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-actin null mouse
* Department of Molecular and Cellular Biology,
Department of Medicine, Section of Cardiovascular Sciences,
Department of Molecular Physiology and Biophysics and
§ Howard Hughes Medical Institute, Department of Genetics, Baylor College of Medicine, Houston, Texas 77030, USA
1Correspondence: Department of Cell Biology, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030, USA. E-mail: schwartz{at}bcm.tmc.edu
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-actin gene activated during the early stages
of embryonic cardiovascular development is switched off in late stage
heart tissue and replaced by cardiac and skeletal -actins. SM
-actin also appears during vascular development, but becomes the
single most abundant protein in adult vascular smooth muscle cells.
Tissue-specific expression of SM -actin is thought to be required
for the principal force-generating capacity of the vascular smooth
muscle cell. We wanted to determine whether SM -actin gene
expression actually relates to an actin isoform’s function. Analysis
of SM -actin null mice indicated that SM -actin is not required
for the formation of the cardiovascular system. Also, SM -actin null
mice appeared to have no difficulty feeding or reproducing. Survival in
the absence of SM -actin may result from other actin isoforms
partially substituting for this isoform. In fact, skeletal -actin
gene, an actin isoform not usually expressed in vascular smooth muscle,
was activated in the aortas of these SM -actin null mice. However,
even with a modest increase in skeletal -actin activity, highly
compromised vascular contractility, tone, and blood flow were detected
in SM -actin-defective mice. This study supports the concept that SM
-actin has a central role in regulating vascular contractility and
blood pressure homeostasis, but is not required for the formation
of the cardiovascular system.—Schildmeyer, L. A., Braun, R.,
Taffet, G., DeBiasi, M., Burns, A. E., Bradley, A., and Schwartz,
R. J. Impaired vascular contractility and blood pressure
homeostasis in the smooth muscle -actin null mouse.
Key Words: SM -actin gene • homologous recombination • vascular tone
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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2
3
4
5)
. The high degree of
sequence conservation between the actin proteins suggests that this
multigene family arose by duplication and subsequent divergence from a
common ancestral gene (1
, 6)
. In the course of these
events, certain regulatory and structural features of the actin loci
diversified to produce the specialized actin genes observed today. In
birds and mammals, actins are represented by at least six different
isoforms, including skeletal and cardiac -actins, vascular and
visceral and 
smooth muscle actins, and the ubiquitous ß- and
-cytoplasmic actins (2
3
4
5)
. These actins are highly
homologous, differing from each other by less than 5% of their amino
acid sequence and encoded by separate unlinked genes (2
, 3)
. Tissue-specific expression of actin isoforms, regulated
expression of isoforms during muscle differentiation, and coexpression
of multiple isoforms in various cell types suggest that actin isoforms
may have specialized functions (5
6
7
8)
. For example, the
cytoplasmic actins, including ß and , are enriched in nonmuscle
cells where they participate in the formation of the cytoskeletal
apparatus and microfilaments that function in cell motility,
endocytosis, and related processes (1)
. In contrast, the
striated cardiac and skeletal -actins and smooth muscle - and
-actins compose contractile sarcomeres and smooth muscle myofibrils
of warm-blooded vertebrate muscle tissues (6
7
8
9
10)
.
The smooth muscle (SM) -actin gene is activated during the early
stages of embryonic heart development (11
12
13
14
15)
The
sequential activation of SM -actin and cardiac -actin gene
activity follows the formation of the definitive heart tube. Since the
earliest myocardial filament bundles are reminiscent of the
organization in smooth muscle, it was suggested that SM -actin might
be important in supporting the early contractile functions of the
developing heart when myofibrillogenesis is just beginning
(11)
. SM -actin is down-regulated during late stage
cardiac morphogenesis, being replaced by cardiac and skeletal
-actins (11
12
13)
. In contrast, SM -actin also
appears early during vascular development, but is retained to become
the single most abundant protein in adult vascular smooth muscle cells.
Skeletal and cardiac -actin transcripts are not detected during
normal vasculogenesis (11)
, and SM -actin is thought to
be required for the principal force-generating capacity of the vascular
smooth muscle cell (8
, 9)
Recent studies support the contention that individual actin isoforms
are involved with specific cellular contractile functions
(16
17
18
19
20)
. Lloyd et al. (18)
showed that the
level of the nonmuscle ß- and -actin protein may determine a
feedback-regulatory response. Also, the up-regulation of skeletal
muscle -actin gene and SM -actin gene activity in hearts of
cardiac -actin null mice supports a cellular sensing mechanism that
might respond to changes in sarcomeric contractility and/or the absence
of cardiac -actin by eliciting compensatory gene activity
(19)
. We wanted to determine whether SM -actin gene
expression relates to an actin isoform’s specialized function or is
simply the manifestation of programmatic gene regulation of the actin
multigene family. This study defines the role of SM -actin in the
cardiovascular system by using gene targeting in mouse embryonic stem
cells to inactivate SM -actin gene transcription.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-actin gene targeting-actin genomic DNA clones were isolated from a 129
SVJ mouse genomic library (Stratagene, San Diego, Calif.). The
targeting vector was constructed by inserting a Pol2NeobpA cassette
(21)
at the +1 site of the SM -actin gene and consisted
of an upstream 2.1 kb EcoRV/SmaI fragment and a
downstream 3.5 kb SmaI/XbaI fragment of the SM
-actin gene (22)
. The targeting plasmid vector was cut
at a unique site before electroporation into AB2.2 ES cells. DNA
isolated from G418-resistant clone was digested with XbaI
and screened using the mini Southern procedure (23)
.
Digestion of wild-type (wt) DNA gave a fragment of 
10 kb, whereas
digestion of DNA from targeted clones gave the expected 10 kb wt
fragment, as well as a 5.5 kb fragment from the targeted allele. A
chimeric male mouse capable of germline transmission was obtained by
injecting targeted cells into 3.5-day-old blastocysts from C57 mice.
The male chimera was mated to produce offspring carrying the mutated
allele.
RNA Northern blot analysis
Mice were killed by CO2 asphyxiation.
Tissues of interest were isolated, rinsed in phosphate buffered
standard saline (0.9% NaCl, 050 M NaPO4, pH
7.2), blotted, frozen in liquid N2, and crushed
using a percussion mortar cooled with liquid N2.
Tissues used for RNA isolation were sections of the aorta that extended
from the aortic arch to the diaphragm, the ventricles of the heart, the
lower part of the large intestine, skeletal muscle from the upper
portion of the hind limb, and the uterus. Total RNA was extracted from
the powdered tissues using methods previously described
(24
25
26)
. RNA samples were electrophoresed on 1.0%
agarose, 2.2% formaldehyde gels, blotted onto Genescreen nylon
membrane (DuPont, NEN, Wilmington, Del.), and UV cross-linked to the
membrane. A probe corresponding to the 3'UTR of the mouse SM -actin
gene (22
; 160 bp fragment) was labeled by random
priming. Probes for the cardiac -actin gene (27
; 140
bp fragment) and the skeletal -actin gene (27
; 200
bp fragment) were labeled as riboprobes. Blots were hybridized 16–18 h
at either 45°C for the random primed probes or 68°C for the
riboprobes in a solution containing 50% formamide, 5x SSPE, 5x
Denhardt’s solution, 0.1% bovine serum albumin, 0.1%
polyvinylpyrrolidone, 0.1% ficoll, 1% sodium dodecyl sulfate (SDS),
10% dextran, and 200 mg/ml sheared salmon sperm. Blots were exposed
with intensifying screens to X-ray film (Kodak X-Omat AR) at -70°C
for 1 to 7 days.
Protein Western blot analysis
As described above, powdered tissues were extracted (10 ml of
extraction solution per 1 g of powdered tissue) in a solution of
25 mM Na2HPO4, 1%
ß-mercaptoethanol, and protease inhibitors at 4°C for 1–2 h. SDS
was added to a final concentration of 1%. The samples were incubated
at 37°C for 20 min and clarified by centrifugation at 14,000
g for 5 min. Total protein concentration was determined by
Bradford assay. Protein samples (10 µg per well) were electrophoresed
on 10% acrylamide-1% SDS gels with a 2.5% acrylamide-1% stacking
gel and then transferred to Immobilon-P transfer membrane (Millipore,
Bedford, Mass.). Blots were probed with mAb 1A412 (28
;
Sigma, St. Louis, Mo.) specific for SM -actin and mAb 5C5
(29
; Sigma), specific for cardiac and skeletal -actins,
and reacted with a horseradish peroxidase secondary antibody detected
by chemiluminescence with the ECL Western Blotting Analysis System
(Amersham, Arlington Heights, Ill.) and exposed to X-ray film (Kodak
X-Omat AR).
Immunohistochemistry
Tissues were fixed in zinc formalin, paraffin embedded, and
sectioned at 5 µm. Tissue sections were then stained with a
combination of trichrome and elastica stains to detect muscle,
connective, epithelial, and elastic tissues. Tissue sections were also
stained using the SM -actin monoclonal antibody 1A4
(28
; dilution 1:1000) visualized by DAB.
Transmission electron microscopy
Aortas were fixed for 2 h in phosphate-buffered saline
(PBS) containing 2.5% glutaraldehyde, transferred to PBS containing
1% tannic acid (5 min), and then postfixed in PBS containing 1%
osmium tetroxide (60 min). This was followed by en bloc
staining with aqueous uranyl acetate (60 min), dehydration through
ethanol, and embedding in LX 112 resin (Polysciences, Inc., Warrington,
Pa.). Ultrathin sections were stained with uranyl acetate and lead
citrate prior to viewing on a JEOL 200Cx electron microscope.
Vascular contractility
Aortas were carefully excised from killed mice, immediately
placed in ice-cold Krebs-Ringer solution, and dissected free of
surrounding tissue. A section of the thoracic/abdominal aorta
(extending from a region just below the aortic arch to the diaphragm)
was cut into 3 approximately equal segments of 2–3 mm each. The aortic
ring sections were mounted in 13 ml tissue baths containing a
Krebs-Ringer bicarbonate solution (110 mM NaCl, 4.7 mM KCl, 1.0 mM
MgSO4, 1.2 mM
KH2PO4, 25 mM
NaHCO3, 11.1 mM glucose, and 2.5 mM
CaCl2) at 37°C and bubbled with a mixture of
95% O2 and 5% CO2.
Isometric tension recordings were made using a Brush Mark 200 chart
recorder. Aortic ring sections were equilibrated under tension for at
least 1 h before experiments were begun. Ability to contract was
evaluated at different initial tensions (0.75 g, 1.0 g, and
1.25 g) using two concentrations of KCl (80 mM and 120 mM) to
initiate aortic contraction.
Blood pressure measurement
Systolic blood pressure measurements were made in conscious nice
by a noninvasive tail cuff method (30
; IITC Inc. Life
Science, Woodland, Calif.). Three weeks prior to taking pressure
measurements, mice were trained by gentle handling and warming to
30°C. Baseline pressure measurements were obtained by taking five
consecutive measurements each day for 7 to 10 days for each mouse.
Blood pressure measurements were also taken after infusion of sodium
nitroprusside (0.5 mg/ml) for 15 min via the tail artery. Blood
pressure measurements were continuously monitored over an hour after
withdrawal of drug.
Tail artery blood flow
Mice were anesthetized with an intraperitoneal injection of a
combination anesthetic (Ketamine, Xylazine, Acepromazine). A
pulsed-Doppler (20 MHz) flow probe placed around the tail at least 1.5
cm from the base of the tail recorded blood flow through the caudal
artery as used in the rat (31)
. A temperature probe
inserted into the rectum measured core body temperature. A 60 W
incandescent light was used to warm the animal. The outputs of the flow
probe were directed into a spectrum analyzer, which performed fast
Fourier transforms on the data. Anesthetized mice were first warmed to
a core basal body temperature of 38°C and the Doppler flow probe was
positioned to obtain maximum flow signals through the caudal artery.
Mice were then cooled to <34°C and allowed to remain 15 min at
this temperature. The mice were warmed again; at approximately every
increase of 0.5° in basal body temperature, the blood flow through
the tail artery was recorded for 1 s, providing a relative measure
of blood flow.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-actin null mutant mice-actin in the
cardiovascular system by using gene targeting in mouse embryonic stem
cells to inactivate SM -actin gene transcription. To block the
accumulation of SM -actin transcripts, a Pol II promoter neomycin
hybrid gene was inserted into the SM -actin gene locus at the +1 cap
site. Electroporation of the targeting vector (Fig. 1a
, b
) into AB2.2 cells resulted in a targeting frequency 1
out of 12 positive homologous recombinants per G418-resistant clones. A
male chimeric mouse obtained from a targeted 129 strain was mated with
C57 females, resulting in seven heterozygous breeding pairs.
Intercrosses between these SM -actin heterozygous mice yielded a
number of offspring that approximated the predicted Mendelian ratio of
1:2: 1 for wt, ±, and -/- mice and about an equal number of males
and females, as shown in Table 1
. Mice identified as null mutants (SM -actin -/-) for the SM
-actin gene by Southern blot analysis presented no obvious phenotype
on gross examination and were fertile, reaching adulthood without
succumbing to abnormalities resulting from the absence of this protein.
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Skeletal -actin up-regulated in SM -actin-defective vessels
The expression of muscle -actin genes in SM -actin
null mice was evaluated by RNA Northern blots using 3' untranslated
region (3' UTR) labeled probes specific for each actin isoform. As
shown in Fig. 1c
, robust SM -actin expression is
demonstrated by the appearance of 1.7 kb RNA species in wild-type
aorta, intestine, and uterus tissues and expressed to a lesser extent
in wt cardiac and skeletal tissue. In comparison, the accumulation of
SM -actin RNA transcripts are blocked by neo+ insertion mutagenesis
in tissues from the SM -actin null mice. Expression of the cardiac
-actin gene was not influenced by the SM -actin-defective gene,
as shown by its comparable mRNA levels in wt and mutant striated muscle
tissues (Fig. 1c
). Although cardiac and skeletal -actin
genes are silent in normal vascular smooth muscle tissues, skeletal
-actin RNA unexpectedly was up-regulated in the majority of SM
-actin -/- aortas examined (Fig. 1c
).
SM -actin was immunodetected by Western blots of
electrophoresed protein extracts of wt aorta, uterus, and intestine
reacted with a well-characterized SM -actin mAb (Fig. 1d
). Reduced amounts of SM -actin were detected
in any wt cardiac and skeletal muscle protein extracts. Under identical
assay conditions, SM -actin immunoreactive species were not detected
in extracts of muscle tissues taken from SM -actin null mutants,
thus confirming the absence of SM -actin gene expression in SM
-actin null mice. Striated -actins from cardiac and skeletal
muscle tissue samples of both wt and mutant mice reacted strongly with
the sarcomeric -actin mAb (Fig. 1d
), but wt aortic
extracts did not demonstrate the presence of striated -actin gene
expression in wt smooth muscle tissues. However, aortic samples from SM
-actin null mice (Fig. 1d
) exhibit sarcomeric -actin
reactive protein, substantiating the novel observation of up-regulation
of skeletal -actin gene activity seen in the Northern blots.
Absence of vascular contractility and lowered basal blood pressure
in SM -actin null mice
SM -actin mAb cross-reactivity marked the medial layers of
normal aorta, carotid artery, and cardiac arteriole (Fig. 2a
, c
, e
) but not -/- vessels (Fig. 2b
, d
, f
).
Aortic tissue sections stained with a combination of trichrome and
elastica (Figs. 2g
, h
) showed the presence of
muscle (red) and elastic (black) components in both wt and SM -actin
-/- tissue. The elastic layers of the wt aorta have a contracted,
ruffled-like appearance (Fig. 2g
) whereas the comparably
stained mutant aorta appears unruffled and flat (Fig. 2h
),
as also seen in the antibody-stained sections (Fig. 2a
, b
, c
, d
, e
, f
). At higher resolution, electron microscopy
revealed disorganization and reduction of myofilamentous arrays in the
SM -actin -/- aortic tissue sections (Fig. 2j
, see
arrows) in comparison to wt tissue sections (Fig. 2i
). The
appearance of the -/- vessels led us to examine the likelihood of
impaired biological function.
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Vascular contractility was used to evaluate the functional consequences
of the SM -actin null mutant in vitro. Aortic rings were
set at resting tensions of 0.75 g, 1.0 g, and 1.25 g and
depolarized by addition of KCl (80 mM and 120 mM). Differences in the
responses of the wt and mutant vessels were quite dramatic. Aortic
rings from wt mice (Fig. 3a
, top three tracings) responded to KCl stimulation whereas
aortic segments from mutant mice (Fig. 3a
, lower three
tracings) showed little or no response. Similar results were obtained
by stimulation with norepinephrine (1x10-5 and
1x10-6 M) and serotonin
(1x10-6 M) (data not shown). Differences in
contractility between wt and -/- aortas were independent of the
agonist and the initial tension at the time of stimulation (Fig. 3b
), suggesting that this phenomenon results from a
fundamental dysfunction of the contractile system, presumably the
absence of SM -actin.
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In vivo studies included measuring the systolic blood
pressure in adult conscious mice to determine the relevance of these
observations in vivo. When the values for each pair of mice
were compared, the mean systolic blood pressure of the mutant mouse was
significantly lower that that of his wt littermate in all five pairs
examined (P<0.05, Table 2
). Overall, the SM -actin null mice showed a significant reduction
(28%) in systolic blood pressure when compared to wild-type mice. We
examined the effect of intravenous infusion of sodium nitroprusside
(SNP) on blood pressure. Typically, SNP produces rapid vasodilatation
followed by a baroreflex-mediated activation of the sympathetic nervous
system, releasing norepinephrine and resulting in increased heart rate
and vasoconstriction. In the wt mice, SNP infusion resulted in a 50%
increase in heart rate (data not shown) and a 45% decrease in
systolic blood pressure (Fig. 4a
). When SNP infusion was stopped, the systolic blood
pressure rapidly returned to predrug levels after a transient
overshoot, reflecting the release of NE at the vascular level. In the
two SM -actin null littermates, SNP induced reduced vasodilatation,
a 26% decrease in systolic blood pressure, and reflex tachycardia.
However, when SNP infusion was stopped, the systolic blood pressure
remained low for over an hour (Fig. 4b
). Thus, resistance
blood vessels were unable to respond in response to NE in the absence
of SM -actin.
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Additional in vivo studies investigated the ability of
contractile vessels to relax and undergo vasodilatation. Vascular
dysfunction in the mutant mice was indicated by a diminished peripheral
blood flow in response to thermal challenge. As shown in Fig. 5
, the relative peak tail artery blood flow velocity is increased in
normal mice in response to increasing core body temperature whereas
mutant mice showed a significantly reduced response. Raising the body
temperature of the mice 2.5°C resulted in a fivefold increase in flow
velocity in normal mice but less than a twofold increase in the mutant
mice, demonstrating impaired vascular regulation and an inability to
modify tone in the SM -actin null mice. Increased heart rate in
response to increase in temperature was similar in both the wt and
mutant mice and does not explain this difference in blood flow. The
null mice recovered without incident, without any detection of
vasospasm, on return to normal body temperature.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-actin gene expression relates to a
specialized function during formation or maturation of the
cardiovascular system. SM -actin is the first muscle actin expressed
during development of the cardiovascular system (11)
, but
we showed that SM -actin does not seem to play an obligatory role in
the formation of the cardiovascular system. Mutant SM -actin mice
appear to have no difficulty feeding or reproducing (Table 1
). Survival in the absence of SM -actin may result from other
actin isoforms partially substituting for this isoform
(19)
. Since SM -actin is the predominant actin isoform
in tissues from the gastrointestinal and urogenital tracts (7
, 13)
, it is not surprising that loss of SM -actin in these
tissues appears to have no effect on a gross level.
The first phenotypic difference discerned between wild-type and SM
-actin null mice (Fig. 2)
was clearly revealed upon histological
examination of muscular arteries and arterioles. Cross-sectioned wt
vessels showed a highly contracted, ruffled and or puckered like
appearance whereas comparably stained mutant aorta appeared smooth.
Electron micrographs also showed disarrayed and reduced myofibrils in
SM -actin null -/- vessels in comparison to wt tissue sections.
Large differences in vascular contractility was observed in aortic ring
assays in which aortic segments from mutant mice (Fig. 3)
showed
essentially no response to KCl depolarization, revealing a fundamental
deficit in the vascular smooth muscle contractile system of the SM
-actin null mouse. Blood pressure is thought to be associated with
the contractile state or mass of vascular smooth muscle by the arterial
side of vasculature in which luminal narrowing serves to increase the
total peripheral resistance (31)
. We observed that the
mean systolic blood pressure in adult conscious mice SM -actin null
mice was significantly reduced by 28% when compared to wt littermates
in all five pairs examined (P<0.05, Table 2
). As shown in
Fig. 4
, infusion of sodium nitroprusside in SM -actin null
littermates had impaired vasodilatation and vasoconstriction
activities when infused with sodium nitroprusside, as demonstrated by
the inability to expeditiously reduce and/or raise blood pressure.
Additional in vivo studies, shown in Fig. 5
, also indicated
a diminished peripheral blood flow in response to thermal challenge.
Therefore, mice lacking the SM -actin demonstrate compromised
vascular responses as well as reduced basal blood pressure, which may
make them more susceptible to cardiovascular stress.
This study and others (18
, 19
, 32)
indicate that smooth
and striated muscular tissues have the capacity to sense and respond to
dramatic changes in either the level of contractile proteins and or
contractile activity. Evaluation of the expression of muscle -actin
genes in SM -actin null mice showed that cardiac -actin gene
activity was not influenced by the SM -actin-defective gene, as
shown by its identical mRNA levels in wt and mutant striated muscle
tissues. Whereas cardiac and skeletal -actin genes are silent in
normal vascular smooth muscle tissues, skeletal -actin RNA was
up-regulated in the majority of SM -actin -/- aortas examined.
Apparently, up-regulation of skeletal -actin gene expression in the
aorta of the mutant mice compensates to some degree for the loss of the
SM -actin. In addition, cardiac -actin knockout mice have been
described (19)
in which increased expression of skeletal
-actin and reactivation of the SM -actin were observed in the
hearts of newborn homozygous mutants. Increased levels of these other
myogenic -actins were insufficient to maintain myofibrillar
integrity in response for the loss of the -cardiac actin, and most
of these cardiac -actin null mice do not survive to birth. Although
mice lacking cardiac -actin can be rescued up to adulthood by the
ectopic expression of enteric smooth muscle -actin using the cardiac
-myosin heavy chain promoter, the hearts of these rescued cardiac
-actin-deficient mice are extremely hypodynamic, considerably
enlarged, and hypertrophied. Furthermore, the transgenic expressed
enteric smooth muscle -actin reduces cardiac contractility in
wild-type and heterozygous mice. The activation and substitution of
alternative actin isoforms are insufficient to correct the deficit of
striated and SM actins, supporting specialized functions of these actin
isoforms. In this regard, the SM -actin null mouse may also serve as
a model to further investigate the role SM -actin plays in wound
repair (33
, 34)
, certain types of cancer
(35
36)
, liver disease (37)
, and kidney
disease (38
, 39)
.
In conclusion, the complete absence of SM -actin transcripts and
protein did not appear to disrupt the formation of the cardiovascular
system. However, we found compromised vascular contractility, lowered
blood pressure, and reduced blood flow in SM -actin null mice.
Unexpectedly, skeletal -actin expression was activated in the
vasculature of these null mice, demonstrating greater flexibility in
the genetic reprogramming of vascular smooth muscle tissues than
previously imagined.
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ACKNOWLEDGMENTS |
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Received for publication October 26, 2000.
Revision received May 16, 2000.
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REFERENCES |
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-smooth muscle (aortic) actin gene: an actin gene which produces multiple mRNAs. J. Biol. Chem. 261,8965-8976-vascular and gamma enteric smooth muscle isoactins: isolation and characterization of rat gamma enteric actin cDNA. Mol. Cell. Biol. 8,5224-5231-actin genes during avian cardiogenesis: vascular smooth muscle -actin gene transcripts mark the onset of cardiomyocyte differentiation. J. Cell Biol. 107,2575-2586-smooth muscle actin is transiently expressed in embryonic rat cardiac and skeletal muscles. Differentiation 39,161-166[Medline]
-sarcomeric actin, -smooth muscle actin and desmin during myogenesis in rat and mouse embryos, in skeletal muscle. Differentiation 44,132-142[Medline]
-actin gene contains evolutionarily conserved sequence motifs within a functional promoter. J. Biol. Chem. 265,16667-16675-smooth muscle actin: a new probe for smooth muscle differentiation. J. Cell Biol. 103,2787-2796This article has been cited by other articles:
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 D. F. Chang, N. S. Belaguli, J. Chang, and R. J. Schwartz LIM-only protein, CRP2, switched on smooth muscle gene activity in adult cardiac myocytes PNAS, January 2, 2007; 104(1): 157 - 162. [Abstract] [Full Text] [PDF]
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M. Takeji, T. Moriyama, S. Oseto, N. Kawada, M. Hori, E. Imai, and T. Miwa Smooth Muscle {alpha}-Actin Deficiency in Myofibroblasts Leads to Enhanced Renal Tissue Fibrosis J. Biol. Chem., December 29, 2006; 281(52): 40193 - 40200. [Abstract] [Full Text] [PDF]
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 A. Hosoya, H. Nakamura, T. Ninomiya, K. Yoshiba, N. Yoshiba, H. Nakaya, S. Wakitani, H. Yamada, E. Kasahara, and H. Ozawa Immunohistochemical Localization of {alpha}-Smooth Muscle Actin During Rat Molar Tooth Development J. Histochem. Cytochem., December 1, 2006; 54(12): 1371 - 1378. [Abstract] [Full Text] [PDF]
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K. Ichimura, H. Kurihara, and T. Sakai Involvement of Mesangial Cells Expressing {alpha}-Smooth Muscle Actin During Restorative Glomerular Remodeling in Thy-1.1 Nephritis J. Histochem. Cytochem., November 1, 2006; 54(11): 1291 - 1301. [Abstract] [Full Text] [PDF]
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2.5°C) resulted in a relative fivefold increase in peak tail
artery blood flow in normal mice whereas in the SM