Date Published: January 19, 2017
Publisher: Public Library of Science
Author(s): Carlos A. Pantoja-Melendez, Antonio Miranda-Duarte, Bladimir Roque-Ramirez, Juan C. Zenteno, Michael Kyba.
Limb-Girdle Muscular Dystrophy type 2 (LGMD2) is a group of autosomally recessive inherited disorders defined by weakness and wasting of the shoulder and pelvic girdle muscles. In the past, several population isolates with high incidence of LGMD2 arising from founder mutation effects have been identified. The aim of this work is to describe the results of clinical, epidemiologic, and molecular studies performed in a Mexican village segregating numerous cases of LGMD2. A population census was conducted in the village to identify all LGMD affected patients. Molecular analysis included genome wide homozygosity mapping using a 250K SNP Affymetrix microarray followed by PCR amplification and direct nucleotide sequencing of the candidate gene. In addition, DNA from 401 randomly selected unaffected villagers was analyzed to establish the carrier frequency of the LGMD2 causal mutation. A total of 32 LGMD2 patients were identified in the village, rendering a disease prevalence of 4.3 (CI: 2.9–5.9) cases per 1,000 habitants (1 in 232). Genome wide homozygosity mapping revealed that affected individuals shared a 6.6 Mb region of homozygosity at chromosome 15q15. The identified homozygous interval contained CAPN3, the gene responsible for LGMD2 type A (LGMD2A). Direct sequencing of this gene revealed homozygosity for a novel c.348C>A mutation (p.Ala116Asp) in DNA from all 20 affected subjects available for genetic screening, except one which was heterozygous for the mutation. In such patient, a heterozygous c.2362AG>TCATCT deletion/insertion was recognized as the second CAPN3 mutation. Western blot and autocatalytic activity analyses in protein lysates from skeletal muscle biopsy obtained from a p.Ala116Asp homozygous patient suggested that this particular mutation increased the autocatalytic activity of CAPN3. Thirty eigth heterozygotes of the p.Ala116Asp mutation were identified among 401 genotyped unaffected villagers, yielding a population carrier frequency of 1 in 11. This study demonstrates that a cluster of patients with LGMD2A in a small Mexican village arises from a novel CAPN3 founder mutation. Evidence of allelic heterogeneity is demonstrated by the recognition of an additional CAPN3 mutation in a single affected. Our study provides an additional example of genetic isolation causing a high prevalence of LGMD and of successful molecular characterization of the disease by means of homozygosity mapping. The identification of a very high carrier frequency of the LGMD2-causing mutation has implications for more rational genetic counseling in this community.
The muscular dystrophies encompass a clinically heterogeneous group of inherited disorders characterized by progressive weakness and degeneration of the skeletal muscles. The distribution and severity of muscular affectation in these conditions as well as their age of onset is highly variable. In some specific forms of the disease, non-skeletal muscles such as cardiac and respiratory muscles could also be affected and other organs such as brain and eyes can be involved. All forms of muscular dystrophy aggravates as muscles progressively degenerate and weaken, and most affected individuals eventually lose their ability to walk . Muscular dystrophies are caused by mutations in any of the dozens of genes encoding proteins needed for muscle integrity and function and are among the most genetically heterogeneous human conditions with up to 165 muscular dystrophies and myopathies-causative genes listed in a recent review .
Mexican mestizos are a recently admixed population composed of Amerindian, European, and, to a lesser extent, African ancestry. Although this ethnic admixture generated considerable genetic heterogeneity between subpopulations from different regions throughout the country , the practice of consanguineous marriages, which is still common in many communities, reduces genetic variation and increases the chances of recessive disease. In this work, the clinical, epidemiological, and molecular characterization of an isolated Mexican population with a high incidence of LGMD type 2A was performed. LGMD2 is a clinically and genetically heterogeneous group of myopathies with substantial prevalence variation in different geographical regions. Previously, the ethnic origin of patients was suggested as a feature to be taken into account for helping in the clinical distinction between LGMD subtypes (Reviewed in ). To date, very few investigations on the clinical and molecular features of Mexican LGMD patients have been performed. In a recent report from Mexico, Gomez-Diaz et al. used immunofluorescence staining to analyze muscle biopsies from 212 patients with various muscular dystrophies and identified that 18% of them corresponded to dysferlinopathies while 11% were calpainopathies .