Brugada syndrome predisposes to sudden death due to disruption of normal cardiac ion channel function, yet our understanding of the underlying cellular mechanisms is incomplete. Commonly used heterologous expression models lack many characteristics of native cardiomyocytes and, in particular, the individual genetic background of a patient. Patient-specific induced pluripotent stem (iPS) cell-derived cardiomyocytes (iPS-CM) may uncover cellular phenotypical characteristics not observed in heterologous models. Our objective was to determine the properties of the sodium current in iPS-CM with a mutation in SCN5A associated with Brugada syndrome.Dermal fibroblasts from a Brugada syndrome patient with a mutation in SCN5A (c.1100G > A, leading to Nav1.5_p.R367H) were reprogrammed to iPS cells. Clones were characterized and differentiated to form beating clusters and sheets. Patient and control iPS-CM were structurally indistinguishable. Sodium current properties of patient and control iPS-CM were compared. These results were contrasted with those obtained in tsA201 cells heterologously expressing sodium channels with the same mutation.Patient-derived iPS-CM showed a 33.1–45.5% reduction in INa density, a shift in both activation and inactivation voltage-dependence curves, and faster recovery from inactivation. Co-expression of wild-type and mutant channels in tsA201 cells did not compromise channel trafficking to the membrane, but resulted in a reduction of 49.8% in sodium current density without affecting any other parameters.Cardiomyocytes derived from iPS cells from a Brugada syndrome patient with a mutation in SCN5A recapitulate the loss of function of sodium channel current associated with this syndrome; including pro-arrhythmic changes in channel function not detected using conventional heterologous expression systems.

All mitochondrial tRNAs in Trypanosoma brucei derive from cytosolic tRNAs that are in part imported into mitochondria. Some trypanosomal tRNAs are thiolated in a compartment-specific manner. We have identified three proteins required for the thio modification of cytosolic tRNA Gln , tRNA Glu , and tRNA Lys . RNA interference-mediated ablation of these proteins results in the cytosolic accumulation non-thio-modified tRNAs but does not increase their import. Moreover, in vitro import experiments showed that both thio-modified and non-thio-modified tRNA Glu can efficiently be imported into mitochondria. These results indicate that unlike previously suggested the cytosolspecific thio modifications do not function as antideterminants for mitochondrial tRNA import. Consistent with these results we showed by using inducible expression of a tagged tRNA Glu that it is mainly the thiolated form that is imported in vivo. Unexpectedly, the imported tRNA becomes dethiolated after import, which explains why the non-thiolated form is enriched in mitochondria. Finally, we have identified two genes required for thiolation of imported tRNA Trp whose wobble nucleotide is subject to mitochondrial C to U editing. Interestingly, downregulation of thiolation resulted in an increase of edited tRNA Trp but did not affect growth.Most protozoa, many fungi, plants, and a few animals lack a variable number of mitochondrial tRNA genes. It has been shown in these organisms that the missing genes are compensated for by import of a small fraction of the corresponding cytosolic tRNAs (1, 2). Among all organisms that import tRNAs trypanosomatids such as Trypanosoma brucei and Leishmania are extreme in that their mitochondrial genomes have lost all tRNA genes. Trypanosomatids therefore need to import the entire set of mitochondrial tRNAs. As a consequence all mitochondrial tRNAs in trypanosomatids derive from eukaryotic-type cytosolic tRNAs that need to function in the context of the bacterial-type translation system of mitochondria (3). Two tRNAs, the initiator tRNA Met and the tRNA Sec , are cytosol-specific. It has been shown that in T. brucei the tRNA import specificity is mediated by binding to cytosolic translation elongation factor 1a (4). The fact that initiator tRNA Met and tRNA Sec do not bind to elongation factor 1a therefore explains their exclusive cytosolic localization.The extent of mitochondrial localization of different trypanosomal tRNAs varies by at least an order of magnitude (5, 6). The same is true for other organisms that import tRNAs (7). Cells therefore need to determine both the tRNA import specificity as well as the extent of mitochondrial localization of each imported tRNA species. How this is achieved is not known in any species.In trypanosomatids mitochondrial tRNAs and their cytosolic counterparts derive from the same nuclear genes. However, due to compartment-specific post-transcriptional nucleotide modifications, the cytosolic and the corresponding imported tRNAs are often physically different. Mitochondria-specifi...

Brugada syndrome is an inherited, rare cardiac arrhythmogenic disease, associated with sudden cardiac death. It accounts for up to 20% of sudden deaths in patients without structural cardiac abnormalities. The majority of mutations involve the cardiac sodium channel gene SCN5A and give rise to classical abnormal electrocardiogram with ST segment elevation in the right precordial leads V1 to V3 and a predisposition to ventricular fibrillation. The pathophysiological mechanisms of Brugada syndrome have been investigated using model systems including transgenic mice, canine heart preparations, and expression systems to study different SCN5A mutations. These models have a number of limitations. The recent development of pluripotent stem cell technology creates an opportunity to study cardiomyocytes derived from patients and healthy individuals. To date, only a few studies have been done using Brugada syndrome patient-specific iPS-CM, which have provided novel insights into the mechanisms and pathophysiology of Brugada syndrome. This review provides an evaluation of the strengths and limitations of each of these model systems and summarizes the key mechanisms that have been identified to date.