Genotype–phenotype relationships involving hypertrophic cardiomyopathy-associated mutations in titin, muscle LIM protein, and telethonin

Abstract

Background

TTN-encoded titin, CSRP3-encoded muscle LIM protein, and TCAP-encoded telethonin are Z-disc proteins essential for the structural organization of the cardiac sarcomere and the cardiomyocyte’s stretch sensor. All three genes have been established as cardiomyopathy-associated genes for both dilated cardiomyopathy (DCM) and hypertrophic cardiomyopathy (HCM). Here, we sought to characterize the frequency, spectrum, and phenotype associated with HCM-associated mutations in these three genes in a large cohort of unrelated patients evaluated at a single tertiary outpatient center.

Methods

DNA was obtained from 389 patients with HCM (215 male, left ventricular wall thickness of 21.6 ± 6 mm) and analyzed for mutations involving all translated exons of CSRP3 and TCAP and targeted HCM-associated exons (2, 3, 4, and 14) of TTN using polymerase chain reaction (PCR), denaturing high performance liquid chromatography (DHPLC), and direct DNA sequencing. Clinical data were extracted from patient records and maintained independent of the genotype.

Results

Overall, 16 patients (4.1%) harbored a Z-disc mutation: 12 had a MLP mutation and 4 patients a TCAP mutation. No TTN mutations were detected. Seven patients were also found to have a concomitant myofilament mutation. Seven patients with a MLP-mutation were found to harbor the DCM-associated, functionally characterized W4R mutation. W4R-MLP was also noted in a single white control subject. Patients with MLP/TCAP-associated HCM clinically mimicked myofilament-HCM.

Conclusions

Approximately 4.1% of unrelated patients had HCM-associated MLP or TCAP mutations. MLP/TCAP-HCM phenotypically mirrors myofilament-HCM and is more severe than the subset of patients who still remain without a disease-causing mutation. The precise role of W4R-MLP in the pathogenesis of either DCM or HCM warrants further investigation.

Introduction

Affecting one in 500 persons, hypertrophic cardiomyopathy (HCM) is a disease associated with remarkable genotypic and phenotypic heterogeneity [1], [2]. Clinical outcomes range from an entirely asymptomatic course with normal longevity to chronic progressive heart failure or sudden cardiac death (SCD). Indeed, HCM is one of the leading causes of SCD in young persons [1].

The most common genetically mediated form of HCM is myofilament-HCM with hundreds of disease-associated mutations in eight genes encoding proteins critical to the sarcomere’s thick—[β-myosin heavy chain (MYH7) [3], regulatory myosin light chain (MYL2), and essential myosin light chain (MYL3)] [4], intermediate—[myosin binding protein C (MYBPC3)] [5], and thin myofilament [cardiac troponin T (TNNT2), α-tropomyosin (TPM1) [6], cardiac troponin I (TNNI3) [7], and actin (ACTC) [8], [9]]. Myofilament-HCM accounts for approximately 40–65% of HCM among cohorts of unrelated patients [10]. In general, patients with myofilament-HCM have greater hypertrophy and present at a younger age than those who remain without an established disease-causing mutation [11]. The two most common genotypes of myofilament-HCM, MYBPC3- and MYH7-HCM, are phenotypically indistinguishable from each other [12], [13], [14], [15], [16], [17], [18], [19], [20].

Besides perturbations involving the sarcomere’s myofilaments, the Z-disc, which comprises a cadre of proteins involved in cardiomyocyte cytoarchitecture and mechano-sensor-signaling, has emerged recently as host to several HCM-associated mutations extending the spectrum of “sarcomeric”-HCM. To date, three genes encoding critical Z-disc proteins: TTN-encoded titin, CSRP3-encoded muscle LIM protein (MLP), and the TCAP-encoded telethonin, have been implicated in the pathogenesis of both dilated cardiomyopathy (DCM) and HCM [21], [22], [23], [24].

As part of the cardiomyocyte stretch response machinery, TTN-encoded titin, which extends throughout half of the sarcomere from the M-line to the Z-disc is the largest of the three proteins; mapped on chromosome 2q31, TTN encodes for a giant 26,926 amino-acid protein with a molecular weight of 2993 kD [25]. CSRP3-encoded MLP and TCAP-encoded telethonin are mapped to chromosome 11p15.1 and 17q12, respectively, and contain 194 and 167 amino acids, respectively [26], [27]. Prior to this study, one HCM-associated mutation in TTN (R740L-TTN) [28], three HCM-associated mutations in MLP (L44P-MLP, C58G-MLP, and S54R/E55G-MLP) [22], and two HCM-associated mutations in TCAP (T137I-TCAP and R153H-TCAP) have been reported [21].

Having completed a comprehensive mutational analysis involving all translated exons of the eight genes responsible for myofilament-HCM [14], [15], [29], [30], we sought to determine the frequency, spectrum, and phenotype associated with these three genes that encode essential Z-disc proteins among a large cohort of unrelated patients diagnosed clinically with HCM.