Expanding SPG18 clinical spectrum: autosomal dominant mutation causes complicated hereditary spastic paraplegia in a large family

Abstract

Background: SPG18 is caused by mutations in the endoplasmic reticulum lipid raft associated 2 (ERLIN2) gene. Autosomal recessive (AR) mutations are usually associated with complicated hereditary spastic paraplegia (HSP), while autosomal dominant (AD) mutations use to cause pure SPG18.

Aim: To define the variegate clinical spectrum of the SPG18 and to evaluate a dominant negative effect of erlin2 (encoded by ERLIN2) on oligomerization as causing differences between AR and AD phenotypes.

Methods: In a four-generation pedigree with an AD pattern, a spastic paraplegia multigene panel test was performed. Oligomerization of erlin2 was analyzed with velocity gradient assay in fibroblasts of the proband and healthy subjects.

Results: Despite the common p.V168M mutation identified in ERLIN2, a phenoconversion to amyotrophic lateral sclerosis (ALS) was observed in the second generation, pure HSP in the third generation, and a complicated form with psychomotor delay and epilepsy in the fourth generation. Erlin2 oligomerization was found to be normal.

Discussion: We report the first AD SPG18 family with a complicated phenotype, and we ruled out a dominant negative effect of V168M on erlin2 oligomerization. Therefore, our data do not support the hypothesis of a relationship between the mode of inheritance and the phenotype, but confirm the multifaceted nature of SPG18 on both genetic and clinical point of view. Clinicians should be aware of the importance of conducting an in-depth clinical evaluation to unmask all the possible manifestations associated to an only apparently pure SPG18 phenotype. We confirm the genotype-phenotype correlation between V168M and ALS emphasizing the value of close follow-up.

Keywords: Amyotrophic lateral sclerosis (ALS); ERLIN2; Hereditary spastic paraplegia (HSP); SPG18.

Fig. 1

Family pedigree. Black filled symbol affected; white symbol, unaffected; diamond shaped symbol, masked gender. Roman numerals represent the generation. Arabic numerals identify individuals. The arrow indicates the proband. Sanger sequencing was performed in subjects with asterisks: all carried the V168M mutation at heterozygous state except for IV-9

Fig. 2

Analysis of erlin-2 structural model. Ribbon representation of the predicted structure of erlin2 (Val23-Thr238). Residues in the sequence are colored as the domains they belong to. Met168 is colored in blue, while the other residues involved in the mutations discussed in our analysis are colored in red. All mutated residues are mapped into the structure and represented as sticks and dots

Fig. 3

Analysis of oligomeric state of erlin2. The oligomeric state of erlin2 was evaluated by sedimentation on velocity gradient [8] (A) or by Blue native electrophoresis page [9] (BN-PAGE) (B). A Healthy subject and patient (III-12) fibroblasts were lysed in buffer containing 0.4% SDS and 0.2% Triton X-100 and run through 5–30% sucrose gradients. Fractions of 500 µl were collected from the top (fraction 1) to the bottom (fraction 9) of the gradients. Proteins were TCA-precipitated and detected by western blotting using a specific erlin2 antibody. The distribution on the gradients of the molecular mass markers is indicated. Densitometric analysis of protein distribution on the gradients, expressed as percentage in each fraction, is shown at the bottom; values are the mean of two different experiments ± S.D. B Healthy subject and patient (III-12) fibroblasts were homogenized in pounce homogenizer (30 strokes) in sucrose buffer (83 mM sucrose, 6.6 mM imidazole/HCL, pH 7.0) and centrifuged for 10 min at 20,000 g. Membrane pellets were extracted in the solubilization buffer (50 mM NaCl, 2 mM aminocaproic acid, 1 mM EDTA, 1% TX-100, 5% glycerol, pH 7.0). Protein samples (40 µg/lane) were resolved on a 6–15% gradient gel, then electroblotted to PVDF membrane and detected using a specific erlin2 antibody. Molecular weight standards are indicated