Cardiac Troponin I Gene Knockout

Abstract

—Troponin I is a subunit of the thin filament–associated troponin-tropomyosin complex involved in calcium regulation of skeletal and cardiac muscle contraction. We deleted the cardiac isoform of troponin I by using gene targeting in murine embryonic stem cells to determine the developmental and physiological effects of the absence of this regulatory protein. Mice lacking cardiac troponin I were born healthy, with normal heart and body weight, because a fetal troponin I isoform (identical to slow skeletal troponin I) compensated for the absence of cardiac troponin I. Compensation was only temporary, however, as 15 days after birth slow skeletal troponin I expression began a steady decline, giving rise to a troponin I deficiency. Mice died of acute heart failure on day 18, demonstrating that some form of troponin I is required for normal cardiac function and survival. Ventricular myocytes isolated from these troponin I–depleted hearts displayed shortened sarcomeres and elevated resting tension measured under relaxing conditions and had a reduced myofilament Ca sensitivity under activating conditions. The results show that (1) developmental downregulation of slow skeletal troponin I occurs even in the absence of cardiac troponin I and (2) the resultant troponin I depletion alters specific mechanical properties of myocardium and can lead to a lethal form of acute heart failure.

Deletion of a Genomic Segment Containing the Cardiac Troponin I Gene Knocks Down Expression of the Slow Troponin T Gene and Impairs Fatigue Tolerance of Diaphragm Muscle

“…This functional importance might have been responsible for the ssTnT gene remaining closely linked to the upstream cTnI gene during evolution, because a destruction of either the 3Ј-coding exons of the cTnI gene or the 5Ј-enhancer region of the ssTnT gene would cause infantile lethality (5,7).…”

“…The genotyping of the double transgenic offspring was performed on genome DNA isolated from tail biopsies using PCR, and the compensation for the loss of endogenous cTnI by cTnI-ND in cardiac muscle was confirmed by Western blotting as described (7,9) (Fig. 2).…”

“…Shown in ex vivo promoter analysis, the 5Ј upstream enhancer region of the ssTnT gene overlaps with the structure of the cTnI gene (6). This structural integration of cTnI and ssTnT genes suggested that a previously developed mouse line in which the entire Tnni3 gene was deleted through embryonic stem cell gene targeting (7) would have lost a large portion of the 5Ј-enhancer region of the Tnnt1 gene (Fig. 1).…”

Muscle-specific RING finger 1 is a bona fide ubiquitin ligase that degrades cardiac troponin I

“…Nevertheless, the effects of MuRF1 on contractility are consistent with the reported effects of loss of troponin I function. Deletion of troponin I results in decreased contractility and cardiac dysfunction in mice (33), and a recessive mutation in troponin I that disrupts assembly of the troponin complex also results in a contractility defect in humans (34). It is tempting to speculate that disassembly of sarcomeres by molecules such as MuRF1 may be a maladaptive response that contributes to wall thinning and contractile dysfunction in dilated cardiomyopathies.…”

The Role of Troponins in Muscle Contraction

“…The protein isoforms and their corresponding mRNAs follow the same pattern of accumulation, suggesting that the transition in troponin expression is regulated at the level of gene transcription (45). A mouse model lacking CTnI demonstrated that a fetal TnI isoform that is identical to STnI, compensated for the absence of CTnI up to 15 days after birth and around day 18, the mice died of acute heart failure (46). This suggested that some form of TnI is required for normal cardiac function and survival.…”