Disease relevance of TTN

• Expression of one cDNA clone in Escherichia coli produced a fusion protein that reacted specifically with titin antibodies, and titin antiserum affinity-purified against this fusion protein reacted specifically with titin on immunoblots of chicken cardiac and skeletal muscle myofibrils [1].

High impact information on TTN

• We have found that avian vascular and visceral smooth muscles contain a novel, megadalton protein, smitin, that is similar to striated muscle titin in molecular morphology, localization in a contractile apparatus, and ability to interact with myosin filaments [2].
• Recruitment of all three elements allows cardiac titin to extend fully reversibly at physiological sarcomere lengths, without the need to unfold Ig domains [3].
• I-band titin in cardiac muscle is a three-element molecular spring and is critical for maintaining thin filament structure [3].
• In Western analysis, a polyclonal antibody raised against brush border T-protein reacted specifically with T-protein in isolated brush borders and cross-reacted with titin in pectoralis and cardiac muscle samples [4].
• Identification and characterization of two huge protein components of the brush border cytoskeleton: evidence for a cellular isoform of titin [4].

Biological context of TTN

• Differences between the rabbit and chicken titin sequences included a unique, serine-rich region in one motif, which represents a potential phosphorylation site [1].
• To characterize the primary structure of titin, three overlapping cDNA clones comprising 2.4 kilobases of avian muscle titin coding sequence were obtained from a cDNA library constructed from embryonic chick cardiac muscle RNA size-selected for large transcripts [1].
• Transfections of primary cultures of embryonic chick cardiomyocytes demonstrated that the z-repeats play the major role in targeting titin fragments to the Z-band [5].
• Immunocytochemical studies of cardiac myofibrillogenesis in early chick embryos. I. Presence of immunofluorescent titin spots in premyofibril stages [6].

Anatomical context of TTN

• Titin is an approximately 3000-kDa polypeptide that constitutes a set of elastic filaments that connect thick filaments to the Z-line in vertebrate striated muscle myofibrils [1].
• Connectin/titin, a 3000 kDa protein, links the Z line to the myosin filament in striated muscle sarcomeres [7].
• No readily discernable specific ultrastructural features were found in titin-positive regions of the 6 somite stage cardiac primodia [8].
• These observations suggest that newly synthesized titin molecules are stable proteins that rapidly associate with the cytoskeleton of developing myotubes [9].
• These preparations and sections revealed that titin, a putative scaffolding protein of sarcomeres, is present in a punctate state and also in a diffuse form throughout the cytoplasm of cardiac myocytes in the premyofibril stages (4-7 somite stages) as well as in the early stages of myofibril formation [6].

Associations of TTN with chemical compounds

• In whole myofibril preparations from skeletal muscles, troponin-C, connectin (titin), and nebulin were intensely fluorescent, being shown to have calcium-binding ability [10].
• Pressure above 2 kbar apparently generated a flexible hydrophobic pocket for tryptophan residues of titin [11].
• Titin antibodies bound to the nitrocellulose immobilized native antigen were directly conjugated with fluorescein isothiocyanate [12].
• Phosphorylation of beta-connectin (titin 2), an elastic protein of chicken breast muscle, occurred in the presence of [gamma-32P] ATP, 0.2 mM CaCl2 and 25 mM phosphate buffer, pH 7 [13].
• In contrast, in C3 exoenzyme treated cells, focal adhesions were disassembled and costamere were absent; in addition, beta-actin-positive, non-striated fibrils were lost and assembly of M-protein, titin, and alpha-actinin into myofibrils was poor, as shown by diffuse and filamentous staining pattern [14].

Physical interactions of TTN

• In striated muscle sarcomeres, interactions between the myosin-binding protein titin and alpha-actinin in the Z-line provide an important structural linkage [15].
• These smitin-binding sites are highly homologous to the titin-binding sites of striated muscle alpha-actinin [15].

Other interactions of TTN

• Within these titin spots, smaller alpha-actinin dots were observed [8].
• When the tropomyosin 3' UTR is expressed in osteoblasts, less skeletal muscle tropomyosin is expressed, and titin expression is delayed [16].
• This indicates that M-protein, along with other thick filament-associated proteins such as C-protein, twichin, and titin, belongs to the superfamily of cytoskeletal proteins with immunoglobulin/fibronectin repeats [17].
• We suggest that assembly of thin and thick filament occurs independently during myofibrillogenesis in situ and that myomesin might be important for integrating thick filaments with the M-line end of titin [18].
• Western blot analysis of these samples revealed several fragments of myosin heavy chain, M-protein, and titin [19].

Analytical, diagnostic and therapeutic context of TTN

• Indirect immunofluorescence localization with the fusion protein-specific antibodies demonstrated that the cDNA sequence was from the region of titin located in the myofibrillar A-band adjacent to the A/I junction [1].
• Immunoelectron microscopy of myofibrils identifies both titin-associated proteins as M band constituents [20].
• TII, the extractable form of titin, was purified from myofibrils and separated by high resolution gel permeation chromatography into two fractions (TIIA and TIIB) [20].
• Using pulse-labeling with [35S]methionine and immunoprecipitation with an anti-titin mAb, we have examined the biosynthesis of titin in synchronized cultures of skeletal muscle cells derived from day 12 chicken embryos [9].
• We determined the actual number of titin molecules in a unit cell from scanning transmission electron microscopy mass measurements of end-filaments [21].

References