementia and of AD in particular (Rasmussen et al., 2020). The association emerged after integrating data from nine APOE variants, including R269G. The frequency of dementia, including AD, amongst older carriers in this study (at least 60 years old), however, was similar to that of non-carriers in the same age group. Also of note, in a subsequent study, this group found an increased risk for ischemic cerebrovascular disease in carriers of rare ApoE variants who had low ApoE levels in plasma (Rasmussen et al., 2023).

R269G’s global frequency in the gnomAD database was 0.00036, including 83 heterozygotes (gnomAD v2.1.1, June 2022). Most of these carriers were of European or Latino/admixed American ancestry.

Non-neurological conditions

This variant was first described in a 51-year-old Caucasian man diagnosed with hyperlipoproteinemia type IV (HLPP4) who had elevated levels of plasma cholesterol and triglycerides (van den Maagdenberg et al., 1993). The carrier’s ApoE proteins migrated on an isoelectric focusing gel to the known positions of the common ApoE3 and ApoE4 isoforms, but upon cysteamine treatment, which adds a positive charge to cysteine residues, the proteins behaved unexpectedly. Instead of both bands shifting to the ApoE4 position, an ApoE3-migrating band remained. This indicated the presence of a new mutation, leading to the identification of the R269G substitution.

A subsequent family study of this same proband revealed three out of four carriers with elevated plasma triglyceride levels, including one individual who also carried the V254E mutation (Zhao et al., 1994), which, coincidentally, also decreases AD risk.  The fourth carrier, who was considerably leaner than the others, however, had a normal lipid profile.

Consistent with these initial findings, subsequent studies have reported variable blood lipid profiles and conditions associated with this mutation. For example, while the mutation correlated with hyperlipidemia and atherosclerosis in a Canadian family (Kang et al., 1997), two French carriers, a woman and her daughter, had normal lipid blood profiles (Richard et al., 1997). Moreover, in a French cohort of patients with primary dyslipidemia, three carriers had elevated low-density lipoprotein (LDL) cholesterol levels and were diagnosed with autosomal dominant hypercholesterolemia, also known as hyperlipoproteinemia type IIa (HLPP2a), while one had elevated triglyceride levels and was diagnosed with familial combined hyperlipidemia, also known as hyperlipoproteinemia type IIb (HLPP2b) (Abou Khalil et al., 2022). Moreover, one of the carriers with HLPP2a had a strong probability of their condition being due to variants in multiple genes (weighted polygenic risk score in the eighth decile).

In the largest study of R269G carriers, comprising 111 individuals, lipid species including LDL cholesterol, remnant cholesterol, high-density lipoprotein (HDL) cholesterol, and triglycerides, were within normal levels, although as mentioned above, ApoE plasma levels were low (Rasmussen et al., 2020, Rasmussen et al., 2023).

R269G has also been reported in carriers of other rare APOE variants, E31K (Rasmussen et al., 2023), V254E, and G145D. In the latter two cases, the carriers had elevated triglycerides, with the V254E carrier having moderately increased plasma levels at age 24 (Zhao et al., 1994), and the G145D carrier being diagnosed with hypertriglyceridemia at age 10 (Richard et al., 1997).

Biological Effect

The biological effects of this variant are unknown, but R269 is part of two important functional sequences—ApoE’s lipid binding region and one of its homo-oligomerization regions—and it is predicted to play a role in determining ApoE’s global structure. Given that R269G is protective for AD, if it affects lipid binding, one might predict it bolsters ApoE4’s reduced lipid binding capacity, making it more like ApoE2 and ApoE3 (Le Guen et al., 2022). If, on the other hand, R269G confers protection by altering oligomerization, one might predict it decreases this capability to reduce harmful aggregation.

However, how this variant might cause either or both effects is unclear. The substitution of a charged arginine with a nonpolar glycine might be expected to enhance lipid binding (Bu, 2022), but increase, rather than decrease, oligomerization (Le Guen et al., 2022). It is also important to consider the interactions between ApoE’s N- and C-termini, which have yet to be determined and likely depend on multiple factors, including the ApoE isoform (E2, E3, or E4) and the protein’s lipidation status. For example, it is possible that R269G alters ApoE’s global conformation by disrupting a proposed salt bridge between R269 and Q116 (Chen et al., 2011), and/or inserting a charge in the vicinity of other salt bridges, including the R79-E273 bridge, which some have proposed is a unique characteristic of ApoE4. This could alter ApoE’s stability in the lipidated state and make it more likely to aggregate (June 2022 news). On the other hand, a study using FRET and computational simulations failed to identify any long-range interactions involving R269 (Stuchell-Brereton et al., 2023). Differences in predictions may be due to the use of different methods and/or different proteins. For example, whereas Chen and colleagues used an ApoE3-like construct mutated to reduce aggregation (Chen et al., 2011), Stuchell-Brereton and co-workers used dilute ApoE4 (Stuchell-Brereton et al., 2023).

Also of note, the increase in ApoE acidity resulting from the substitution of an arginine with a glycine has been noted as potentially protective of endolysosomal trafficking—a process that can be stalled by the loss of solubility of proteins with isoelectric points close to the pH of early endosomes, such as ApoE4 (Vance et al., 2024).

Information on this variant’s ability to bind cell surface receptors is limited, but very low-density lipoprotein (VLDL) particles isolated from a carrier bound normally to macrophages in vitro (Kang et al., 1997). 

In silico algorithms, including Polyphen, Polyphen2, SIFT, Mutation Taster, and Provean, predicted this variant likely affects protein function (Marduel et al., 2013, Abou Khalil et al., 2022). Its PHRED-scaled CADD score (23.1), which integrates diverse information in silico, was above the commonly used threshold of 20 for predicting functional effects (CADD v.1.6, May 2022).

Nomenclature

When it was first discovered, this variant was named APOE*3(Cys112-->Arg; Arg251-->Gly) because its isoelectric migration pattern was similar to that of ApoE3 (van den Maagdenberg et al., 1993). More recently, to denote its linkage to APOE4, it has sometimes been referred to as APOE4 [R251G] (Le Guen et al., 2022). In both cases, the amino acid was identified by its position in the mature ApoE protein (299 amino acids long).

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References

News Citations

1. Two ApoE Mutations Decrease Risk for Alzheimer's Disease 10 Jun 2022

Mutations Citations

1. APOE C130R (ApoE4)
2. APOE R176C (ApoE2)
3. APOE E31K
4. APOE G145D

Paper Citations

1. Le Guen Y, Belloy ME, Grenier-Boley B, de Rojas I, Castillo-Morales A, Jansen I, Nicolas A, Bellenguez C, Dalmasso C, Küçükali F, Eger SJ, Rasmussen KL, Thomassen JQ, Deleuze JF, He Z, Napolioni V, Amouyel P, Jessen F, Kehoe PG, van Duijn C, Tsolaki M, Sánchez-Juan P, Sleegers K, Ingelsson M, Rossi G, Hiltunen M, Sims R, van der Flier WM, Ramirez A, Andreassen OA, Frikke-Schmidt R, Williams J, Ruiz A, Lambert JC, Greicius MD, Members of the EADB, GR@ACE, DEGESCO, DemGene, GERAD, and EADI Groups, Arosio B, Benussi L, Boland A, Borroni B, Caffarra P, Daian D, Daniele A, Debette S, Dufouil C, Düzel E, Galimberti D, Giedraitis V, Grimmer T, Graff C, Grünblatt E, Hanon O, Hausner L, Heilmann-Heimbach S, Holstege H, Hort J, Jürgen D, Kuulasmaa T, van der Lugt A, Masullo C, Mecocci P, Mehrabian S, de Mendonça A, Moebus S, Nacmias B, Nicolas G, Olaso R, Papenberg G, Parnetti L, Pasquier F, Peters O, Pijnenburg YA, Popp J, Rainero I, Ramakers I, Riedel-Heller S, Scarmeas N, Scheltens P, Scherbaum N, Schneider A, Seripa D, Soininen H, Solfrizzi V, Spalletta G, Squassina A, van Swieten J, Tegos TJ, Tremolizzo L, Verhey F, Vyhnalek M, Wiltfang J, Boada M, García-González P, Puerta R, Real LM, Álvarez V, Bullido MJ, Clarimon J, García-Alberca JM, Mir P, Moreno F, Pastor P, Piñol-Ripoll G, Molina-Porcel L, Pérez-Tur J, Rodríguez-Rodríguez E, Royo JL, Sánchez-Valle R, Dichgans M, Rujescu D. Association of Rare APOE Missense Variants V236E and R251G With Risk of Alzheimer Disease. JAMA Neurol. 2022 Jul 1;79(7):652-663. PubMed.
2. Rasmussen KL, Tybjaerg-Hansen A, Nordestgaard BG, Frikke-Schmidt R. APOE and dementia - resequencing and genotyping in 105,597 individuals. Alzheimers Dement. 2020 Dec;16(12):1624-1637. Epub 2020 Aug 18 PubMed.
3. Rasmussen KL, Luo J, Nordestgaard BG, Tybjærg-Hansen A, Frikke-Schmidt R. APOE and vascular disease: Sequencing and genotyping in general population cohorts. Atherosclerosis. 2023 Nov;385:117218. Epub 2023 Aug 9 PubMed.
4. van den Maagdenberg AM, Weng W, de Bruijn IH, de Knijff P, Funke H, Smelt AH, Gevers Leuven JA, van't Hooft FM, Assmann G, Hofker MH. Characterization of five new mutants in the carboxyl-terminal domain of human apolipoprotein E: no cosegregation with severe hyperlipidemia. Am J Hum Genet. 1993 May;52(5):937-46. PubMed.
5. Kang AK, Jenkins DJ, Wolever TM, Huff MW, Maguire GF, Connelly PW, Hegele RA. Apolipoprotein E R112; R251G: a carboxy-terminal variant found in patients with hyperlipidemia and coronary heart disease. Mutat Res. 1997 Sep;382(1-2):57-65. PubMed.
6. Richard P, Beucler I, Pascual De Zulueta M, Biteau N, De Gennes JL, Iron A. Compound heterozygote for both rare apolipoprotein E1 (Gly127-->Asp, Arg158-->Cys) and E3(Cys112-->Arg, Arg251-->Gly) alleles in a multigeneration pedigree with hyperlipoproteinaemia. Clin Sci (Lond). 1997 Jul;93(1):89-95. PubMed.
7. Abou Khalil Y, Marmontel O, Ferrières J, Paillard F, Yelnik C, Carreau V, Charrière S, Bruckert E, Gallo A, Giral P, Philippi A, Bluteau O, Boileau C, Abifadel M, Di-Filippo M, Carrié A, Rabès JP, Varret M. APOE Molecular Spectrum in a French Cohort with Primary Dyslipidemia. Int J Mol Sci. 2022 May 21;23(10) PubMed.
8. Zhao SP, Van den Maagdenberg AM, Vroom TF, Van 't Hooft FM, Gevers Leuvens JA, Havekes LM, Frants RR, Van der Laarse A, Smelt AH. Lipoprotein profiles in a family with two mutants of apolipoprotein E: possible association with hypertriglyceridaemia but not with dysbetalipoproteinaemia. Clin Sci (Lond). 1994 Mar;86(3):323-9. PubMed.
9. Bu G. APOE targeting strategy in Alzheimer's disease: lessons learned from protective variants. Mol Neurodegener. 2022 Aug 3;17(1):51. PubMed.
10. Chen J, Li Q, Wang J. Topology of human apolipoprotein E3 uniquely regulates its diverse biological functions. Proc Natl Acad Sci U S A. 2011 Sep 6;108(36):14813-8. Epub 2011 Aug 22 PubMed.
11. Stuchell-Brereton MD, Zimmerman MI, Miller JJ, Mallimadugula UL, Incicco JJ, Roy D, Smith LG, Cubuk J, Baban B, DeKoster GT, Frieden C, Bowman GR, Soranno A. Apolipoprotein E4 has extensive conformational heterogeneity in lipid-free and lipid-bound forms. Proc Natl Acad Sci U S A. 2023 Feb 14;120(7):e2215371120. Epub 2023 Feb 7 PubMed.
12. Vance JM, Farrer LA, Huang Y, Cruchaga C, Hyman BT, Pericak-Vance MA, Goate AM, Greicius MD, Griswold AJ, Haines JL, Tcw J, Schellenberg GD, Tsai LH, Herz J, Holtzman DM. Report of the APOE4 National Institute on Aging/Alzheimer Disease Sequencing Project Consortium Working Group: Reducing APOE4 in Carriers is a Therapeutic Goal for Alzheimer's Disease. Ann Neurol. 2024 Apr;95(4):625-634. Epub 2024 Jan 5 PubMed.
13. Marduel M, Ouguerram K, Serre V, Bonnefont-Rousselot D, Marques-Pinheiro A, Erik Berge K, Devillers M, Luc G, Lecerf JM, Tosolini L, Erlich D, Peloso GM, Stitziel N, Nitchké P, Jaïs JP, French Research Network on ADH, Abifadel M, Kathiresan S, Leren TP, Rabès JP, Boileau C, Varret M. Description of a large family with autosomal dominant hypercholesterolemia associated with the APOE p.Leu167del mutation. Hum Mutat. 2013 Jan;34(1):83-7. Epub 2012 Oct 11 PubMed.

Other Citations

1. V254E

Further Reading