TNNI3K is a novel mediator of myofilament function and phosphorylates cardiac troponin I

Hui Wang,Yan-Wan Zhang,Rui-Xia Xu,Lin Wang,Li Song,Xian-Min Meng,Na Shi,Jue Ye

doi:10.1590/1414-431x20122515

Abstract

The phosphorylation of cardiac troponin I (cTnI) plays an important role in the contractile dysfunction associated with heart failure. Human cardiac troponin I-interacting kinase (TNNI3K) is a novel cardiac-specific functional kinase that can bind to cTnI in a yeast two-hybrid screen. The purpose of this study was to investigate whether TNNI3K can phosphorylate cTnI at specific sites and to examine whether the phosphorylation of cTnI caused by TNNI3K can regulate cardiac myofilament contractile function. Co-immunoprecipitation was performed to confirm that TNNI3K could interact with cTnI. Kinase assays further indicated that TNNI3K did not phosphorylate cTnI at Ser23/24 and Ser44, but directly phosphorylated Ser43 and Thr143 in vitro. The results obtained for adult rat cardiomyocytes also indicated that enhanced phosphorylation of cTnI at Ser43 and Thr143 correlated with rTNNI3K (rat TNNI3K) overexpression, and phosphorylation was reduced when rTNNI3K was knocked down. To determine the contractile function modulated by TNNI3K-mediated phosphorylation of cTnI, cardiomyocyte contraction was studied in adult rat ventricular myocytes. The contraction of cardiomyocytes increased with rTNNI3K overexpression and decreased with rTNNI3K knockdown. We conclude that TNNI3K may be a novel mediator of cTnI phosphorylation and contribute to the regulation of cardiac myofilament contraction function.

Highlights

The human cardiac troponin I-interacting kinase (TNNI3K) gene expresses a novel cardiac-specific functional kinase
The results showed that TNNI3K coimmunoprecipitated cardiac troponin I (cTnI), while no significant bands were detected in the control (Figure 1)
As an important post-translational modification, phosphorylation of cTnI and other myofilament proteins plays a major role in the dynamic modulation of contractile function and in the regulation of thin filament function in the transition from compensated hypertrophy to heart failure [6,11,18,19]

Summary

Introduction

The human cardiac troponin I-interacting kinase (TNNI3K) gene expresses a novel cardiac-specific functional kinase. It was first cloned from an adult heart cDNA library based on large-scale expressed sequence tag sequencing [1]. The TNNI3K protein contains three domains, including seven N-terminal ankyrin repeats, a protein kinase (PK) domain that contains motifs conserved in both serine/threonine and tyrosine PKs, and a C-terminal Ser-rich domain. TNNI3K belongs to a new family of kinases, called the mixed lineage kinase family in the tyrosine kinase-like group, based on a sequence comparison of the catalytic domain together with knowledge of sequence similarity and domain structures outside the catalytic domain [2]. The TNNI3K C-terminal Ser-rich domain was previously used as bait to perform a yeast two-hybrid screen of a human cardiac library, and cardiac troponin I (cTnI) was identified as a TNNI3K-interacting protein [1]

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