Tnni3k alleles influence ventricular mononuclear diploid cardiomyocyte frequency

Recent evidence implicates mononuclear diploid cardiomyocytes as a proliferative and regenerative subpopulation of the postnatal heart. The number of these cardiomyocytes is a complex trait showing substantial natural variation among inbred mouse strains based on the combined influences of multiple polymorphic genes. One gene confirmed to influence this parameter is the cardiomyocyte-specific kinase Tnni3k. Here, we have studied Tnni3k alleles across a number of species. Using a newly-generated kinase-dead allele in mice, we show that Tnni3k function is dependent on its kinase activity. In an in vitro kinase assay, we show that several common human TNNI3K kinase domain variants substantially compromise kinase activity, suggesting that TNNI3K may influence human heart regenerative capacity and potentially also other aspects of human heart disease. We show that two kinase domain frameshift mutations in mice cause loss-of-function consequences by nonsense-mediated decay. We further show that the Tnni3k gene in two species of mole-rat has independently devolved into a pseudogene, presumably associated with the transition of these species to a low metabolism and hypoxic subterranean life. This may be explained by the observation that Tnni3k function in mice converges with oxidative stress to regulate mononuclear diploid cardiomyocyte frequency. Unlike other studied rodents, naked mole-rats have a surprisingly high (30%) mononuclear cardiomyocyte level but most of their mononuclear cardiomyocytes are polyploid; their mononuclear diploid cardiomyocyte level (7%) is within the known range (2–10%) of inbred mouse strains. Naked mole-rats provide further insight on a recent proposal that cardiomyocyte polyploidy is associated with evolutionary acquisition of endothermy.
Author summary Embryonic cardiomyocytes have one diploid nucleus (like most cells of the body), but most adult cardiomyocytes are polyploid. Most adult cardiomyocytes are also post-mitotic and nonregenerative, and as a result, heart injury (such as from a heart attack) is followed by scarring and impaired function rather than by regeneration. A subset of cardiomyocytes in the adult heart remains mononuclear diploid, and recent evidence indicates that this subpopulation has proliferative and regenerative capacity. Our previous work in mice showed that the percentage of this cell population in the adult heart is a complex trait subject to the combined influence of a number of polymorphic genes. One gene that influences variation in this trait is a kinase gene known as Tnni3k. This study addresses the consequences of a number of Tnni3k alleles, both newly engineered in mice and naturally occurring in a number of species, including human and mole-rat, and studied at the phenotypic and biochemical level. These results provide insight into inter- and intra-species variation in the cardiomyocyte composition of the adult heart, and may be relevant to understanding heart regenerative ability in humans and across other species.