Myofibril-Inducing RNA (MIR) is essential for tropomyosin expression and myofibrillogenesis in axolotl hearts

The Mexican axolotl, Ambystoma mexicanum, has proven to be a unique animal model in the study of cardiac development. The axolotl (a neotenous salamander) carries a naturally occurring recessive mutation, gene c, first discovered and characterized by Humphrey [1], which results in abnormal cardiac development in homozygous recessive "c/c" embryos. The mutant embryonic hearts develop, but fail to beat, making them distinguishable from normal embryonic hearts which start to beat at stage 35. The myocytes of the mutant hearts fail to form organized myofibrils and the embryos survive only to stage 42, the hatching stage, due to a lack of circulation.Among the various myofibril structural proteins, tropomyosin has been shown by SDS-PAGE [2], radio-immunoassay [3], 2D gel electrophoresis [4] and confocal microscopy of whole hearts to be drastically reduced in the mutants [5-7]. Interestingly, other myofibril structural proteins such as actin, myosin and myosin binding protein C, however, were found to be at or near normal levels in the mutant hearts [8-10].Using this animal model, Myofibril-Inducing RNA (MIR), a small bioactive RNA, was shown in previous studies to be able to restore tropomyosin protein synthesis, promote myofibrillogenesis, and initiate heartbeat in the mutant embryonic hearts in organ culture [11]. The MIR appears to function through its unique secondary structure since it is a non-coding RNA [7,11].In mammals, birds and amphibians, altogether four different types of tropomyosin genes have been identified: alpha gene (TPM1), beta gene (TPM2), gamma gene (TPM3) and TM4 type gene (TPM4) [12]. More recently, TM4, a cytoskeletal tropomyosin, also has been associated with growth and regeneration in response to injury, disease state and stress in skeletal muscle of mouse and humans [13]. Moreover, in zebrafish embryos, a heart specific isoform of TM4 is essential for normal myofibril formation and developing a heartbeat [14]. In addition, it has been found re