Catchin, a novel protein in molluscan catch muscles, is produced by alternative splicing from the myosin heavy chain gene

Yamada, Akira; Matsumoto, Yoshio; Oiwa, Kazuhiro; Nyitray, László

doi:10.1006/jmbi.1999.3349

Cited by 56 publications

(45 citation statements)

References 35 publications

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“…Peptide Synthesis-The myosin loop 2 peptide of ABRM myosin heavy chain (residues 390 -408, CAQNKEAETTGTH-KKRKSSA) was synthesized (AJ249991) (14) with an additional Cys at the N terminus (M r 2175). The actin subdomain I peptide, AGFAGDDAP, corresponding to residues 21-29 of chicken skeletal actin, was synthesized (M r 819).…”

Section: Methodsmentioning

confidence: 99%

Myosin Loop 2 Is Involved in the Formation of a Trimeric Complex of Twitchin, Actin, and Myosin

Funabara

Osawa

Ueda

et al. 2009

Journal of Biological Chemistry

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Molluscan smooth muscles, such as mussel anterior byssus retractor muscle (ABRM) 2 and adductor muscle, exhibit a low energy cost phase of tension maintenance termed catch. Catch muscle develops active tension following an increase of the intracellular [Ca 2ϩ ] induced by secretion of acetylcholine. Myosin is activated by direct binding of Ca 2ϩ to the regulatory myosin light chain and initiates a relative sliding between thick and thin filaments (1). After a decrease of intracellular [Ca 2ϩ ] to resting levels, the catch state is formed where tension is maintained over long periods of time with little energy consumption (2, 3). Catch tension is abolished by secretion of serotonin and an increase of intracellular [cAMP] with the resulting activation of cAMP-dependent protein kinase and phosphorylation of twitchin (4, 5). Twitchin phosphorylation is required for relaxation of the muscle from catch. For this cycle to repeat, dephosphorylation of twitchin is necessary (6). Thus, in this scheme, twitchin is a major regulator of the catch state.Molluscan twitchin is known as a myosin-binding protein belonging to the titin/connectin superfamily. It is a single polypeptide of 530 kDa containing multiple repeats of immunoglobulin (Ig) and fibronectin type 3-like motifs in addition to a single kinase domain homologous to the catalytic domain of myosin light chain kinase of vertebrate smooth muscle (7). There are several possible phosphorylation sites in molluscan twitchin recognized by cAMP-dependent protein kinase, and two, D1 and D2, have been identified. The D1 phosphorylation site (Ser-1075) is in the linker region between the 7th and 8th Ig motifs (numbering from the N terminus). The D2 site (Ser-4316) is in the linker region between the 21st and 22nd Ig motifs. Additional sites are found close to D1, but are thought not to be vital for catch regulation.The molecular mechanisms underlying development and maintenance of the catch state have been controversial for several years. One theory proposes that catch reflected attached frozen or slowly cycling cross-bridges (8, 9). What distinguished the attached cross-bridge from the detached relaxed state is not clear. Also it was suggested that interactions between thick filaments, other than cross-bridges, or between thin and thick filaments are responsible for the catch contraction (10). In either of the latter cases, the cross-bridge (myosin head) was not involved.Recently we found that a twitchin fragment including the D2 phosphorylation site and its flanking Ig motifs (TWD2-S) interacted with myosin and actin in a phosphorylation-sensitive manner, and it was suggested that this trimeric complex contributed to tension maintenance in catch (11). TWD2-S bound to a region of the actin molecule known also to interact with loop 2 of myosin that is involved in the ATP-driven movement of myosin with actin (12). In the present study, we show that the myosin loop 2 binds to TWD2-S using competitive cosedimentation assays and isothermal titration calorimetry (ITC). Th...

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Section: Methodsmentioning

confidence: 99%

Myosin Loop 2 Is Involved in the Formation of a Trimeric Complex of Twitchin, Actin, and Myosin

Funabara

Osawa

Ueda

et al. 2009

Journal of Biological Chemistry

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“…This protein consists almost entirely of coiled coil and can co-assemble into thick filaments. Similarly, use of an alternative promotor in a molluscan myosin II gene leads to the transcription of catchin, a rod-like protein found in catch muscles (Yamada et al, 2000). In both of these examples, the exons that contain initiation methionine codons are found within large introns of the corresponding muscle myosin genes, and both proteins contain unique N-terminal domains.…”

Section: Discovery Of a Headless Myo10 In Brainmentioning

confidence: 99%

Myo10 in brain: developmental regulation, identification of a headless isoform and dynamics in neurons

Sousa

Berg

Robertson

et al. 2006

Journal of Cell Science

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Although Myo10 (myosin-X) is an unconventional myosin associated with filopodia, little is known about its isoforms and roles in the nervous system. We report here that, in addition to full-length Myo10, brain expresses a shorter form of Myo10 that lacks a myosin head domain. This `headless' Myo10 is thus unable to function as a molecular motor, but is otherwise identical to full-length Myo10 and, like it, contains three pleckstrin homology (PH) domains, a myosin-tail homology 4 (MyTH4) domain, and a band-4.1/ezrin/radixin/moesin (FERM) domain. Immunoblotting demonstrates that both full-length and headless Myo10 exhibit dramatic developmental regulation in mouse brain. Immunofluorescence with an antibody that detects both isoforms demonstrates that Myo10 is expressed in neurons, such as Purkinje cells, as well as non-neuronal cells, such as astrocytes and ependymal cells. CAD cells, a neuronal cell line, express both full-length and headless Myo10, and this endogenous Myo10 is present in cell bodies, neurites, growth cones and the tips of filopodia. To investigate the dynamics of the two forms of Myo10 in neurons, CAD cells were transfected with GFP constructs corresponding to full-length or headless Myo10. Only full-length Myo10 localizes to filopodial tips and undergoes intrafilopodial motility, demonstrating that the motor domain is necessary for these activities. Live cell imaging also reveals that full-length Myo10 localizes to the tips of neuronal filopodia as they explore and interact with their surroundings, suggesting that this myosin has a role in neuronal actin dynamics.

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“…They consisted of the main components of native thick filaments twitchin, myosin (a heavy chain and light chains), catchin (16), and paramyosin, with a small amount of actin (Fig. 4B, lane 1).…”

Section: Reconstitution Of Catch Statementioning

confidence: 99%

“…These in vitro experiments are much more useful than those using muscle fibers to identify components essential for the catch state and to reveal phosphorylation/dephosphorylation cascades relaying signals onward since the catch state can be reconstituted with purified proteins, and each interaction is separately analyzed without interference of other components. Indeed, our group showed that the myofilament components essential for the catch state are myosin, twitchin, and actin, whereas other components such as paramyosin, catchin (16), and tropomyosin are not (15).…”

mentioning

confidence: 99%

Protein Phosphatase 2B Dephosphorylates Twitchin, Initiating the Catch State of Invertebrate Smooth Muscle

Yamada

Matsumoto

Nakamura

et al. 2004

Journal of Biological Chemistry

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"Catch" is the state where some invertebrate muscles sustain high tension for long periods at low ATP hydrolysis rates. Physiological studies using muscle fibers have not yet fully provided the details of the initiation process of the catch state. The process was extensively studied by using an in vitro reconstitution assay with several phosphatase inhibitors. Actin filaments bound to thick filaments pretreated with the soluble protein fraction of muscle homogenate and Ca 2؉ (catch treatment) in the presence of MgATP at a low free Ca 2؉ concentration (the catch state). Catch treatment with >50 M okadaic acid, >1 M microcystin LR, 1 M cyclosporin A, 1 M FK506, or 0.2 mM calcineurin autoinhibitory peptide fragment produced almost no binding of the actin filaments, indicating protein phosphatase 2B (PP2B) was involved. Use of bovine calcineurin (PP2B) and its activator calmodulin instead of the soluble protein fraction initiated the catch state, indicating that only PP2B and calmodulin in the soluble protein fraction are essential for the initiation process. The initiation was reproduced with purified actin, myosin, twitchin, PP2B, and calmodulin. 32 P autoradiography showed that only twitchin was dephosphorylated during the catch treatment with either the soluble protein fraction or bovine calcineurin and calmodulin. These results indicate that PP2B directly dephosphorylates twitchin and initiates the catch state and that no other component is required for the initiation process of the catch state.Invertebrate muscles such as the smooth adductor and byssus retractor muscles of bivalves maintain high tension with little energy expenditure after active contractions (1, 2). This high tension state, called the "catch state," is terminated by a neurotransmitter, 5-hydroxytryptamine (3), activating cAMPdependent protein kinase (PKA) 1 through an increase in the intracellular concentration of cAMP. When a catalytic subunit of PKA was added to chemically demembranated catch muscles, the passive tension of the catch state diminished (4). Phosphorylation of a target protein(s) by this kinase thus causes relaxation of the catch tension.Twitchin is a candidate for this target protein. Originally identified in the nematode Caenorhabditis elegans (5, 6), it is also called "mini-titin" and is abundantly present in molluscan muscles (7,8). Studies with the anterior byssus retractor muscle of the mussel suggested that its phosphorylation by PKA was involved in the relaxation of the catch state (9). The catalytic subunit of PKA phosphorylates 2-3 serine residues of the mussel catch muscle twitchin (10, 11). In contrast to the relaxation of the catch state, the regulation of the reverse reaction, i.e. initiation of the catch state, has not been well characterized yet, since the reaction in vivo seems to occur in parallel with the activation of muscle contraction. Therefore, how initiation signals are relayed onward and amplified in cascades and which types of phosphatases dephosphorylate twitchin to initiate the catch state are stil...

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Catchin, a novel protein in molluscan catch muscles, is produced by alternative splicing from the myosin heavy chain gene

Cited by 56 publications

References 35 publications

Myosin Loop 2 Is Involved in the Formation of a Trimeric Complex of Twitchin, Actin, and Myosin

Myosin Loop 2 Is Involved in the Formation of a Trimeric Complex of Twitchin, Actin, and Myosin

Myo10 in brain: developmental regulation, identification of a headless isoform and dynamics in neurons

Protein Phosphatase 2B Dephosphorylates Twitchin, Initiating the Catch State of Invertebrate Smooth Muscle

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