Interaction of Contractile Proteins with DNA

Abstract

The interaction of contractile proteins (myosin, actin, tropomyosin and troponin) with DNA was studied in vitro using a nitrocellulose filter binding technique. The data indicate a high affinity of myosin and troponin for DNA, a lesser interaction between DNA and tropomyosin and the absence of binding of actin to DNA. When binding to DNA was detected, the interaction was higher with single-stranded DNA than with RNA or double-stranded DNA, although in some conditions myosin binds equally as well to native as to denatured eukaryotic DNA.Myosin binds better to eukaryotic than to phage native DNA.The characterization of non-histone chromatin proteins and their possible correlation with proteins of known biological functions is a current problem. There is some evidence that contractile proteins, such as myosin, actin, and others [I -41, exist among the nonhistone chromatin proteins. These proteins are present not only in the muscle but in other tissues; for example, myosin, tropomyosin, actin and a troponin C subunit-like protein have been found in the cytoplasmic and nuclear fractions from fibroblasts [5 -81. Other authors have also reported the existence of myosin and actin in the mitotic spindle [9,10], and recently Cande et al. [ l l ] have shown the existence of actin in the fibers running from the kinetochore of the chromosome to the pole in the mitotic spindle. This evidence suggests the possibility of the interaction of contractile proteins with the chromosome. This interaction could occur during the whole cell cycle or, perhaps, in one phase only.If a protein is associated to chromatin it must have affinity for DNA, another chromosomal protein or other chromosomal components. Therefore, the affinity for purified DNA can give some insight on the possible interaction of a protein with chromatin. In this report we show some DNA-binding properties of contractile proteins (myosin, actin, tropomyosin and troponin) isolated from muscle microfilaments. Our results indicate that troponin and myosin have a high affinity for DNA, tropomyosin has a weaker interaction with DNA and actin does not bind to DNA. Abhrericrzion. EGTA, [ethylenebis(oxyethylenenitrilo)]tetraace-Enzjwes. Myosin ATPase (EC 3.6.1.3); nuclease Sl (EC tic acid. 3.1.4.21).Moreover, we show that myosin binds better to eukaryotic than to phage DNA. MATERIALS AND METHODS Protein PreparationsMyosin was purified from pig leg muscle following the procedure of Richards et al. [12]. Ground muscle was extracted with a solution containing 0.67 M KC1, 5 mM KH,PO,/Na,HPO,, 5 mM MgCl, and 10 mM sodium pyrophosphate. Myosin was precipitated by adding 9 vol. of water, resuspended in 0.3 M KC1 and dialyzed against 0.15 M potassium phosphate, pH 7.5 (25 "C). 40 mg of protein were chromatographed on a 15-ml DEAE-Sephadex column as described by Godfrey and Harrington [13]. The eluted myosin was dialyzed against 0.15 M potassium phosphate, pH 7.5, 10 mM EDTA, and 50% glycerol and stored at -20 "C. The usual specific activity of the myosin ATPase was approximately 50 nm...

Microtubule-associated protein MAP2 preferentially binds to a dA/dT sequence present in mouse satellite DNA.

“…This might explain why, in the division of hybrid cells, the chromosomes from different species are segregated by the microtubules of the same cell, as well as the preferential segregation of certain chromosomes. We also would like to discuss the results presented here with respect to two previous results: (a) MAP2 preferentially binds to the synthetic polymer poly(dA)/poly(dT) and, in the presence of this polymer, the critical concentration needed for tubulin assembly is lower than in the presence of other synthetic polymers, including poly(dAdT)/poly(dAdT) (Corces et al, 1980); (b) MAP2 or related proteins are not only present in brain but also in non-neural tissues (Valdivia et al, 1982). These Fig.…”

Cytoplasmic nucleic acid-based XNAs directly enhance live cardiac cell function by a Ca2+ cycling-independent mechanism via the sarcomere

“…1B), provides evidence that ssDNAs have a direct effect on the myofilamentbased contractile apparatus. To further test this possibility, we determined whether XNAs could bind with high affinity to purified sarcomeric proteins, as suggested previously 25 . Fluorescence polarization and capillary electrophoresis data showed low nM affinity of random sequence ssDNA to both the bovine and human sarcomeric troponin complex (Fig.…”