ANXA11 mutations prevail in Chinese ALS patients with and without cognitive dementia

Abstract

Objective: To investigate the genetic contribution of ANXA11, a gene associated with amyotrophic lateral sclerosis (ALS), in Chinese ALS patients with and without cognitive dementia.

Methods: Sequencing all the coding exons of ANXA11 and intron-exon boundaries in 18 familial amyotrophic lateral sclerosis (FALS), 353 unrelated sporadic amyotrophic lateral sclerosis (SALS), and 12 Chinese patients with ALS-frontotemporal lobar dementia (ALS-FTD). The transcripts in peripheral blood generated from a splicing mutation were examined by reverse transcriptase PCR.

Results: We identified 6 nonsynonymous heterozygous mutations (5 novel and 1 recurrent), 1 splice site mutation, and 1 deletion of 10 amino acids (not accounted in the mutant frequency) in 11 unrelated patients, accounting for a mutant frequency of 5.6% (1/18) in FALS, 2.3% (8/353) in SALS, and 8.3% (1/12) in ALS-FTD. The deletion of 10 amino acids was detected in 1 clinically undetermined male with an ALS family history who had atrophy in hand muscles and myotonic discharges revealed by EMG. The novel p. P36R mutation was identified in 1 FALS index, 1 patient with SALS, and 1 ALS-FTD. The splicing mutation (c.174-2A>G) caused in-frame skipping of the entire exon 6. The rest missense mutations including p.D40G, p.V128M, p.S229R, p.R302C and p.G491R were found in 6 unrelated patients with SALS.

Conclusions: The ANXA11 gene is one of the most frequently mutated genes in Chinese patients with SALS. A canonical splice site mutation leading to skipping of the entire exon 6 further supports the loss-of-function mechanism. In addition, the study findings further expand the ANXA11 phenotype, first highlighting its pathogenic role in ALS-FTD.

Figure 1. Mutations identified in ANXA11 in ALS and ALS-FTD patients

Mutations found in the previous study and in the present study are marked in black and in red, respectively. A deletion mutation associated to a clinically undetermined patient is marked in blue. Conservation of amino acid across species is shown at the bottom. ALS = amyotrophic lateral sclerosis; FTD = frontotemporal lobe dementia.

Figure 2. Updated mutation spectrum of FALS and SALS in Chinese populations

SOD1 is the most common mutant gene in FALS, and ANXA11 in SALS. FALS = familial amyotrophic lateral sclerosis; SALS = sporadic amyotrophic lateral sclerosis.

Figure 3. Transcripts of the splicing mutation (c.174-2A>G) detected by reverse transcription PCR analysis

A 1.5% agarose gel fractionation of RT-PCR products of blood RNA shows the distinct fragments of cDNA obtained by specific primers. Lane WT represents a normal transcript in a healthy control. Lane MU represents an aberrant transcript in a patient carrying the c.174-2A>G mutation, which contains 3 different bans. Sequencing results show that the band located at 750 bp implies a normal transcript; the band located at 250 bp implies an aberrant transcript, which was caused by exon 6 skipping; and the band located at 500 bp is confirmed to be a result of formation of heterodimer consisting of 1 normal transcript and 1 aberrant transcript. ALS = amyotrophic lateral sclerosis; cDNA = complementary DNA; FTD = frontotemporal lobe dementia; RT-PCR = reverse transcription PCR.