Resources and insights

A Genome Wide Association Study (GWAS) of Recurrent Exertional Rhabdomyolysis (RER) identified a region on chromosome 16 as the only part of the genome associated with RER [1]. One of the populations in the study showed an association between RER and five Single Nucleotide Polymorphisms (SNPs) that define a 71.6 kb interval within the 822 kb CACNA2D3 gene. The CACNA2D3 gene encodes a regulatory subunit of the voltage-gated calcium channel called the dihydropyridine receptor (DHPR) that functions in excitation-contraction coupling, the process through which signals from motor neurons initiate muscle contraction [2]. The alpha(2)delta subunit encoded by CACNA2D3 sets the abundance of voltage-gated calcium channels by acting as a molecular chaperone. The alpha(2)delta subunit is bound to the plasma membrane through a glycosylphosphatidylinositol (GPI) anchor [3,4]. The level of expression of alpha(2)delta subunits sets the abundance of voltage-gated calcium channels in both neuronal and transfected nonneuronal cells [5,6]. The dihydropyridine receptor, of which the CACNA2D3 protein is a part, is known to physically interact with the ryanodine receptor encoded by the RYR1 gene. Malignant Hyperthermia (MH) in horses is caused by mutations in the RYR1 gene [7]. The RYR1-R2454G allele that causes MH in horses acts as an enhancer of the GYS1-R309H allele of glycogen synthase…

Several recent studies address the mechanism of muscle damage in filaminopathy. Filaminopathy is a term that describes muscle disorders resulting from mutations in the FLNC gene, which encodes filamen C. Two different clinical disease states are included in filaminopathy: Myofibrillar Myopathy 5 (MFM5) and Distal Myopathy 4 (DM4). Studies on the mechanism of muscle damage in filaminopathy suggest that there is an overlap of symptoms between these two disease states, and that the same mechanism of muscle damage takes place in both. Muscle tissue from patients with MFM5 shows cytoplasmic protein aggregates and rimmed vacuoles; electron microscopy shows Z disc fragmentation, Z disc streaming, nemaline rods, and autophagic vacuoles. Muscle tissue from patients with DM4 shows nonspecific changes, not including the Z disc fragmentation and protein aggregates seen in MFM5. Studies of Human Patients A study of muscle tissue from human filaminopathy patients with the W2710X or V930-T933del mutations shows that the large protein aggregates that are seen in Myofibrillar Myopathy contain not only Z disc proteins, but also proteins involved in clearing protein aggregates [1]. Double immunofluorescence with antibodies to either FLNC or MYOT and a series of other proteins shows that the aggregates contain heat shock proteins (HSP20, HSP27, HSP40,…

How can you figure out the chances of different genotypes when breeding horses? Monohybrid Cross It’s easy for one gene. Let’s say that there is a stallion that has one copy of one of the genetic variants associated with PSSM2, the P2 variant (MYOT-S232P). This stallion is heterozygous (n/P2), meaning that he has one normal copy of the MYOT gene and one copy with the P2 mutation. If the stallion is bred to a mare that is clear (n/n) for the P2 allele, the Punnett square below shows the odds. Gametes (sperm and eggs) contain a single copy of each gene in the horse. The stallion produces two kinds of sperm in equal frequency: sperm with P2, and sperm with the normal allele (n). The mare produces eggs with one of the two normal alleles (n). This means that there is a 50% chance of a foal that is n/P2, and a 50% chance of a foal that is n/n. Dihybrid Cross What if we are following two genes? In general, two gene pairs will show independent assortment. This means that the segregation of one pair of alleles into gametes will not influence the segregation of another pair of alleles….

Filamins are a family of actin-binding proteins. In humans and other mammals, there are three filamins: filamin A, filamin B, and filamin C. Filamins A and B are widely expressed, while filamin C expression appears limited to skeletal and cardiac muscle. The figure below (adapted from [1]) shows the domain structure of filamin C. Filamins have an amino-terminal actin-binding domain consisting of two calponin-homology (CH) domains. The rest of the protein consists of 24 immunoglobulin-like filamin domains with two hinge domains. Filamin C differs from filamins A and B in a novel domain 20 that interacts with Z disc proteins, including myotilin and myozenins. While filamin domains 1-15 form a rod, domain 20 is folded over domains 19 and 21 [2]. Filamin domain 24 is required for homodimerization. The figure also shows the positions of human mutations associated with various types of cardiomyopathy and skeletal muscle myopathy, as summarized in the table below. Human FLNC Alleles Associated with Cardiomyopathy Figurea Alleleb Domainc Mutation Typed Citation 1 Y7Tfs*51 ABD frameshift 1 2 Y83* ABD termination 3 25 F106L ABD missense 4 3 E108* ABD termination 5 26 V123A ABD missense 5 4 E238Rfs*14 ABD frameshift 1 5 R269* Filamin 1 termination…

In human patients, mutations in the gene encoding filamin C (FLNC) are associated with Myofibrillar Myopathy 5 [1]. In a prior blog post, we reviewed the human mutations in FLNC associated with the disease state Myofibrillar Myopathy 5 (MFM5) and various cardiomyopathies. We also presented the specific changes associated with the equine P3 allele of FLNC, a haplotype that bears two different missense alleles of FLNC: E753K in Ig-like domain 6 and A1270T in Ig-like domain 11. In this blog post, we review bioinformatic evidence associating the P3 allele of FLNC with Myofibrillar Myopathy in horses. The association of the P3 allele of FLNC with Myofibrillar Myopathy in horses was first made in a horse diagnosed with Myofibrillar Myopathy (MFM) through the identification of desmin-positive inclusions observed in muscle tissue. The horse displayed symptoms of exercise intolerance. A genetic test for GYS1-R309H (P1) ruled out Polysaccharide Storage Myopathy type 1 (PSSM1). The desmin staining technique was relatively new at the time of the muscle biopsy on this horse; the presence of desmin-positive inclusions was diagnostic of MFM in distinction to other types of Polysaccharide Storage Myopathy type 2 (PSSM2). The association of the P3 allele of FLNC with Myofibrillar Myopathy…

Is PSSM2 genetic? By genetic, we mean that it is an inherited condition, transmitted from parents to offspring rather than resulting exclusively from environmental conditions, such as diet or exposure to toxins or pathogens. What do we mean by PSSM2? PSSM2 is an abbreviation for Polysaccharide Storage Myopathy type 2 [1]. It is a disease state in horses initially defined by the following criteria: symptoms of exercise intolerance, a negative DNA test for GYS1-R309H (a genetic variant of the gene encoding glycogen synthase that is associated with the disease state PSSM1), and abnormalities observed in muscle biopsy [1]. The first evidence suggesting that PSSM2 is genetic was presented in 2007, when Dr. Molly McCue presented a pedigree of affected Quarter Horses related by descent from a common ancestor in her PhD dissertation. Related horses were diagnosed with PSSM2 by muscle biopsy. Polysaccharide Storage Myopathy type 1 (PSSM1) was shown to be associated with GYS1-R309H in 2008 [2]. The GYS1-R309H variant has been shown to encode a constitutively activated glycogen synthase [3]. The improper regulation of this enzyme pushes the equilibrium between glycogen synthesis and degradation in the direction of glycogen synthesis, even when the affected horse’s energy needs would normally…

Myotilin is a muscle protein encoded by the MYOT gene. Myotilin is a structural protein of muscle that localizes to the Z discs. Myotilin contains two Immunoglobulin-like (Ig-like) domains similar to those of palladin, myopalladin, titin, and other proteins. The amino-terminal portion of myotilin contains a large number of serine residues and is not similar to any other known protein. Human patients with Limb-Girdle Muscular Dystrophy (LGMD) or Myofibrillar Myopathy (MFM) have been found to have mutations in MYOT. Both LGMD and MFM are disease states associated with mutations in a number of different genes. A substantial fraction of human patients with LGMD or MFM do not receive a molecular diagnosis, suggesting that there are undiscovered genetic variants responsible for some of these cases. Human patients with LGMD that have mutations in MYOT are diagnosed with Limb-Girdle Muscular Dystrophy 1A (LGMD1A), while human patients with MFM that have mutations in MYOT are diagnosed with Myofibrillar Myopathy 3 (MFM3). Pathogenic mutations in MYOT are dominant missense alleles. Known pathogenic alleles include S55F, T57I, S60C, S60F, and S95I [1, 2, 3]. There is a substantial difference in the age of onset and the symptoms of human patients with MYOT mutations, even among…