DRESS syndrome associated with nevirapine therapy.
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Adult↗
Biomedical subjects
A F Martin
Publications and source records attributed to A F Martin.
Expression of slow skeletal troponin I in adult transgenic mouse heart muscle reduces the force decline observed during acidic conditions.
1. Acidosis in cardiac muscle is associated with a decrease in developed force. We hypothesized that slow skeletal troponin I (ssTnI), which is expressed in neonatal hearts, is responsible for the observed decreased response to acidic conditions. To test this hypothesis directly, we used adult transgenic (TG) mice that express ssTnI in the heart. Cardiac TnI (cTnI) was completely replaced by ssTnI either with a FLAG epitope introduced into the N-terminus (TG-ssTnI) or without the epitope (TG-ssTnI) in these mice. TG mice that express cTnI were also generated as a control TG line (TG-cTnI). Non-transgenic (NTG) littermates were used as controls. 2. We measured the force-calcium relationship in all four groups at pH 7.0 and pH 6.5 in detergent-extracted fibre bundles prepared from left ventricular papillary muscles. The force-calcium relationship was identical in fibre bundles from NTG and TG-cTnI mouse hearts, therefore NTG mice served as controls for TG-ssTnIand TG-ssTnI mice. Compared to NTG controls, the force generated by fibre bundles from TG mice expressing ssTnI was more sensitive to Ca(2+). The shift in EC(50) (the concentration of Ca(2+) at which half-maximal force is generated) caused by acidic pH was significantly smaller in fibre bundles isolated from TG hearts compared to those from NTG hearts. However, there was no difference in the force-calcium relationship between hearts from the TG-ssTnIand TG-ssTnI groups. 3. We also isolated papillary muscles from the right ventricle of NTG and TG mouse hearts expressing ssTnI and measured isometric force at extracellular pH 7.33 and pH 6.75. At acidic pH, after an initial decline, twitch force recovered to 60 +/- 3 % (n = 7) in NTG papillary muscles, 98 +/- 2 % (n = 5) in muscles from TG-ssTnIand 96 +/- 3 % (n = 7) in muscles from TG-ssTnI hearts. Our results indicate that TnI isoform composition plays a crucial role in the determination of myocardial force sensitivity to acidosis.
Acidosis↗
Phosphorylation of troponin I by protein kinase A accelerates relaxation and crossbridge cycle kinetics in mouse ventricular muscle.
Phosphorylation of cardiac myofibrils by cAMP-dependent protein kinase (PKA) can increase the intrinsic rate of myofibrillar relaxation, which may contribute to the shortening of the cardiac twitch during beta-adrenoceptor stimulation. However, it is not known whether the acceleration of myofibrillar relaxation is due to phosphorylation of troponin I (TnI) or of myosin binding protein-C (MyBP-C). To distinguish between these possibilities, we used transgenic mice that overexpress the nonphosphorylatable, slow skeletal isoform of TnI in the myocardium and do not express the normal, phosphorylatable cardiac TNI: The intrinsic rate of relaxation of myofibrils from wild-type and transgenic mice was measured using flash photolysis of diazo-2 to rapidly decrease the [Ca(2+)] within skinned muscles from the mouse ventricles. Incubation with PKA nearly doubled the intrinsic rate of myofibrillar relaxation in muscles from wild-type mice (relaxation half-time fell from approximately 150 to approximately 90 ms at 22 degrees C) but had no effect on the relaxation rate of muscles from the transgenic mice. In parallel studies with intact muscles, we assessed crossbridge kinetics indirectly by determining f(min) (the frequency for minimum dynamic stiffness) during tetanic contractions. Stimulation of beta-adrenoceptors with isoproterenol increased f(min) from 1.9 to 3.1 Hz in muscles from wild-type mice but had no effect on f(min) in muscles from transgenic mice. We conclude that the acceleration of myofibrillar relaxation rate by PKA is due to phosphorylation of TnI, rather than MyBP-C, and that this may be due, at least in part, to faster crossbridge cycle kinetics.
Adrenergic beta-Agonists↗
Myosin isoforms in female human detrusor.
The aim of this study was to document the relative proportions of two isoforms of myosin heavy chain in detrusor smooth muscle of women with detrusor overactivity and in asymptomatic controls. Women aged 35-65 with documented detrusor overactivity and without a history of neurologic disease, prior incontinence surgery, elevated post-void residual urine volume, or indwelling urinary catheter were eligible for the study. Full-thickness biopsies of extraperitoneal bladder dome were obtained at the time of laparotomy in six patients with documented detrusor overactivity and in a control group of eight continent patients. Biopsies were frozen in liquid nitrogen, crushed with a frozen mortar and pestle at -80 degrees C, and homogenized in buffer, and the extracts were electrophoresed on 6% polyacrylamide sodium dodecyl sulfate gels and stained with Coomassie blue. The gels were de-stained and then the protein bands were scanned with a densitometer. The mean patient age was 48 years (range, 36-59). Seven patients were Caucasian and seven patients were African American. Detrusor smooth muscle contains a mean of 34% (range, 27-43%) SM1 and 66% (range, 57-73%) SM2 isoforms. There was no difference in isoform composition when patients were compared according to urogynecologic diagnosis or according to race. In detrusor biopsies from women, approximately 34% of myosin is of the SM1 isoform and approximately 66% is of the SM2 isoform. This ratio is relatively constant in the two races studied and unchanged in women with detrusor overactivity. Animal models utilizing outlet obstruction of the bladder to provoke detrusor instability and detrusor hypertrophy are known to alter myosin isoform distribution and may not be appropriate models of detrusor instability in human females.
Adult↗
Localization of regions of troponin I important in deactivation of cardiac myofilaments by acidic pH.
Ca2+-activation of cardiac muscle myofilaments is more sensitive to depression by acidic pH than is the case with skeletal myofilaments. We tested the hypothesis that this difference is related to specific regions of the TnI (troponin I) isoforms in these muscles. We exchanged native Tn complex in detergent-extracted fiber bundles from mouse ventricles with Tn containing various combinations of fast (fsTnI) or slow skeletal (ssTnI) complexed with either cardiac TnC (cTnC) or fsTnC, and with cTnC complexed with the following chimeras: (1) fsTnI N-terminal region (fN) plus cTnI inhibitory peptide (cIp) and cTnI C-terminal region (cC); and (2) cTnI N-terminal region (cN)-cIp-fsTnI C-terminal region (fC). We determined the change in half maximal Ca2+(DeltaEC50) for tension activation at pH 7.0 and pH 6.5. Similar DeltaEC50 values were obtained for unextracted controls (5.53+/-0.30 microm), for preparations containing cTnI-cTnC (5.74+/-0.40 microm), and preparations exchanged with cTnI-fsTnC (5.63+/-0.40 microm). However, replacement of cTnI with fsTnI significantly decreased DeltaEC50 to 3.95+/-0.17 microm. Replacement of cTnI with ssTnI also significantly depressed DeltaEC50 to 2.07+/-0.15 microm. Results of studies using the chimeras demonstrated that the C-terminal domains of cTnI and fsTnI are responsible for these differences. This conclusion also fits with data from experiments in which we measured Ca2+-binding to the regulatory site of cTnC in binary complexes containing cTnC with cTnI, fsTnI, or the chimeras. Our results localize a region of TnI important in effects of acidosis on cardiac myofilaments and extend our earlier data indicating that C-terminal regions of cTnI outside the Ip are critical for activation by Ca2+.
Acid-Base Equilibrium↗
Impaired cardiomyocyte relaxation and diastolic function in transgenic mice expressing slow skeletal troponin I in the heart.
1. To assess the specific functions of the cardiac isoform of troponin I (cTnI), we produced transgenic mice that expressed slow skeletal troponin I (ssTnI) specifically in cardiomyocytes. Cardiomyocytes from these mice displayed quantitative replacement of cTnI with transgene-encoded ssTnI. 2. The ssTnI transgenic mice were viable and fertile and did not display increased mortality or detectable cardiovascular histopathology. They exhibited normal ventricular weights and heart rates. 3. Permeabilized transgenic cardiomyocytes demonstrated an increased Ca2+ sensitivity of tension and a lack of contractile responsiveness to cAMP-dependent protein kinase (PKA). Isolated cardiomyocytes from transgenic mice had normal velocities of unloaded shortening but unlike wild-type controls exhibited no enhancement of the velocity of shortening in response to treatment with isoprenaline. Transgenic cardiomyocytes exhibited greater extents of shortening than non-transgenic cardiomyocytes at baseline and after treatment with isoprenaline. 4. The rates of rise of intracellular [Ca2+] and the peak amplitudes of the intracellular [Ca2+] transients were similar in transgenic and wild-type myocytes. However, the half-time of intracellular [Ca2+] decay was significantly greater in the transgenic myocytes. This change in decay of intracellular [Ca2+] was correlated with an increase in the re-lengthening time of the transgenic cells. 5. These changes in cardiomyocyte function in vitro were manifested in vivo as impaired diastolic function both at baseline and after stimulation with isoprenaline. 6. Thus, cTnI has important roles in regulating the Ca2+ sensitivity of cardiac myofibrils and controlling cardiomyocyte relaxation and cardiac diastolic function. cTnI is also required for the normal responsiveness of cardiomyocytes to beta-adrenergic receptor stimulation.
Animals↗
Interactions at the NH2-terminal interface of cardiac troponin I modulate myofilament activation.
Cardiac troponin I (cTnI) is an essential element in activation of myofilaments by Ca2+ binding to cardiac troponin C (cTnC). Yet, its role in transduction of the Ca2+ binding signal to cardiac troponin T (cTnT) and tropomyosin-actin remain poorly understood. We have recently discovered that regions of cTnI C-terminal to a previously defined inhibitory peptide are essential for full inhibitory activity and Ca(2+)-sensitivity of cardiac myofilaments (Rarick et al., 1997). However, apart from its role in structural binding to cTnC, there is little knowledge concerning the role of the N-terminus of cTnI in the activation and regulation of cardiac myofilaments. To address this question, we generated wild-type mouse cardiac TnI (WT-cTnI; 211 residues) and two N-terminal deletion mutants of mouse cTnI, cTnI54-211 (missing 53 residues), and cTnI80-211 (missing 79 residues). The cTnI54-211 mutant retained the ability to bind to cTnT, but lost the ability to bind to cTnC, whereas the cTnI80-211 mutant lost the ability to bind to cTnT, but bound weakly to cTnC. Both mutants bound to F-actin. In the absence of Ca2+, cTnI54-211 was able to inhibit the unregulated MgATPase activity of myofibrils lacking endogenous cTnI-cTnC to the same extent as WT-cTnI, whereas cTnI80-211 had some impairment of its inhibitory capability. Reconstitution with cTnI54-211/cTnC complex did not restore Ca(2+)-activation of myofibrillar MgATPase activity at all, however, the cTnI80-211/cTnC complex restored Ca(2+)-activation to nearly 50% of that obtained with WT-cTnI/cTnC. These data provide the first evidence of a significant function of a cTnT-binding domain on cTnI. They also indicate that the structural cTnC binding site on cTnI is required for Ca(2+)-dependent activation of cardiac myofilaments, and that cTnT binding to the N-terminus of cTnI is a negative regulator of activation.
Actin Cytoskeleton↗
Cardiac myofilament protein function is altered during sepsis.
Male Sprague-Dawley rats (350-500 g) were made septic by intraperitoneal injection of 200 mg/kg cecal material in 5% dextrose in water (D5W; 5 ml/kg). Control rats (n = 11) received D5W. Preparations were studied on days 1 (n = 7), 3 (n = 7), and 7 (n = 8) of sepsis. In isolated hearts, ventricular function was depressed on days 3 and 7 of sepsis. Densitometric analysis of myofilament proteins from septic rats separated by SDS-PAGE showed no differences in relative amounts of actin, troponin, tropomyosin and myosin light chains compared to control. Myofilament function, assessed by measuring ATPase activities, was altered during sepsis. CA(2+)-independent Mg-ATPase activity was elevated on days 1 and 3 of sepsis, returning toward control by day 7. Maximal ATPase activity was unchanged on day 1, but was increased on days 3 and 7 sepsis. Myofibrillar myosin K(EDTA)-, Ca(2+)-, and Mg(2+)-ATPase activities were not altered, nor were there any apparent changes in myosin heavy chain isoform populations. Our data are the first to demonstrate alterations in minimal and maximal ATPase activities and myofilament CA(2+)-sensitivity during chronic peritoneal sepsis. These alterations may contribute to observed changes in ventricular function.
Animals↗
The C terminus of cardiac troponin I is essential for full inhibitory activity and Ca2+ sensitivity of rat myofibrils.
Although the C terminus of troponin I is known to be important in myofilament Ca2+ regulation in skeletal muscle, the regulatory function of this region of cardiac troponin I (cTnI) has not been defined. To address this question, the following recombinant proteins were expressed in Escherichia coli and purified: mouse wild-type cTnI (WT cTnI; 211 residues), cTnI-(1-199) (missing 12 residues), cTnI-(1-188) (missing 23 residues), and cTnI-(1-151) (missing 60 residues). The inhibitory activity of cTnI and the mutants was tested in myofibrils, from which cTnI.cTnC was extracted by exchanging endogenous cardiac troponin with exogenous cTnT causing the Ca2+ sensitivity of the myofibrils to be lost. Addition of increasing amounts of exogenous WT cTnI or cTnI-(1-199) to cTnT-treated myofibrils at pCa 8 caused a concentration-dependent inhibition of the maximum ATPase activity. However, cTnI-(1-188) and cTnI-(1-151) inhibited this activity to about 75% and 50% of that of the WT cTnI, respectively. We also formed a complex of either WT cTnI or each of the mutants with cTnC, reconstituted the complex into the cTnT-treated myofibrils, and measured the Mg2+-ATPase activity as a function of pCa. We found that the cTnI-(1-188).cTnC complex only partially restored Ca2+ sensitivity, whereas the cTnI-(1-151).cTnC complex did not restore any Ca2+ sensitivity. Each cTnI C-terminal deletion mutant was able to bind to cTnC, as shown by urea-polyacrylamide gel-shift analysis and size exclusion chromatography. Each mutant also co-sedimented with actin. Our results indicate that residues 152-199 (C-terminal to the inhibitory region) of cTnI are essential for full inhibitory activity and Ca2+ sensitivity of myofibrillar ATPase activity in the heart.
Amino Acid Sequence↗
C-terminal isoforms of the myosin heavy chain and smooth muscle function.
Two myosin heavy chain isoforms expressed in smooth muscle, SM1 (204 kDa) and SM2 (200 kDa), are derived from alternate splicing that results in different amino acid sequences at their non-helical C-terminal tail regions. These isoforms are developmentally regulated and differentially expressed in various smooth muscle tissues. The functional role of myosin isoforms differing at the C-terminal tail has been investigated both in vitro and in vivo. Removal of the C-terminal tail of SM1 by chymotrypsin activates the ATPase of myosin at low Mg2+ but does not change the maximum activity. Addition of peptides, mimicking C-terminal tail regions specific to the SM1 and SM2 isoforms, to permeabilized taenia coli smooth muscle fibers inhibits maximum shortening velocity (Vm) and decreases Ca2+ sensitivity but has no effect on maximum force. The inhibition of Vm by the SM1-peptide was not reversed on washout, whereas Vm inhibition by the SM2-peptide is reversible. We demonstrated that the SM1 peptide specifically bound to myosin at the subfragment 2-light meromyosin (S2-LMM) junction using crosslinking and immunomicroscopy. Modification at this site could have a direct effect on crossbridge function. The relation between C-terminal myosin isoforms and contractile function in vivo was examined using estrogen administration to ovariectomized rats to increase the relative expression of the SM1 C-terminal isoform in uterine smooth muscle. This increase in SM1 was significantly correlated with an increase in Vm. In contrast, the high ATPase N-terminal isoform was decreased by administration of estrogen to ovariectomized rats. Thus, changes in C-terminal isoform distribution appear to affect contractile function in vivo. We propose a mechanism whereby the interactions between the C-terminal tail of one myosin molecule and the S2-LMM region of another in the thick filament can modulate contractility in an isoform specific manner. Further work is needed to unequivocally identify the function of smooth muscle myosin isoforms. However, our evidence suggests that the C-terminal heavy chain isoforms may be important modulators of smooth muscle contractility.
Amino Acid Sequence↗
Smooth muscle contractility is modulated by myosin tail-S2-LMM hinge region interaction.
The functional significance of two major smooth muscle myosin isoforms, which differ in the nonenzymic COOH-terminal tail region, is not known. We report here that a 13-amino acid peptide, which mimics a region of the tail unique to the SM1 myosin isoform, inhibits contraction velocity in permeabilized smooth muscle. This peptide is shown to bind to the S2-light meromyosin (LMM) hinge region of myosin using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, photoaffinity labeling, and immunoelectron microscopy. Our results suggest that novel intermolecular contacts between the tail and S2-LMM hinge regions of adjacent myosin molecules in the thick filament may modulate contractility and provide a basis for distinct isoform function.
Animals↗
Correlations between myosin heavy chain isoforms and mechanical parameters in rat myometrium.
1. The relations between mechanical parameters and myosin heavy chain isoforms were studied in myometrial smooth muscle from ovariectomized rats (O) and oestrogen-treated, ovariectomized rats (E). 2. Treatment of the rats for three days with beta-oestradiol (2 micrograms kg-1 day-1) 2-4 weeks postsurgery, produced maximal changes in uterine mass and myosin content of approximately threefold. 3. Myosin heavy chain isoform SM1 (204 kDa) was increased from 65.5 +/- 0.8% to 72.9 +/- 0.6% of the total isoform species (P < 0.001, n = 24, O and E respectively) after oestrogen treatment. 4. To avoid complications associated with activation processes, mechanical parameters were measured in permeabilized myometrial fibre bundles activated at a calcium concentration of 12.6 microM. After oestrogen treatment the maximum velocity of shortening (Vmax) measured by the slack test increased from 0.044 +/- 0.006 of the reference length (Lo) s-1 to 0.101 +/- 0.006 Lo s-1, and maximal isometric force (Pmax) increased from 23.3 +/- 4.4 mN mm-2 to 74.1 +/- 13.9 mN mm-2 (P < 0.001, n = 24, respectively). Series elasticity and the half-time to peak force were not significantly altered. 5. Both Vmax and Pmax correlated significantly with percentage SM1 in O and E fibre bundles (r = 0.61 and 0.56, n = 48 fibres; or r = 0.87 and 0.89, n = 8 grouped data per rat). Vmax, however, was only weakly correlated with Pmax (r = 0.39, n = 48). 6. To assess the relative significance of the correlation between Vmax and the percentage of SM1 and that between Vmax and Pmax, we used a multiple regression analysis with the model Vmax = intercept + beta 1 x % SM1 + beta 2 x Pmax, where intercept, beta 1 and beta 2 are regression parameters. This analysis (n = 48) indicated that Vmax was significantly dependent on the percentage of SM1 (P < 0.0002) but not on Pmax (P < 0.61). 7. There were no significant differences in the levels of myosin light chain phosphorylation between O and E fibre bundles, indicating that light chain phosphorylation is unlikely to be the basis for the differences in mechanical parameters demonstrated by these fibres.
Animals↗
Identification of a novel smooth muscle myosin heavy chain cDNA: isoform diversity in the S1 head region.
Smooth muscle myosin heavy chain (SMHC) isoforms, SM1 and SM2, are the products of alternative splicing from a single gene (P. Babij and M. Periasamy. J. Mol. Biol. 210: 673-679, 1989). We have previously shown that this splicing occurs at the 3'-end of the mRNA, resulting in proteins that differ at the carboxyterminal (R. Nagai, M. Kuro-o, P. Babij, and M. Periasamy. J. Biol. Chem. 264: 9734-9737, 1989). In the present study we demonstrate that additional SMHC isoform diversity occurs in the globular head region by isolating and characterizing two distinct rat SMHC cDNA (SMHC-11 = SM1B and SMHC-5 = SM1A). Sequence comparison of the two clones reveals that they are completely identical in their coding regions, except at the region encoding the 25/50 kDa junction of the myosin head, where the SM1B isoform contains an additional seven amino acids. This divergent region is located adjacent to the Mg(2+)-ATPase site, and differences in this region may be of functional importance. Ribonuclease protection analysis demonstrates that the corresponding SM1B and SM1A mRNA messages are coexpressed in all smooth muscle tissues; however, the proportion of the two mRNA present differs significantly between tissues. The SM1A-type mRNA predominates in most smooth muscle tissues, with the exception of intestine and urinary bladder, which contain greater proportions of the SM1B message. The differential distribution of these two isoforms may provide important clues toward understanding differences in smooth muscle contractile properties.
Alternative Splicing↗
Cleavage of a smooth muscle myosin heavy chain near its C terminus by alpha-chymotrypsin. Effect on the properties of myosin.
Limited proteolysis of gizzard myosin by alpha-chymotrypsin converted the heavy chain doublet pattern, seen by gel electrophoresis, to a single band. Light chain degradation was not observed and only minor cleavage occurred at other heavy chain sites. Using a polyclonal antibody raised against a unique sequence from the slower-migrating heavy chain (SM1) it was shown that this conversion was due to the loss of a peptide approximately 4000 daltons from the C terminus of SM1. The peptide was isolated and sequenced, and the cleavage site was identified between phenylalanine 1943 and alanine 1944. Addition of antibody before protease protected SM1 from cleavage. The following changes were observed (a) the Mg2(+)-dependence of actin-activated ATPase of digested phosphorylated myosin was altered and activity was relatively high at low Mg2+ levels, i.e. similar to phosphorylated heavy meromyosin; (b) the KCl dependence of Mg2(+)-ATPase of the digested myosin, particularly the phosphorylated form, showed an altered pattern consistent with the stabilization of the 6 S conformation; (c) the tendency for aggregation was increased by proteolysis of phosphorylated myosin. These results show that the C-terminal region of a gizzard myosin heavy chain can modify some of the properties of myosin. It is suggested that the observed modifications reflect an enhanced tendency of the digested myosin to aggregate.
Actin Cytoskeleton↗
Myosin heavy chain isoforms and smooth muscle function.
Using isoform specific antibodies we have verified the presence of two distinct muscle type myosin heavy chain isoforms in rat uterine muscle. We have shown that an endogenous protease can cleave a small 4 kDa region from the C-terminal of the SM1 isoform which generates a pSM1 species which comigrates with the SM2 isoform on low density SDS gels. While this cleavage can complicate isoform identification, more importantly, this cleavage was associated with a substantial increase in the actomyosin ATPase. Thus we have identified a domain at the C-terminal which may be involved in regulation of the ATPase activity. Interestingly, it is at this C-terminal, tail region of the smooth muscle myosin molecule where the only known isoform specific sequence differences are located. In skinned smooth muscle fibers of rat uterine muscle, we have also shown that differences in myosin heavy chain distribution, induced by beta-estradiol treatment of ovariectomized rats, are correlated with changes in unloaded shortening velocity. Thus our work suggests that the functional significance of myosin heavy chain isoforms in smooth muscle may be similar to that observed in striated muscle.
Animals↗
Molecular cloning and developmental expression of the rat cardiac-specific isoform of troponin I.
Troponin I is the subunit of the troponin complex in striated muscle which inhibits actomyosin ATPase activity. We have isolated a full-length cDNA clone for rat cardiac troponin I and determined its nucleic acid sequence. The amino acid sequence deduced from this clone shows 88%-92% similarity with previously reported amino acid sequences for rabbit (Wilkinson and Grand, 1978) and bovine (Leszyk et al.) cardiac troponin I. Examination of cardiac troponin I mRNA abundance during development revealed a 15-fold induction in its expression in the adult heart compared to that in embryonic (14 day) heart muscle. Furthermore, expression of cardiac troponin I mRNA was restricted to heart muscle and was not detected in skeletal muscle at any developmental stage.
Amino Acid Sequence↗
Identification and functional significance of troponin I isoforms in neonatal rat heart myofibrils.
We investigated the mechanism(s) responsible for differences in the effects of acidic pH on Ca2+ activation of the activity of adult and neonatal rat heart myofilaments. Studies on preparations of myofilaments reconstituted with adult troponin-tropomyosin (Tn-Tm) and either adult or neonatal thick filaments indicated that the difference in effect of acidic pH is related to differences in Tn-Tm and not other myofilament proteins. Immunoblotting analysis showed that development of the rat heart myofibrils is associated with isoform switching from slow skeletal TnI to cardiac TnI and from a slow mobility isoform of TnT (TnT1) to a faster Mr isoform (TnT2. Expression of slow skeletal TnI was associated with a relative insensitivity of myofilament Ca2+ activation to deactivation by acidic pH. Moreover, the effect of acidic pH on Ca2+ activation of ATPase activity of soleus myofibrils, which contain cardiac TnC and slow skeletal TnI, was essentially the same as the effect of acidic pH on rat cardiac myofibrils in the early neonatal period. Neonatal myofilaments also contained a relative abundance of a set of polypeptides copurifying with the thin filaments. We have identified these proteins as histones. The relative amount of histones among a variety of preparations from different species was not correlated with the pH sensitivity of myofibrillar Ca2+ activation. Shifts in TnT isoforms among these species were also not correlated with an altered response to acidic pH. Our data provide evidence in support of the hypothesis that the relative insensitivity of neonatal myofilament activity to acidic pH is due to the presence of slow skeletal TnI in the thin-filament regulatory complex.
Acids↗
Myocardial Na, K-ATPase in one-kidney, one-clip hypertensive rats.
Myocardial ventricular Na, K-ATPase activity of normotensive rats was compared with that of healthy rats with chronic benign one-kidney, one-clip hypertension. The yield of protein (mg/g wet wt left plus right ventricles) in microsomal and sarcolemmal membrane fractions was the same for both normotensive and hypertensive rat ventricles. However, the yield of protein (mg/ventricle) was 26% greater in the hypertensive relative to the normotensive animals, consistent with the presence of hypertrophy, as also indicated by an increase in the ratio of ventricular to body weight and a shift in the isomyosin composition. Na, K-ATPase activity, sodium-dependent phosphorylation and ouabain binding were significantly (P less than 0.05) decreased (by 20%, 40%, and 45%, respectively) in the hypertensive rat ventricles when the data were expressed in units/g tissue wet weight. However, when expressed in units per ventricle, values in normotensive and hypertensive animals were similar. The molecular activity or turnover number of ventricular (and also renal) Na, K-ATPase activity was the same in both groups of animals. These results suggest that the decrease in myocardial specific Na, K-ATPase activity in the rat made hypertensive by removing one kidney and constricting the renal artery of the other kidney is related to the presence of cardiac hypertrophy.
Animals↗