Disruption of Striated Preferentially Expressed Gene Locus Leads to Dilated Cardiomyopathy in Mice

Abstract

Background-The striated preferentially expressed gene (Speg) generates 4 different isoforms through alternative promoter use and tissue-specific splicing. Depending on the cell type, Speg isoforms may serve as markers of striated or smooth muscle differentiation. Methods and Results-To elucidate function of Speg gene isoforms, we disrupted the Speg gene locus in mice by replacing common exons 8, 9, and 10 with a lacZ gene. ␤-Galactosidase activity was detected in cardiomyocytes of the developing heart starting at day 11.5 days post coitum (dpc). ␤-Galactosidase activity in other cell types, including vascular smooth muscle cells, did not begin until 18.5 dpc. In the developing heart, protein expression of only Speg␣ and Speg␤ isoforms was present in cardiomyocytes. Homozygous Speg mutant hearts began to enlarge by 16.5 dpc, and by 18.5 dpc, they demonstrated dilation of right and left atria and ventricles. These cardiac abnormalities in the absence of Speg were associated with a cellular hypertrophic response, myofibril degeneration, and a marked decrease in cardiac function. Moreover, Speg mutant mice exhibited significant neonatal mortality, with increased death occurring by 2 days after birth. Conclusions-These findings demonstrate that mutation of the Speg locus leads to cardiac dysfunction and a phenotype consistent with a dilated cardiomyopathy. Key Words: cardiomyopathy Ⅲ hypertrophy Ⅲ myocytes Ⅲ genes M yosin light chain kinases (MLCK) are a family of proteins that are important for myocyte function, including structure and regulation of the actin-based cytoskeleton. 1,2 One member of this family, striated preferentially expressed gene (Speg), has isoforms in both striated and smooth muscle cells. 3 The Speg locus contains 2 transcriptional start sites, and through alternative promoter use and splicing in a tissue-specific manner, it generates 4 different isoforms. 3 Speg␣ and Speg␤ are expressed in striated muscle (cardiac and skeletal), aortic preferentially expressed gene (Apeg-1) is expressed in smooth muscle (predominantly vascular), and brain preferentially expressed gene (Bpeg) is expressed in the brain and aorta. [3][4][5] Speg␣ and Speg␤ share homology with MLCK family members. Specifically, the Speg isoforms, along with obscurin-MLCK, are unique members of the MLCK family, because they contain 2 tandemly arranged serine/threonine kinase (MLCK) domains. 2 Beyond the MLCK domains, Speg␣ and Speg␤ also contain immunoglobulin and fibronectin domains that are characteristic of the MLCK family of proteins. Prior investigations in our laboratory revealed that Speg␣ and Speg␤ are sensitive markers of striated muscle differentiation and that Speg colocalizes with desmin in the sarcomeric Z disc 3 ; however, the functional significance of Speg isoforms is yet to be elucidated. Editorial p 213 Clinical Perspective p 268In contrast to smooth muscle cells, striated muscle cells of both cardiac and skeletal origin undergo terminal differenti- Received June 15, 2008; accepted October 14, 2008. From Pulmona...

SPEG (Striated Muscle Preferentially Expressed Protein Kinase) Is Essential for Cardiac Function by Regulating Junctional Membrane Complex Activity

“…Quantitative PCR revealed an 83% reduction in SPEG transcripts compared to non-failing (NF) human hearts (Figure 2A). To circumvent prenatal lethality in germline Speg knockout mice 10 and study the role of SPEG in cardiac myocytes, we obtained generated conditional cardiac myocyte-restricted tamoxifen-inducible Speg knockout (MCM- Speg fl/fl ) mice by crossing Speg -floxed (fl) mice with Myh6 -driven mER-Cre-mER (MCM) mice. Upon administration of tamoxifen for 5 days, SPEG protein levels were assessed using Western blotting in the hearts of MCM- Speg fl/fl and MCM control mice (Figure 2B).…”

“…These findings are consistent with the prior observation that rare Speg mutations are associated with a genetic disease called centronuclear myopathy, which is often associated with cardiomyopathy. 11 Since complete SPEG deficiency is embryonically lethal, 10 we developed a novel inducible, tissue-specific Speg knockout model. By crossing the tamoxifen-inducible cardiac myocyte-specific Cre transgenic model (αMHC-MerCreMer) with a floxed SPEG fl/fl mouse, we were able to quickly downregulate SPEG levels by 80% in the mouse heart.…”

“…Disruption of the SPEG gene locus in developing mouse hearts resulted in cardiac myocyte hypertrophy, myofibril degeneration, and impaired cardiac function, leading to increased neonatal mortality. 10 It had yet to be determined whether loss of SPEG in mature myocardium affected cardiac contractility and cardiac myocyte organization. Our study is the first to show that deletion of SPEG in the adult heart has a profound impact on cardiac function and calcium handling.…”

“…It has been previously demonstrated that Speg is important for cardiac development, as a germline knockout of all Speg isoforms resulted in embryonic dilated cardiomyopathy and early postnatal death. 10 Additionally, rare Speg mutations have been implicated in centronuclear myopathy, a genetic disease of skeletal and cardiac muscle. 11 Importantly, however, nothing is known to date about the potential role of Speg in the adult heart, or the mechanisms by which altered Speg levels modulate cardiac dysfunction.…”

Modifier locus of the skeletal muscle involvement in Emery–Dreifuss muscular dystrophy

“…So far, we excluded the presence of modifier variants in the analyzed regions. However, some regions remain to be analyzed, in particular the MYL1 gene promoter, the SPEG1 gene coding and regulatory regions specifically expressed in skeletal and cardiac muscles (Liu et al 2009). Thus, additional experiments are needed to identify the modifier gene in the EMD1 family and to understand the mechanism by which this gene influences muscular related phenotype variability in the family.…”