RESUMO
BACKGROUND: Cardiac transverse tubules (T-tubules) are anchored to sarcomeric Z-discs by costameres to establish a regular spaced pattern. One of the major components of costameres is the dystrophin-glycoprotein complex (DGC). Nevertheless, how the assembly of the DGC coordinates with the formation and maintenance of T-tubules under physiological and pathological conditions remains unclear. METHODS: Given the known role of Ptpn23 (protein tyrosine phosphatase, nonreceptor type 23) in regulating membrane deformation, its expression in patients with dilated cardiomyopathy was determined. Taking advantage of Cre/Loxp, CRISPR/Cas9, and adeno-associated virus 9 (AAV9)-mediated in vivo gene editing, we generated cardiomyocyte-specific Ptpn23 and Actn2 (α-actinin-2, a major component of Z-discs) knockout mice. We also perturbed the DGC by using dystrophin global knockout mice (DmdE4*). MM 4-64 and Di-8-ANEPPS staining, Cav3 immunofluorescence, and transmission electron microscopy were performed to determine T-tubule structure in isolated cells and intact hearts. In addition, the assembly of the DGC with Ptpn23 and dystrophin loss of function was determined by glycerol-gradient fractionation and SDS-PAGE analysis. RESULTS: The expression level of Ptpn23 was reduced in failing hearts from dilated cardiomyopathy patients and mice. Genetic deletion of Ptpn23 resulted in disorganized T-tubules with enlarged diameters and progressive dilated cardiomyopathy without affecting sarcomere organization. AAV9-mediated mosaic somatic mutagenesis further indicated a cell-autonomous role of Ptpn23 in regulating T-tubule formation. Genetic and biochemical analyses showed that Ptpn23 was essential for the integrity of costameres, which anchor the T-tubule membrane to Z-discs, through interactions with α-actinin and dystrophin. Deletion of α-actinin altered the subcellular localization of Ptpn23 and DGCs. In addition, genetic inactivation of dystrophin caused similar T-tubule defects to Ptpn23 loss-of-function without affecting Ptpn23 localization at Z-discs. Last, inducible Ptpn23 knockout at 1 month of age showed Ptpn23 is also required for the maintenance of T-tubules in adult cardiomyocytes. CONCLUSIONS: Ptpn23 is essential for cardiac T-tubule formation and maintenance along Z-discs. During postnatal heart development, Ptpn23 interacts with sarcomeric α-actinin and coordinates the assembly of the DGC at costameres to sculpt T-tubule spatial patterning and morphology.
RESUMO
Keloids are characterized by fibroblast hyperproliferation and excessive accumulation of extracellular matrix (ECM) and are a major global health care burden among cutaneous diseases. However, the function of long noncoding RNA (lncRNA)-mediated ECM remodeling during the pathogenesis of keloids is still unclear. Herein, we identified a long noncoding transcript, namely, lymphocyte-specific protein 1 pseudogene 5 (LSP1P5), that modulates ECM component deposition in keloids. First, high-throughput transcriptome analysis showed that LSP1P5 was selectively upregulated in keloids and correlated with more severe disease in a clinical keloid cohort. Therapeutically, the attenuation of LSP1P5 significantly decreased the expression of ECM markers (COL1, COL3, and FN1) both in vitro and in vivo. Intriguingly, an antifibrotic gene, CCAAT enhancer binding protein alpha (CEBPA), is a functional downstream candidate of LSP1P5. Mechanistically, LSP1P5 represses CEBPA expression by hijacking Suppressor of Zeste 12 to the promoter of CEBPA, thereby enhancing the polycomb repressive complex 2-mediated H3K27me3 and changing the chromosomal opening status of CEBPA. Taken together, these findings indicate that targeting LSP1P5 abrogates fibrosis in keloids through epigenetic regulation of CEBPA, revealing a novel antifibrotic therapeutic strategy that bridges our current understanding of lncRNA regulation, histone modification and ECM remodeling in keloids.