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C Klotz

Publications and source records attributed to C Klotz.

36 records · Page 2Linked to original sources

A protein of 175,000 daltons associated with striated rootlets in ciliated epithelia, as revealed by a monoclonal antibody.

Basal bodies from laying quail oviduct were semipurified and used as immunogen to produce monoclonal antibodies. On 38 clones obtained and among those staining the apical pole of the ciliated cell, CC-310 was chosen because it labeled the apical region with a punctuated aspect, suggesting a staining of basal bodies or of basal body-associated structures; the basal pole was also labeled. The ultrastructural localization performed by the immunogold technique showed that the labeling was mainly associated with the striated rootlets. The basal feet, the side of the basal bodies, and the basal poles of the demembranated cells were also decorated. The identification of the antigen performed by immunoblots of deciliated cortices revealed two proteins of 175,000 and 40,000, whereas immunoblots of basal bodies showed only the 175,000-mw protein. The possibility of these two proteins sharing the same epitope, located at both poles of the cell, is discussed. Immunofluorescence ascertained that CC-310 decorated the striated rootlets in ciliated epithelia from other species: mussel, frog, and human tissue. Finally, when tested on cultured cell lines, CC-310 labeled the centrosome and its associated rootlets on PtK2 during interphase. During mitosis the poles of the mitotic spindle were stained without any apparent rootlet-like structure.

Animals↗

Myosin at the apical pole of ciliated epithelial cells as revealed by a monoclonal antibody.

A monoclonal antibody (CC-212), obtained in a fusion experiment in which basal bodies from quail oviduct were used as immunogen, has been shown to label the apical pole of ciliated cells and to react with a 200-kD protein. This monoclonal antibody was demonstrated to be an anti-myosin from smooth muscle or from nonmuscular cells using the following criteria: On Western blots it reacted with the myosin heavy chains from gizzard and platelet extracts and from cultured cell line extracts, but did not react with striated muscle myosin heavy chains. By immunofluorescence it decorated the stress fibers of well-spread cells with a characteristic striated pattern, while it did not react with myotubes containing organized myofibrils. On native ciliated cells as well as on Triton-extracted ciliated cortices from quail oviduct, this monoclonal antibody decorated the apical pole with a stronger labeling of the periphery of the apical area. Ultrastructural localization was attempted using the immunogold technique on the same preparation. Myosin was associated with a filamentous material present between striated rootlets and the proximal extremities of the basal bodies. No labeling of the basal body itself or of axoneme was observed.

Animals↗

Comparative sequence of myosin light chains from normal and hypertrophied human hearts.

Myosin light chains from normal and hypertrophied human hearts were partially sequenced in order to see whether structural modifications of these light subunits could provide a molecular basis for the changes observed in heart properties and in myosin enzymatic activity. Normal light chains were prepared form hearts taken at autopsy, weighing 350 g or less and apparently devoid of myocardial disease. "Hypertrophied cardiac myosin light chains" were prepared from two greatly hypertrophied hearts, weighing 600 and750 g. No amino acid substitutions, deletions, or additions were observed in the light chains from hypertrophied hearts. The third light chain previously reported in human cardiac myosin and related to hypertrophy was found to be a proteolytic product of LC2. The comparison between human and beef cardiac myosin light chains indicated that the sequences of these subunits of the myosin molecule are highly conserved.

Amino Acid Sequence↗

Evidence for new forms of cardiac myosin heavy chains in mechanical heart overloading and in ageing.

Heavy chains of myosin rods and subfragment 1 were isolated from normal hearts and from mechanically overloaded hearts of young and older rats. These myosin heavy-chain fragments were cleaved by cyanogen bromide or partially proteolysed by pronase and by chymotrypsin after denaturation with sodium dodecyl sulfate. The peptides, analyzed by electrophoresis on a one-dimensional polyacrylamide slab gel, varied depending on the origin of the cardiac myosin heavy chains. Some bands present in the peptide patterns of the normal heart of young rats were missing from the pattern of greatly hypertrophied hearts and vice versa. We conclude that mechanical overloading of the heart stimulates the synthesis of cardiac myosin 'isozyme' with a heavy-chain primary structure which is different from that observed in the normal heart of young rat. The patterns from myosin heavy-chain peptides from the hearts of older rats were different from those for peptides from young rat hearts; these results also indicate the presence of a new myosin heavy chain specific to ageing. No difference was detected between the peptide patterns of heavy chains isolated from hypertrophied hearts of young and older rats, and those isolated from normal hearts of older rats.

Aging↗

Absence of phosphates from the myosin heavy chains of striated muscles.

This work aimed to determine whether the heavy chains of myosin from different striated muscle were phosphorylated. Myosin and its heavy chains were prepared from cardiac and skeletal muscles of rats injected in vivo with radioactive phosphates. The results for radioactive phosphate localization indicate the absence of phosphate from pure heavy chains and from any of their purified fragments, whatever the striated muscle used. In addition, phosphates are present in the myosin phosphorylated light chain and in a contaminating protein closely associated to the myosin heavy chain.

Animals↗

The separation of the myosin light chains on agarose beads.

The myosin light chains from rabbit skeletal muscle and bovine and human hearts are separated according to their molecular weights by filtration on agarose beads. The purity of the isolated components and the yield of such columns are compared to the results obtained by other techniques of separation.

Animals↗

[Biochemistry of myocardium taken at autopsy. Preliminary report].

The findings after biochemical analysis of heart muscle taken at autopsy are given in this preliminary communication. Human myosin is made up of two heavy sub-units and two light sub-units: it is similar to cardiac myosin found in other mammals, but is different in certain characteristics, particularly immunological ones. Tropomyosin is made up of two different sub-units. The normal human heart contains 1 mg of collagen and 130 microgram of desoxyribonucleic acid (DNA) per 100 mg of fresh tissue. The degree of cardiac hypertrophy correlates with the increase total DNA within the heart, and with the lowering of myofibrillary Ca2+ ATPase, the concentration in the collagen remaining unchanged providing there is no ischaemic heart disease. These techniques may be used to quantify several factors, such as the degree of sclerosis or the nuclear mass in ill-understood conditions such as the primary cardiomyopathies.

Actins↗

Preparation of specific immune sera against rabbit skeletal, pig and human cardiac heavy meromyosins.

Antibodies were produced in guinea-pigs against heavy-meromyosin (HMM) and tested by immunodiffusion and microcomplement fixation (MCF). A similar response was obtained for HMM extracted either from human and pig left ventricles, or from rabbit white skeletal muscles. For each muscle, the antisera cross-react quantitatively with the native intact myosin and are highly specific. Thus, this procedure permits the preparation of muscle type-specific anti myosin antibodies, by a simpler means than those previously published.

Animals↗

A comparative study of reactive--SH groups of cardiac and skeletal muscle myosins.

Cardiac and skeletal muscle myosins have been treated by N-ethylmaleimide in presence or absence of Mg-ADP. The variations of Ca2+ and K+-ATPase activities and the incorporation of N-[14C]ethylmaleimide into the whole myosin molecule and into its separated subunits (heavy and light chains) have been measured with N-ethylmaleimide treatment for different lengths of time. The results reported here show the following: 1. The Ca2+-ATPase activity of cardiac myosin is activated by N-ethylmaleimide treatment to a lesser extent than that of skeletal myosin. 2. The K+-ATPase activity of both myosins is inhibited in the same quantitative way. 3. The cardiac light chain L1 contains one highly reactive thiol group which is absent from the skeletal light chains. 4. The labelling of three SH-groups localized in the heavy subunits of both myosins induced the same degree of inactivation. 5. The difference observed between the degree of inhibition of the Ca2+-ATPase activity for the two types of myosin with longer treatments appears to be due to differences in the reactivity of the fourth--SH group labelled on the heavy chains.

Adenosine Triphosphatases↗

Human cardiac myosin ATPase and light subunits. A comparative study.

Myosin was extracted from normal human hearts (autopsy material) and compared to that of pig heart and rabbit white skeletal muscle. Myosin light subunits were isolated by a preparative urea gel electrophoresis. These subunits were shown by urea and sodium dodecylsulfate gel electrophoresis to be only slightly affected by the time lapse between death and the beginning of myosin extraction. This was also true for myosin ATPases. The Ca-2+-activated ATPases of pig and human heart myosins have the same apparent Km and V, whereas white skeletal muscle myosin ATPase has the same Km with a higher V. Human myosin light subunits, when compared to those of pig heart possess: (i) different molecular weights: 27 999 and 18 000 datlons for pig heart, and 25 000 and 19 000 daltons for human heart. (ii) for both the light chains, different ultraviolet spectra and a higher helical content for the subunit molecular weight 25 000. (iii) a different composition for several amino acids (Tyr, Pro, Lys). A third light subunit (molecular weight 15 000) was occasionally seen in human as well as pig heart myosin. It concentration varied inversely with that of the subunit molecular weight 27 000-25 000, and so was probably a degradation product of the heaviest subunit.

Adenosine Triphosphatases↗

Cardiac myosin: preparation, ATPase in chronic heart hypertrophy.

A low myofibrillar ATPase seems to be established definitely in several experimental models of chronic heart hypertrophy as well as in humans, but the biochemical pathogenesis of this defect is still unclear. Three different preparations of myosin were studied. Their purity was estimated by measuring MgATPase or by polyacrylamide gel electrophoresis. The first preparation was highly contaminated by actin and tropomyosin; the second was rather pure, and the third (chromatography on DEAE-Sephadex) was pure but slightly denaturated. Heart myosin CaATPase (ionic strength 0.6 or 0.06) was decreased in chronic aortic insufficiency in the rabbits (CAI) when all three preparations were tested. Two (molecular weight 18,000 and 26,000), sometimes three light subunits were found in heart myosin. Their charge and molecular weight are normal in CAI. The third subunit (molecular weight 15,500) was found in control as well as in CAI. Search for an inhibitor was unsuccessful since the two myosin ATPases are additive. The nucleoprotein peak separated from myosin during chromatography was identical in control and CAI. Therefore, myosin seems to be abnormal in CAI.

Adenosine Triphosphatases↗

Heart contractile proteins.

That several proteins of the sarcomere differ from one muscle to the next is well documented, and it is becoming evident that homogeneous muscles, like the heart, are also species specific. 1) Clear-cut evidence is available concerning myosin, and, to date, several types of molecules have been described. a) The myosins of white skeletal, heart, and smooth muscle differ in the activity of their Ca2+ and K+ATPases, as also in the structure of their light subunits. b) The Ca2+ATPases of the various cardiac myosins have been shown to exhibit species differences and correlate with the speed of shortening of the muscle. 2) The structures of tropomyosin, some troponin components, and alpha actinin (but not actin) appear to be unlike in the different types of muscle. 3) These phylogenic modifications may be related to the changes characteristic of the particular muscles under pathological conditions, which are accompanied by substantial increase in protein synthesis.

Actins↗