Aberrant developmental titin splicing and dysregulated sarcomere length in Thymosin β4 knockout mice

Nicola Smart,Johannes Riegler,Cameron W Turtle,Craig A Lygate,Debra J Mcandrew,Katja Gehmlich,Karina N Dubé,Anthony N Price,Vivek Muthurangu,Andrew M Taylor,Mark F Lythgoe,Charles Redwood,Paul R Riley

doi:10.1016/j.yjmcc.2016.10.010

Nicola Smart, Johannes Riegler + Show 11 more

Open Access

https://doi.org/10.1016/j.yjmcc.2016.10.010

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Abstract

Sarcomere assembly is a highly orchestrated and dynamic process which adapts, during perinatal development, to accommodate growth of the heart. Sarcomeric components, including titin, undergo an isoform transition to adjust ventricular filling. Many sarcomeric genes have been implicated in congenital cardiomyopathies, such that understanding developmental sarcomere transitions will inform the aetiology and treatment. We sought to determine whether Thymosin β4 (Tβ4), a peptide that regulates the availability of actin monomers for polymerization in non-muscle cells, plays a role in sarcomere assembly during cardiac morphogenesis and influences adult cardiac function. In Tβ4 null mice, immunofluorescence-based sarcomere analyses revealed shortened thin filament, sarcomere and titin spring length in cardiomyocytes, associated with precocious up-regulation of the short titin isoforms during the postnatal splicing transition. By magnetic resonance imaging, this manifested as diminished stroke volume and limited contractile reserve in adult mice. Extrapolating to an in vitro cardiomyocyte model, the altered postnatal splicing was corrected with addition of synthetic Tβ4, whereby normal sarcomere length was restored. Our data suggest that Tβ4 is required for setting correct sarcomere length and for appropriate splicing of titin, not only in the heart but also in skeletal muscle. Distinguishing between thin filament extension and titin splicing as the primary defect is challenging, as these events are intimately linked. The regulation of titin splicing is a previously unrecognised role of Tβ4 and gives preliminary insight into a mechanism by which titin isoforms may be manipulated to correct cardiac dysfunction.

Highlights

The contractile properties of cardiac muscle influence the filling behaviour of the heart, determining the maximal diastolic volume, thereby providing a set point for systolic performance via the Frank-Starling mechanism [1]
We reveal that mice lacking the actin monomer binding protein, Thymosin β4 (Tβ4), fail to appropriately regulate sarcomere length, due to a precocious titin isoform transition postnatally and predominance of the less compliant isoforms through to adulthood
Under conditions of altered cardiac demand, Tβ4 −/Y mice were limited in their ability to increase heart rate and ejection fraction in response to the β1-adrenergic receptor agonist dobutamine

Summary

Introduction

The contractile properties of cardiac muscle influence the filling behaviour of the heart, determining the maximal diastolic volume, thereby providing a set point for systolic performance via the Frank-Starling mechanism [1]. Protein components, which dynamically adapts throughout perinatal heart development to accommodate physiological hypertrophic cardiac growth During this period, various myocardial proteins, including myosin heavy chain (MyHC) [4], troponins [5], tropomyosin [6] and titin [7], undergo an isoform switch from foetal to adult type in order to adjust ventricular filling. Titin acts as a template to ensure the regular interdigitation and centring of thick and thin filaments; it undergoes significant isoform transition within the first few weeks of postnatal life, from the longer, more compliant foetal/N2BA isoforms to a predominance of the shorter, stiffer N2B isoform [15] This coincides with the period of thin filament elongation to set adult sarcomere length according to the extensibility of the predominant titin isoform [2,15]

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