Muscle Myosin Heavy Chain mRNA Research Articles

NADPH oxidase (NOX) derived superoxide (O2.‐) upregulates intermediate‐conductance Ca2+ ‐ activated K+ channel (KCa3.1) expression in vascular smooth muscle cells (VSMCs)

Reactive oxygen species (ROS), predominantly from NOX, mediate VSMC phenotypic modulation known to play a role in hypertension, atherosclerosis, and hypertrophy. We have previously shown that upregulation of KCa3.1 via increased AP‐1 activity is required for phenotypic modulation of porcine coronary smooth muscle cells (PCSMC). However, the role of ROS in KCa3.1 regulation remains unknown. Our objective was to determine if NOX regulates KCa3.1 expression via AP‐1. Basic fibroblast growth factor (bFGF, 25ng/mL) increased O2.−production in PCSMC, while the NOX inhibitor apocynin (Apo, 1mM) completely abolished O2.− production. bFGF increased KCa3.1 mRNA levels 2.5 fold; while decreasing smooth muscle myosin heavy chain mRNA levels 80% (P< 0.05) in PCSMC. Apo completely inhibited KCa3.1 mRNA upregulation indicating that NOX derived O2.−is required for KCa3.1 upregulation. Acute bFGF significantly increased K+ current, which was inhibited ~50% by the KCa3.1 specific channel blocker TRAM‐34 (100nM) indicating that bFGF causes an upregulation in KCa3.1 channel activity. bFGF increased AP‐1 promoter activity 2.5 fold, while Apo completely inhibited the response to bFGF. These results support a novel mechanism by which NOX derived O2.− regulates KCa3.1 expression through AP‐1.

Nitric oxide synthase inhibition prevents activity‐induced NFAT nuclear accumulation and skeletal muscle adaptation

Involvement of calcineurin/NFAT signaling in the regulation of skeletal muscle myosin heavy chain (MHC)‐based fast‐to‐slower isoform adaptations (F‐S) is recognized. Less is known about the role nitric oxide (NO) plays in this process.OBJECTIVETo investigate the necessity of NO in activity‐induced skeletal muscle F‐S in vivo.METHODSEndogenous NO production was blocked by administering L‐NAME (0.75 mg ml−1; ~100 mg kg−1 day−1) during 0, 1, 2, 5 or 10 days of chronic low‐frequency stimulation (CLFS; 10 Hz, 12 h d−1) applied to the tibialis anterior and extensor digitorum longus muscles of rats (L‐Stim; n=6 each group). Control rats received only CLFS (Stim; n=6 each group).RESULTSCLFS induced increases in NFATc1 nuclear localization in Stim, which did not occur in L‐Stim at any time point. These results were confirmed by western blot analyses. Moreover, MHC mRNA, protein and fibre type analyses revealed CLFS‐induced F‐S occurred in Stim, but were completely abolished in L‐Stim.CONCLUSIONSNO may be regulating activity‐induced MHC‐based F‐S at the transcriptional level via NFAT nuclear accumulation in vivo. *Authors contributed equally. Funded by NSERC, AHFMR, CIHR and CRC.

Characterization of Primary Bladder Neck Obstruction: Immunohistochemical and Gene Expression Analysis

Background: Pathophysiology of primary bladder neck obstruction(PBNO) remains poorly understood. We postulatedabnormal smooth muscle (SM) differentiation and disorganizationof bladder neck musculature as key features of PBNOpathophysiology. Materials and Methods: Immunohistochemicalanalysis and gene expression profiles of bladderneck specimens derived from 11 patients urodynamically diagnosedwith PBNO, 4 patients with benign prostatic hypertrophy(BPH) and 2 normal controls were performed. Consecutivesections were processed for immunohistochemicalcharacterization using anti-SM a-actin, h-caldesmon, collagentype IV, and GAP-43 antibodies. cDNA, synthesized fromisolated RNA of the specimens, were polymerase chain reactionamplified to assess for expression of smooth musclemyosin heavy chain, collagen types I and III, non-musclemyosin heavy chain, and a1A adrenergic receptor mRNA.Results: While SM disorganization in PBNO specimens comparedwith normal bladder neck tissue was observed, PBNObladder neck specimens stained positively for SM markers:a-actin, h-caldesmon and collagen type IV, suggesting theirSM origin. Staining for GAP-43, a phosphoprotein associatedwith the growth of axons and the modulation of new nerve/muscle connection, was also positive in PBNO tissue. Reversetranscription and polymerase chain reaction measurementsof SM related gene expression indicated extremely low levelsof smooth muscle myosin heavy chain mRNA expressionin PBNO specimens when compared to BPH or normal tissue.Expression of a1A adrenergic receptor was maintainedin both PBNO and normal bladder neck specimens. Conclusion:Expression of SM specific markers in the PBNO tissuewas diminished compared to bladder neck tissue from BPHor normal controls, suggesting SM dedifferentiation or transdifferentiation.

Changes in the Expression of Smooth Muscle Myosin Heavy Chain mRNA following Partial Bladder Obstruction or Spinal Cord Injury in Rat: A Preliminary Study

Purpose: The smooth muscle myosin heavy chain (SMMHC) isoform composition has been actively researched in a partial bladder obstruction (PBO) or spinal cord injury (SCI) model. Even though rat is an ideal animal for studying bladder physiology, there were very few reports about the changes of the SMMHC isoforms in the PBO or SCI injured bladder of rat. We developed two polymerrase chain reaction (PCR) primer sets to amplify the isoforms and we applied the primers to the PBO and SCI rat models. Materials and Methods: Female rats had their bladder necks surgically obstructed or they were subjected to spinal cord injury. Six weeks after the event, the bladders were excised. The expression of the C-terminal (SM1 and SM2) and N-terminal (SM-A and SM-B) isoforms of SMMHC was analyzed by performing reverse transcriptase-PCR (RT-PCR). Results: The control bladder showed only the SM-B isoform in the C-terminal. However, the bladder after SCI showed an increased SM-A to SM-B ratio. In case of PBO, the ratios were variable. A decreased SM1 expression was noted in the PBO and SCI groups when compared to the control group (p<0.05). Conclusions: Our female rat models for PBO or SCI demonstrates changes in the expression of smooth muscle myosin heavy chain isoforms. We will apply this primer set for studying of rat muscular physiology in PBO or SCI model. (Korean J Urol 2007;48:522-526)

Myosin heavy chain mRNA expression correlates higher with muscle protein accretion than growth in Atlantic salmon, Salmo salar

In order to link growth and protein accretion in Atlantic salmon the mRNA expression of muscle myosin heavy chain ( MyHC) was analysed. MyHC gene is expressed throughout muscle development and is consistent with the hypertropic growth in fish. Total RNA was isolated from white muscle tissues ( N = 32) from salmon fed a fish meal based diet with three levels of solubilised protein incorporated as fish protein hydrolysate (FPH) control (0 g kg − 1 ), medium (180 g kg − 1 ) and high (300 g kg − 1 ) FPH inclusion. The salmon were PIT-tagged and fed the experimental diets for a period of 68 days. A high level of dietary FPH resulted in significant up-regulation of MyHC by a factor of 2.4 compared to fish fed the medium FPH inclusion. The fish with highest MyHC expression also contained significant higher levels of muscle protein. MyHC expression correlated higher with protein accretion than individual specific growth rate (SGR) in salmon. These observations indicate that MyHC mRNA expression can be a useful marker for understanding of growth and protein accretion in Atlantic salmon.

Changes in detrusor smooth muscle myosin heavy chain mRNA expression following spinal cord injury in the mouse.

Smooth muscle myosin heavy chain (SMMHC) isoform composition has been shown to be developmentally regulated and to be associated with functional changes in smooth muscle activity. In this study, we sought to determine expression patterns of SMMHC isoforms in a murine model of spinal cord injury (SCI) and to compare these expression patterns to neurologic, cytometric, and morphometric findings. Baseline cystometry was performed on adult, female mice followed by either thoracic spinal cord transection (SCI) or sham operation (Sham). At 1, 3, or 6 weeks postoperatively neurologic evaluation and cystometry were performed, bladders were harvested, and expression patterns of SMMHC isoforms (SM1 vs. SM2 and SMA vs. SMB) were assessed by RT-PCR. Morphometrics utilizing computer-assisted color image analysis was also performed on all bladders. There was a significant increase in bladder weight and capacity 1 week following SCI which normalized over time, however, morphometric analysis did not reveal an alteration in tissue composition amongst the three groups. One week following SCI, SM1 was predominantly expressed over SM2 and began to normalize at 3 weeks. This coincided with the emergence of reflex voiding and detrusor overactivity. SMA was expressed following SCI only, and the number of bladders found to express SMA decreased with increasing duration since SCI. Smooth muscle myosin heavy chain mRNA expression patterns appear to be affected by SCI. We believe the induction of SMA may be a factor in altered bladder function following injury.

The effect of androgen status on skeletal muscle myosin heavy chain mRNA and protein levels in rats recovering from immobilization

Immobilization rapidly alters skeletal muscle. The aim of the present study was to determine whether testosterone administration or, in contrast, hypogonadism affects the recovery of muscle mass and myosin heavy chain (MHC) profile at both the mRNA and protein level, after 1 week of immobilization. Male rats were assigned to one of five groups: control (C), hindlimb-immobilized (IMM), and recovery (REC; where animals were allowed 2 weeks of free cage-activity after immobilization). The recovery group was further divided to eugonadal (REC-C), castrated (REC-GDX), and a testosterone-treated (REC-T). In all groups except REC-T, the body masses after immobilization were smaller than in C, although after immobilization the body mass in REC-T recovered at a slower rate than in the other two REC groups. The gastrocnemius mass and the amount of type IIa MHC mRNA decreased during immobilization, but the control levels were regained after recovery. The amount of type IIb mRNA was reduced in REC-GDX compared to C and IMM. The changes in the relative distribution of MHC mRNA were in line with these results. After recovery, the proportion of type IIx MHC protein increased and type IIb protein decreased, although in REC-T the changes were not statistically significant. The proportion of type IIa MHC protein increased only in REC-GDX. In summary, during recovery from immobilization it seems that muscle mass increases and the MHC mRNA and protein profile tend to change toward a slower phenotype, primarily as a result of the decrease in type IIb MHC. However, these changes occur rather independently of the testosterone status.

Passive stretch modulates denervation induced alterations in skeletal muscle myosin heavy chain mRNA levels.

The effect of denervation and denervation combined with immobilisation in either the shortened or lengthened position (passive stretch) upon myosin heavy chain (MyHC) mRNA levels was examined in three rat hind-limb muscles with differing phenotypes. Denervation alone caused a reduction in type I and type IIa MyHC transcripts in all three muscles. In contrast denervation caused a 72% increase in type IIb in the slow postural soleus muscle only which was prevented by immobilisation in the lengthened position. In the same muscle passive stretch also significantly retarded the effects of denervation upon the type I transcript (from 38% below control levels to 24% below) and type IIa transcript (from 59% to 32% below control levels). The levels of both type I and IIa transcripts, in the fast phasic plantaris muscle, were both unaffected by stretch combined with denervation when compared to denervation alone. In the mixed gastrocnemius muscle stretch affected the level of the type I but not the type IIa transcript. These data suggest that passive stretch can modulate MyHC gene expression independently of innervation but that it does so in a muscle-specific manner.

Passive stretch modulates denervation induced alterations in skeletal muscle myosin heavy chain mRNA levels

Passive stretch modulates denervation induced alterations in skeletal muscle myosin heavy chain mRNA levels

Unique expression patterns of myosin heavy chain genes in the ductus arteriosus and uterus of rabbits.

In smooth muscle tissue, two smooth muscle myosin heavy chain (MHC) isoforms (SM1, SM2) and two non-muscle MHC isoforms (NMA, NMB) have been identified. The purpose of our study was to clarify whether smooth muscle MHC mRNA expression reflects the physiological and functional state of the muscle. We studied the expression pattern of MHC mRNAs, using the S1-nuclease mapping procedure, in functionally and morphologically changeable organs; the ductus arteriosus (DA) during development (25 and 29 days of gestation, and from 3-day-old neonates) and uteri from virgin, day-10 pregnant (P10) and day-29 pregnant (P29) rabbits. The results demonstrated that SM2 expression was greater in the fetal DA than in the fetal aortic and pulmonary arteries, but that it decreased significantly following closure of DA. In the gravid uterus, SM1 expression was significantly (P<0.05) strong compared to other MHC mRNAs from virgin to P10 rabbits. During pregnancy, NMB expression showed a tendency to increase until P10, and after P10, SM2 expression increased dramatically and NMB expression decreased to give almost a mirror image of the SM2 expression. Smooth muscle type (SM1, SM2) was significantly (P<0.05) strong compared to non-muscle type expression (NMA, NMB) at P29. These data suggest that smooth muscle MHC mRNA, especially SM2 expression reflects the physiological and functional state of the smooth muscle.

Molecular Cloning and Expression of Murine Smooth Muscle Myosin Heavy Chains

The nucleotide and deduced amino acid sequences of two isoforms of mouse smooth muscle myosin heavy chain (SM1 and SM2) were determined. SM1 (6175 bp) and SM2 (6214 bp) cDNA contained a single open reading frame that encodes 1972 and 1938 amino acids (227,056 Da and 223,294 Da), respectively. Smooth muscle myosin heavy chain mRNA was expressed highly in smooth muscle tissue (small intestine) and weakly in heart and lung. Each of SM1 and SM2 cDNA was transfected and expressed in CHO cells. The expressed myosin heavy chains were detected with an antibody raised against smooth muscle myosin heavy chains and showed the same mobility as the native smooth muscle myosin heavy chains in SDS–PAGE.

Interaction of thyroid hormone and functional overload on skeletal muscle isomyosin expression.

This study examined the interaction of exogenous thyroid hormone 3,5,3'-triiodothyronine (T3) and functional overload on skeletal muscle myosin heavy chain (MHC) expression, studied at both the protein and mRNA level of analysis. Animals were allocated to the following groups: 1) normal control, 2) overload control, 3) hyperthyroid control, and 4) hyperthyroid+overload. Overload of the rat plantaris was accomplished by surgical removal of its synergists (soleus and gastrocnemius), and the animals were made hyperthyroid by injections of T3 (350 micrograms/kg every other day). After overload of 8 wk, muscle enlargement occurred by 53% for both overload groups (P < 0.05). This was accompanied by a 330 and 82% increase in the relative content of type I and IIa MHC, respectively, and a corresponding decrease by 16 and 44% in type IIx and IIb MHC, respectively, in the overload control group (P < 0.05 vs. normal control). Changes in the relative and absolute content of mRNA for these MHCs paralleled the protein response. Exogenous T3 completely reversed the upregulation of type I MHC and the downregulation of type IIx associated with overload at both the protein and mRNA level (P < 0.05). However, T3 was only partially effective in blunting the downregulation of IIb MHC and the upregulation of IIa MHC (protein and mRNA) accompanying the overload.(ABSTRACT TRUNCATED AT 250 WORDS)

Interactions between Growth Hormone and Nutrition in Hypophysectomized Rats: Skeletal Muscle Myosin Heavy Chain mRNA Levels

The aim of this study was to examine the role of growth hormone (GH) in regulating muscle phenotype and to determine how this is modulated by altered nutrition. Total RNA was extracted from gastrocnemius muscles of hypophysectomised rats treated with saline, GH or GH but fed a restricted intake. Type 1, 2A, 2B, embryonic and neonatal myosin heavy chain mRNA levels were estimated by slot blot hybridization. Hypophysectomy reduced the concentrations of types 1, 2A and embryonic mRNAs and dramatically elevated types 2B and neonatal compared to control levels, but this was time-dependant. All MHC mRNA levels were partially restored to control levels in the GH-treated rats except for type 1; the level of this transcript was only elevated by GH in the restricted intake group. Restricted food intake modulated the effects of GH administration for all other MHC mRNA concentrations.

Smooth muscle myosin regulation by serum and cell density in cultured rat lung connective tissue cells

RNA and protein analyses were used to detect expression of SM1 and SM2 smooth muscle myosin heavy chain (MHC) in cultured adult rat lung connective tissue cells (RL-90). Smooth muscle MHC mRNA expression in confluent cells grown in 10% serum was approximately 50% of the level in adult stomach. Similar results were obtained in cells cultured at low density (25% confluency) in 1% serum. However, in low-density cultures transferred to 10% serum for 24 h, the level of MHC mRNA decreased to approximately 20% of that in adult stomach. Smooth muscle alpha-actin showed a pattern of expression similar to that for smooth muscle MHC. Expression of nonmuscle MHC-A mRNA was higher in all culture conditions compared to stomach. MHC-A mRNA expression was less in low-density cultures in low serum and increased when low-density cultures were transferred to 10% serum for 24 h. MHC-B mRNA expression was less in low- vs. high-density cultures. In contrast to MHC-A, however, MHC-B mRNA expression in low-density cultures was higher in low serum. Immunofluorescence and immunoblotting with SM1-specific antibody demonstrated the presence of the SM1 protein isoform as well as reactivity to a protein band migrating slightly faster than SM2. These results demonstrate that cultured rat lung connective tissue cells express smooth muscle MHC and that expression is modulated by culture conditions.

An insert of seven amino acids confers functional differences between smooth muscle myosins from the intestines and vasculature

The molecular mechanisms underlying the heterogeneity in contractile properties observed among smooth muscle tissues are unknown. We examined whether part of this diversity might be intrinsic to myosin by comparing structural and enzymatic properties of myosins from two physiologically diverse tissues. Using the reverse transcriptase polymerase chain reaction, we compared avian intestinal smooth muscle and vascular smooth muscle myosin heavy chain (MHC) mRNA. We found that intestinal, but not vascular, MHC mRNA contains an insert of 21 nucleotides, encoding 7 amino acids, in a region near the ATP binding site in the myosin head. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of purified myosin revealed that the relative mobilities of the previously described intestinal MHC isoforms SM1 (204 kDa) and SM2 (200 kDa) were slower than the corresponding vascular SM1 and SM2 isoforms. Furthermore, antibodies raised against a synthetic peptide corresponding to the deduced amino acid sequence of the intestinal insert strongly recognized intestinal SM1 and SM2 but only weakly recognized the vascular isoforms. The presence of the insert in intestinal myosin correlated with a higher velocity of movement of actin filaments in vitro and a higher actin-activated Mg(2+)-ATPase activity, compared with vascular myosin. Other than the MHC insert, one other structural difference distinguished intestinal and vascular myosins: two isoforms of the 17-kDa myosin light chain were found in vascular myosin, whereas a single isoform was found in intestinal myosin. Exchange of the intestinal myosin light chains onto the vascular MHC did not alter its activity in the in vitro motility assay, suggesting that the 7-amino acid MHC insert is responsible for the different enzymatic activities of vascular and intestinal myosins.

Mitogen stimulation affects contractile protein mRNA abundance and translation in embryonic quail myocytes.

In cultures of differentiated, fusion-blocked muscle cells obtained from embryonic Japanese quail (Coturnix coturnix japonica), mitogen stimulation leads to an immediate reduction in the rates of synthesis of skeletal muscle myosin heavy chain (MHC) and alpha-actin. The molecular mechanisms responsible for this downregulation were examined. The cellular abundances of the alpha-actin and MHC mRNAs were affected differently by mitogen stimulation; alpha-actin mRNA abundance declined by an amount which quantitatively accounted for the observed decrease in alpha-actin synthesis, whereas MHC mRNA abundance remained virtually unchanged during the first 6 h following mitogen stimulation, a period during which MHC synthesis declined by more than 70%. MHC mRNA abundance did decline between 6 and 12 h after mitogen stimulation. Downregulation of MHC synthesis therefore involves an initial block in mRNA translation combined with a later loss of MHC mRNA from the cytoplasma, while alpha-actin synthesis is regulated at the level of mRNA abundance. These observations are consistent with the hypothesis that, in addition to transcriptional activation of muscle-specific genes, skeletal muscle differentiation normally involves cell cycle-dependent modulations in cellular factors which control message stability and message translation.

Mitogen stimulation affects contractile protein mRNA abundance and translation in embryonic quail myocytes.

In cultures of differentiated, fusion-blocked muscle cells obtained from embryonic Japanese quail (Coturnix coturnix japonica), mitogen stimulation leads to an immediate reduction in the rates of synthesis of skeletal muscle myosin heavy chain (MHC) and alpha-actin. The molecular mechanisms responsible for this downregulation were examined. The cellular abundances of the alpha-actin and MHC mRNAs were affected differently by mitogen stimulation; alpha-actin mRNA abundance declined by an amount which quantitatively accounted for the observed decrease in alpha-actin synthesis, whereas MHC mRNA abundance remained virtually unchanged during the first 6 h following mitogen stimulation, a period during which MHC synthesis declined by more than 70%. MHC mRNA abundance did decline between 6 and 12 h after mitogen stimulation. Downregulation of MHC synthesis therefore involves an initial block in mRNA translation combined with a later loss of MHC mRNA from the cytoplasma, while alpha-actin synthesis is regulated at the level of mRNA abundance. These observations are consistent with the hypothesis that, in addition to transcriptional activation of muscle-specific genes, skeletal muscle differentiation normally involves cell cycle-dependent modulations in cellular factors which control message stability and message translation.

Analysis of the 5' end of the Drosophila muscle myosin heavy chain gene. Alternatively spliced transcripts initiate at a single site and intron locations are conserved compared to myosin genes of other organisms.

We have localized the transcription start site of the Drosophila melanogaster muscle myosin heavy chain (MHC) gene and find that all forms of the alternatively spliced MHC mRNA initiate at the same location. Therefore the alternative inclusion/exclusion of the 3' penultimate exon in transcripts from this gene (Bernstein, S.I., Hansen, C.J., Becker, K.D., Wassenberg, D.R., II, Roche, E.S., Donady, J.J., and Emerson, C. P., Jr. (1986) Mol. Cell. Biol. 6, 2511-2519; Rozek, C.E., and Davidson, N. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 2128-2134) does not result from the use of different 5' transcription initiation sites. This gene is the first invertebrate MHC gene shown to have TATA and CAAT box consensus sequences and a noncoding 5' exon, properties that are shared with some vertebrate and invertebrate contractile protein genes. The intron that splits the 5' noncoding region of the Drosophila MHC gene contains no major conserved elements relative to other Drosophila contractile protein genes. The introns within the coding region near the 5' end of the Drosophila MHC gene are located at the same sites as nematode and vertebrate MHC gene introns, indicating that these MHC genes are derived from a common ancestral sequence. The putative ATP binding domain encoded in the fourth exon of the Drosophila MHC gene is highly conserved relative to vertebrate, invertebrate, and non-muscle MHC genes suggesting that each of these myosins bind ATP by the same mechanism. Two divergent copies of the third exon are present within the 5' region of the Drosophila MHC gene, suggesting that alternative splicing produces MHC isoforms with different globular head regions.

Sequential accumulation of mRNAs encoding different myosin heavy chain isoforms during skeletal muscle development in vivo detected with a recombinant plasmid identified as coding for an adult fast myosin heavy chain from mouse skeletal muscle.

In order to study developmental transitions of myosin heavy chain gene expression, we have cloned from newborn mouse skeletal muscle a recombinant plasmid (plasmid MHC 32) that contains an insertion coding for the COOH-terminal portion of an adult fast myosin heavy chain isoform of mouse skeletal muscle. By Northern blots and dot blots, it has been shown that the MHC 32 sequence reveals a broad cross-hybridization with RNA from different mammalian striated muscle tissues. Southern blots with mouse genomic DNA show only one homologous gene, but cross-hybridization at lower stringency to seven to eight different bands, some containing multiple genomic fragments, among which are probably the genes encoding the different striated muscle isoforms. S1 protection experiments with RNA from mouse skeletal muscle before and after birth demonstrate that plasmid MHC 32 is homologous to a major mRNA species of adult skeletal muscle. This adult mRNA is a predominant sequence within 5-6 days after birth. It begins to accumulate at 1-3 days; at the 18th day fetal stage, another major mRNA species is detected as partially homologous with the adult MHC 32 sequence. This fetal myosin heavy chain mRNA is still predominant at 1-3 days after birth, but is rapidly (by 5-6 days) replaced by the adult MHC sequence. There is thus a rapid transition after birth from fetal to adult skeletal muscle myosin heavy chain mRNA sequences.

Cytoplasmic processing of myosin heavy chain messenger RNA: evidence provided by using a recombinant DNA plasmid.

A recombinant DNA plasmid, designated pMHC25, has been constructed that contains structural gene sequences for rat skeletal muscle myosin heavy chain (MHC). The identity of the MHC sequence insert in pMHC25 was determined by muscle-tissue specificity, inhibition of MHC protein synthesis in vitro by hybrid-arrested translation, purification of mRNA that directs the synthesis of MHC protein in vitro, and hybridization to a 33S cytoplasmic mRNA found only in differentiated muscle cells. pMHC25-DNA-excess filter hybridizations were used to show that more than 90% of the newly synthesized MHC mRNA that appears in the cytoplasm of differentiated L6E9 myotubes contains a long 3' poly(A) tail. In contrast, 90% of the MHC mRNA that accumulates in the cytoplasm of these same cells during myogenic differentiation lacks this long 3' poly(A) tail. These results suggest the occurrence of a posttranscriptional event in differentiated L6E9 myotubes that involves the cytoplasmic processing of poly(A)+ MHC mRNA to poly(A)- or poly(A)-short MHC mRNA.