Structural interpretation of the two-site binding of troponin on the muscle thin filament

Mak, Alan S.; Smillie, Lawrence B.

doi:10.1016/0022-2836(81)90486-1

Cited by 182 publications

(98 citation statements)

References 35 publications

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“…Furthermore, there are two regions on the tropomyosin molecule that interact with troponin T; one located in the proximity of and the other at the carboxy-terminal end of the protein (41,54,(70)(71)(72). These two troponin T-binding sites correspond to the two regions where the skeletal muscle beta-tropomyosin and rat embryonic fibroblast tropomyosin 1 differ.…”

Section: Beta-tropomyosin and Fibroblast Tropomyosin 1 Cdnas Wementioning

confidence: 99%

Nonmuscle and muscle tropomyosin isoforms are expressed from a single gene by alternative RNA splicing and polyadenylation.

Helfman¹,

Cheley²,

Kuismanen³

et al. 1986

Mol. Cell. Biol.

175

122

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The molecular basis for the expression of rat embryonic fibroblast tropomyosin 1 and skeletal muscle beta-tropomyosin was determined. cDNA clones encoding these tropomyosin isoforms exhibit complete identity except for two carboxy-proximal regions (amino acids 189 to 213 and 258 to 284) and different 3'-untranslated sequences. The isoform-specific regions delineate the troponin T-binding domains of skeletal muscle tropomyosin. Analysis of genomic clones indicates that there are two separate loci in the rat genome that contain sequences complementary to these mRNAs. One locus is a pseudogene. The other locus contains a single gene made up of 11 exons and spans approximately 10 kilobases. Sequences common to all mRNAs were found in exons 1 through 5 (amino acids 1 to 188) and exons 8 and 9 (amino acids 214 to 257). Exons 6 and 11 are specific for fibroblast mRNA (amino acids 189 to 213 and 258 to 284, respectively), while exons 7 and 10 are specific for skeletal muscle mRNA (amino acids 189 to 213 and 258 to 284, respectively). In addition, exons 10 and 11 each contain the entire 3'-untranslated sequences of the respective mRNAs including the polyadenylation site. Although the gene is also expressed in smooth muscle (stomach, uterus, and vas deferens), only the fibroblast-type splice products can be detected in these tissues. S1 and primer extension analyses indicate that all mRNAs expressed from this gene are transcribed from a single promoter. The promoter was found to contain G-C-rich sequences, a TATA-like sequence TTTTA, no identifiable CCAAT box, and two putative Sp1-binding sites.

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Section: Beta-tropomyosin and Fibroblast Tropomyosin 1 Cdnas Wementioning

confidence: 99%

Nonmuscle and muscle tropomyosin isoforms are expressed from a single gene by alternative RNA splicing and polyadenylation.

Helfman¹,

Cheley²,

Kuismanen³

et al. 1986

Mol. Cell. Biol.

175

122

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“…Based on binding studies with TM and Tn peptides, Mak and Smillie (16) proposed that the ␣-TM-Tn interaction in the region of Cys-190 is weakened in the presence of Ca 2ϩ (Ca 2ϩ -sensitive), whereas that at the COOH terminus and the overlap region is strong in the presence and absence of Ca 2ϩ (Ca 2ϩ -independent). Substantial evidence has since supported this model.…”

Section: Tropomyosins (Tm)mentioning

confidence: 99%

The Sequence of the Alternatively Spliced Sixth Exon of α-Tropomyosin Is Critical for Cooperative Actin Binding but Not for Interaction with Troponin

Hammell

Hitchcock‐DeGregori

1997

Journal of Biological Chemistry

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Tropomyosins, a family of highly conserved coiled-coil actin binding proteins, can differ as a consequence of alternative expression of several exons (Lees-Miller, J., and Helfman, D. (1991) BioEssays 13, 429 -437). Exon 6, which encodes residues 189 -213 in long, 284-residue tropomyosins, has two alternative forms, exon 6a or 6b, both highly conserved throughout evolution. In ␣-tropomyosin, exon 6a or 6b is not specific to any one of the nine isoforms. Exon 6b encodes part of a putative Ca 2؉ -sensitive troponin binding site in striated muscle tropomyosins, suggesting that the exon 6-encoded region may be specialized for certain tropomyosin functions.A series of recombinant, unacetylated tropomyosin exon 6 deletion and substitution mutants and chimeras was expressed in Escherichia coli to determine the requirements of exon 6 for tropomyosin function. Functional properties of the tropomyosins were defined by actin affinity measured by cosedimentation, troponin T affinity using a newly developed biosensor assay, and regulation of the actomyosin MgATPase. The region of tropomyosin encoded by exon 6 affects actin affinity but not thin filament assembly, troponin T binding, or regulation with troponin. The tropomyosins with exon 6a or 6b function normally whether a striated muscle exon 9a or smooth/non-muscle exon 9d is present. However, the effect of deleting 21 amino acids encoded by exon 6 or replacing it with a GCN4 leucine zipper sequence depends on the COOH-terminal sequence. Tropomyosins (TM)1 are a family of highly conserved coiledcoil actin binding proteins present in most eukaryotic cells. At least 15 different isoforms arise through the use of alternative promoters and alternative RNA splicing of the transcripts of a small number of genes (three to four in vertebrates; reviewed in Ref. 1). These isoforms are expressed in developmentally and tissue-specific patterns and differ in actin affinity.A function common to all TMs is cooperative binding to F-actin (2). Tropomyosin molecules are aligned head-to-tail in the grooves of the helical actin filament (3, 4). The role of TM is best understood in striated muscle where it regulates Ca 2ϩ -dependent muscle contraction with Tn (reviewed in Refs. 5-7). Structural studies have shown that Tn, found only in striated muscles, extends along at least the COOH-terminal third of the TM molecule (8 -13). Troponin I, TnC, and the COOH terminus of TnT are positioned near Cys-190 of TM. The elongated NH 2 terminus of TnT extends beyond the COOH terminus of one TM to the NH 2 -terminal 10 -30 residues of the next TM along the actin filament (13,14).Troponin greatly increases the affinity of TM for actin in the presence of Ca 2ϩ , with a further increase upon removal of Ca 2ϩ (15). Based on binding studies with TM and Tn peptides, Mak and Smillie (16) proposed that the ␣-TM-Tn interaction in the region of Cys-190 is weakened in the presence of Ca 2ϩ (Ca 2ϩ -sensitive), whereas that at the COOH terminus and the overlap region is strong in the presence and absence of Ca 2ϩ (...

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“…In particular, Ca 2ϩ weakens the actin binding of the troponin subunit, TnI, 1 and the tropomyosin binding of the carboxyl-terminal region of TnT. In this widely held scheme of thin filament function, troponin remains anchored to the thin filament by relatively Ca 2ϩ -insensitive binding of the aminoterminal region of TnT to the carboxyl-terminal region of tropomyosin (Pato, et al, 1981;Mak and Smillie, 1981;Tanokura, et al, 1983;Cho and Hitchcock-DeGregori, 1990;Ishii and Lehrer, 1991). Indeed, rotary-shadowed electron micrographs of troponin and of TnT (Flicker, et al, 1982) show a highly extended molecule with a narrow tail mostly attributable to the amino-terminal portion of TnT (White et al, 1987) and a distinctly more globular head region that is likely to include portions of all three subunits.…”

mentioning

confidence: 99%

NH2-terminal Truncation of Skeletal Muscle Troponin T Does Not Alter the Ca2+ Sensitivity of Thin Filament Assembly

Fisher

Wang

Tobacman

1995

Journal of Biological Chemistry

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Structural interpretation of the two-site binding of troponin on the muscle thin filament

Cited by 182 publications

References 35 publications

Nonmuscle and muscle tropomyosin isoforms are expressed from a single gene by alternative RNA splicing and polyadenylation.

Nonmuscle and muscle tropomyosin isoforms are expressed from a single gene by alternative RNA splicing and polyadenylation.

The Sequence of the Alternatively Spliced Sixth Exon of α-Tropomyosin Is Critical for Cooperative Actin Binding but Not for Interaction with Troponin

NH2-terminal Truncation of Skeletal Muscle Troponin T Does Not Alter the Ca2+ Sensitivity of Thin Filament Assembly

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