A Gene for Autosomal Recessive Limb-Girdle Muscular Dystrophy in Manitoba Hutterites Maps to Chromosome Region 9q31-q33: Evidence for Another Limb-Girdle Muscular Dystrophy Locus

SummaryCharacterized by proximal muscle weakness and wasting, limb-girdle muscular dystrophies (LGMDs) are a heterogeneous group of clinical disorders. Previous reports have documented either autosomal dominant or autosomal recessive modes of inheritance, with genetic linkage studies providing evidence for the existence of at least 12 distinct loci. Gene products have been identified for five genes responsible for autosomal recessive forms of the disorder. We performed a genome scan using pooled DNA from a large Hutterite kindred in which the affected members display a mild form of autosomal recessive LGMD. A total of 200 markers were used to screen pools of DNA from patients and their siblings. Linkage between the LGMD locus and D9S302 (maximum LOD score 5.99 at recombination fraction .03) was established. Since this marker resides within the chromosomal region known to harbor the gene causing Fukuyama congenital muscular dystrophy (FCMD), we expanded our investigations, to include additional markers in chromosome region 9q31-q34.1. Haplotype analysis revealed five recombinations that place the LGMD locus distal to the FCMD locus. The LGMD locus maps close to D9S934 (maximum multipoint LOD score 7.61) in a region that is estimated to be ∼4.4 Mb (Genetic Location Database composite map). On the basis of an inferred ancestral recombination, the gene may lie in a 300-kb region between D9S302 and D9S934. Our results provide compelling evidence that yet another gene is involved in LGMD; we suggest that it be named “LGMD2H.

TRIM32 Regulates Skeletal Muscle Stem Cell Differentiation and Is Necessary for Normal Adult Muscle Regeneration

Limb girdle muscular dystrophy type 2H (LGMD2H) is an inherited autosomal recessive disease of skeletal muscle caused by a mutation in the TRIM32 gene. Currently its pathogenesis is entirely unclear. Typically the regeneration process of adult skeletal muscle during growth or following injury is controlled by a tissue specific stem cell population termed satellite cells. Given that TRIM32 regulates the fate of mammalian neural progenitor cells through controlling their differentiation, we asked whether TRIM32 could also be essential for the regulation of myogenic stem cells. Here we demonstrate for the first time that TRIM32 is expressed in the skeletal muscle stem cell lineage of adult mice, and that in the absence of TRIM32, myogenic differentiation is disrupted. Moreover, we show that the ubiquitin ligase TRIM32 controls this process through the regulation of c-Myc, a similar mechanism to that previously observed in neural progenitors. Importantly we show that loss of TRIM32 function induces a LGMD2H-like phenotype and strongly affects muscle regeneration in vivo. Our studies implicate that the loss of TRIM32 results in dysfunctional muscle stem cells which could contribute to the development of LGMD2H

Erratum to: Mutation screening in 86 known X-linked mental retardation genes by droplet-based multiplex PCR and massive parallel sequencing

Item does not contain fulltext[This corrects the article DOI: 10.1007/s11568-010-9137-y.].1 december 200

Limb-Girdle Muscular Dystrophy Type 2H Associated with Mutation in TRIM32, a Putative E3-Ubiquitin–Ligase Gene

Limb-girdle muscular dystrophy type 2H (LGMD2H) is a mild autosomal recessive myopathy that was first described in the Manitoba Hutterite population. Previous studies in our laboratory mapped the causative gene for this disease to a 6.5-Mb region in chromosomal region 9q31-33, flanked by D9S302 and D9S1850. We have now used additional families and a panel of 26 microsatellite markers to construct haplotypes. Twelve recombination events that reduced the size of the candidate region to 560 kb were identified or inferred. This region is flanked by D9S1126 and D9S737 and contains at least four genes. Exons of these genes were sequenced in one affected individual, and four sequence variations were identified. The families included in our study and 100 control individuals were tested for these variations. On the basis of our results, the mutation in the tripartite-motif–containing gene (TRIM32) that replaces aspartate with asparagine at position 487 appears to be the causative mutation of LGMD2H. All affected individuals were found to be homozygous for D487N, and this mutation was not found in any of the controls. This mutation occurs in an NHL (named after the proteins NCL1, HT2A, and LIN-41) domain at a position that is highly conserved. NHL domains are known to be involved in protein-protein interactions. Although the function of TRIM32 is unknown, current knowledge of the domain structure of this protein suggests that it may be an E3-ubiquitin ligase. If proven, this represents a new pathogenic mechanism leading to muscular dystrophy

Intragenic deletion of TRIM32 in compound heterozygotes with sarcotubular myopathy/LGMD2H

In 2005 the commonality of sarcotubular myopathy (STM) and limb girdle muscular dystrophy type 2H (LGMD2H) was demonstrated, as both are caused by the p D487N missense mutation in TRIM32 originally found in the Manitoba Hutterite population. Recently, three novel homozygous TRIM32 mutations have been described in LGMD patients. Here we describe a three generation Swedish family clinically presenting with limb girdle muscular weakness and histological features of a microvacuolar myopathy. The two index patients were compound heterozygotes for a frameshift mutation in TRIM32 (c.1560delC ) and a 30 kb intragenic deletion, encompassing parts of intron 1 and the entire exon 2 of TRIM32. In these patients, no full-length or truncated TRIM32 could be detected. Interestingly, heterozygous family members carrying only one mutation showed mild clinical symptoms and vacuolar changes in muscle. In our family, the phenotype encompasses additionally a mild demyelinating polyneuropathic syndrome. Thus STM and LGMD2H are the result of loss of function mutations that can be either deletions or missense mutations

Degradation of c-Myc is critical for TRIM32 induced differentiation.

<p>(a) C2C12 cells were transfected with the indicated constructs (left grey boxes) and cultivated for three days in 2.5% HS. Immunostainings of the cells labelled with the indicated markers (upper grey boxes) are shown. (b) Diagram showing the fraction of transfected cells that undergo muscular differentiation (Myosin positive). (mean ± std; *P<0.001 compared to EGFP).</p

TRIM32 is induced during muscle regeneration <i>in vivo</i>.

<p>(a) and (c) Hematoxylin and Eosin (H&E) staining of regenerating TA muscle from wild type mice 3 days post cardiotoxin (CTX) injection. Arrows in (a) indicate the regions of CTX-induced muscle regeneration. (c) shows enlarged region of the boxed area in (a). (b) and (d) RNA in situ hybridization showing <i>TRIM32</i> expression (blue) in wild type mouse TA muscle 3 days post CTX injection. (d) shows enlarged regions of the boxed area in (b). (e) H&E staining of regenerating TA muscle from wild type mice 7 days post CTX injection. (f) RNA in situ hybridization showing <i>TRIM32</i> expression (blue) in wild type mouse TA muscle 7 days post CTX injection.</p

<i>TRIM32−/−</i> mice show delayed and impaired skeletal muscle regeneration.

<p>(a)–(d) H&E staining of transverse cross sections through CTX-injured TA muscles 5 days post injection from wild type (a) and (c) and <i>TRIM32−/−</i> (b) and (d) mice. (c) and (d) show enlarged regions of the boxed areas in (a) and (b), respectively. (e)–(h). H&E staining of transverse cross sections through CTX-injured TA muscles 16 days post injection from wild type (e) and (g) and <i>TRIM32−/−</i> (f) and (h) mice. (g) and (h) show enlarged regions of (e) and (f), respectively. Arrow in (g) indicates centrally nucleated myofiber characteristic of efficient muscle regeneration in wild type TA. In contrast, some regions of injured <i>TRIM32−/−</i> TA display small centrally nucleated myofibers (h, arrowhead), adipocyte-like cells (f, arrow) or fibrosis (h, arrow).</p