Mutation that dramatically alters rat titin isoform expression and cardiomyocyte passive tension

Abstract

Titin is a very large alternatively spliced protein that performs multiple functions in heart and skeletal muscle. A rat strain is described with an autosomal dominant mutation that alters the isoform expression of titin. While wild type animals go through a developmental program where the 3.0 MDalton N2B becomes the major isoform expressed by two to three weeks after birth (~85%), the appearance of the N2B is markedly delayed in heterozygotes and never reaches more than 50% of the titin in the adult. Homozygote mutants express a giant titin of the N2BA isoform type (3.9 MDalton) that persists as the primary titin species through ages of more than one and half years. The mutation does not affect the isoform switching of troponin T, a protein that also is alternatively spliced with developmental changes. The basis for the apparently greater size of the giant titin in homozygous mutants was not determined, but additional length was not due to inclusion of sequence from larger numbers of PEVK exons or the Novex III exon. Passive tension measurements using isolated cardiomyocytes from homozygous mutants showed that cells could be stretched to sarcomere lengths greater than 4 µm without breakage. This novel rat model should be useful for exploring the potential role of titin in the Frank-Starling relationship and mechano-sensing/signaling mechanisms.

Historical perspective on heart function: the Frank–Starling Law

“…Along these lines, a correlation between enhanced length-dependent activation and higher levels of passive tension has been reported (Cazorla et al 1999(Cazorla et al , 2001Fukuda et al 2003). A recent study reporting a reduced myofilament force development and impaired length-dependent activation (Mateja et al 2012) in a rat with a homozygous autosomal mutation expressing a giant titin isoform (N2BA-G,~3.9 MDa) (Greaser et al 2008) appears to support this idea.…”

Nuances of electrophoresis study of titin/connectin

“…Giant titin isoforms expressed in rat striated muscles with an RBM20 autosomal dominant mutation were recently reported (Greaser et al 2008;Li et al 2012;Guo et al 2012). The molecular masses of these isoforms were estimated from their electrophoretic mobility in 1% vertical agarose gel to be 3750 kDa and 3830 kDa ).…”

The Giant Protein Titin: A Regulatory Node That Integrates Myocyte Signaling Pathways

“…The functional implications of these isoform transitions have been discussed elsewhere (12). Interestingly, an unusually large, ϳ3900-kDa, cardiac N2BA titin has been identified in a spontaneous mutant rat model, but the origin of the 200-kDa "extra" mass in this isoform is unknown (19).…”