Search PubMed⌕ Search

Biomedical subjects

E N Olson

Publications and source records attributed to E N Olson.

At least 163 records · Page 9Linked to original sources
PubMed1996

SM22 alpha, a marker of adult smooth muscle, is expressed in multiple myogenic lineages during embryogenesis.

SM22 alpha is a calponin-related protein that is expressed specifically in adult smooth muscle. To begin to define the mechanisms that regulate the establishment of the smooth muscle lineage, we analyzed the expression pattern of the SM22 alpha gene during mouse embryogenesis. In situ hybridization demonstrated that SM22 alpha transcripts were first expressed in vascular smooth muscle cells at about embryonic day (E) 9.5 and thereafter continued to be expressed in all smooth muscle cells into adulthood. In contrast to its smooth muscle specificity in adult tissues, SM22 alpha was expressed transiently in the heart between E8.0 and E12.5 and in skeletal muscle cells in the myotomal compartment of the somites between E9.5 and E12.5. The expression of SM22 alpha in smooth muscle cells, as well as early cardiac and skeletal muscle cells, suggests that there may be commonalities between the regulatory programs that direct muscle-specific gene expression in these three myogenic cell types.

Amino Acid Sequence↗
PubMed1996

Myocyte enhancer binding factor-2 expression and activity in vascular smooth muscle cells. Association with the activated phenotype.

Proliferation and phenotypic modulation of smooth muscle cells (SMCs) are major components of the vessel's response to injury in experimental models of restenosis. Some of the growth factors involved in restenosis have been identified, but to date little is known about the transcription factors that ultimately regulate this process. We examined the expression of the four members of the myocyte enhancer binding factor-2 (MEF2) family of transcription factors in cultured rat aortic SMCs (RASMCs) and a rat model of restenosis because of their known importance in regulating the differentiated phenotype of skeletal and cardiac muscle. In skeletal and cardiac muscle, the MEF2s are believed to be important for activating the expression of contractile protein and other muscle-specific genes. Therefore, we anticipated that the MEF2s would be expressed at high levels in medial SMCs that are producing contractile proteins and that they would be downregulated along with the contractile protein genes in neointimal SMCs. On the contrary, we observe that MEF2A, MEF2B, and MEF2D mRNAs are upregulated in the neointima, with the highest levels in the layer of cells nearest to the lumen, whereas MEF2C mRNA levels do not appreciably increase. Moreover, few cells in the media are making MEF2 proteins detectable by immunohistochemistry, whereas large numbers of neointimal cells are positive for all four MEF2s. These data suggest that the MEF2s are involved in the activated smooth muscle phenotype and not in the maintenance of contractile protein gene expression.

Animals↗
PubMed1996

Sclerotome-related helix-loop-helix type transcription factor (scleraxis) mRNA is expressed in osteoblasts and its level is enhanced by type-beta transforming growth factor.

Scleraxis is a recently identified transcription factor with a basic helix-loop-helix motif, which is expressed in sclerotome during embryonic development. We have examined the expression of scleraxis mRNA in rat osteoblastic cells and found that the scleraxis gene was expressed as a 1.2 kb mRNA species in osteoblastic osteosarcoma ROS17/2.8 cells. The scleraxis mRNA expression was enhanced by type-beta transforming growth factor (TGF beta) treatment. The TGF beta effect was observed in a dose-dependent manner starting at 0.2 ng/ml and saturating at 2 ng/ml. The effect was time-dependent and was first observed within 12 h and peaked at 24 h. The TGF beta effect was blocked by cycloheximide, while no effect on scleraxis mRNA stability was observed. TGF beta treatment enhanced scleraxis-E box (Scx-E) binding activity in the nuclear extracts of ROS17/2.8 cells. Furthermore, TGF beta enhanced transcriptional activity of the CAT constructs which contain the Scx-E box sequence. TGF beta treatment also enhanced scleraxis gene expression in osteoblast-enriched cells derived from primary rat calvaria. These findings indicated for the first time that the novel helix-loop-helix type transcription factor (scleraxis) mRNA is expressed in osteoblasts and its expression is regulated by TGF beta.

Animals↗
PubMed1995

Cooperative activation of muscle gene expression by MEF2 and myogenic bHLH proteins.

Members of the myocyte enhancer factor-2 (MEF2) family of MADS domain transcription factors cannot induce myogenesis in transfected fibroblasts, but when coexpressed with the myogenic basic-helix-loop-helix (bHLH) proteins MyoD or myogenin they dramatically increase the extent of myogenic conversion above that seen with either myogenic bHLH factor alone. This cooperativity required direct interactions between the DNA-binding domains of MEF2 and the myogenic bHLH factors, but only one of the factors needed a transactivation domain, and only one of the factors needed to be bound to DNA. These interactions allow either factor to activate transcription through the other's binding site and reveal a novel mechanism for indirect activation of gene expression via protein-protein interactions between the DNA-binding domains of heterologous classes of transcription factors.

Amino Acid Sequence↗
PubMed1995

A subclass of bHLH proteins required for cardiac morphogenesis.

Skeletal muscle development is controlled by a family of muscle-specific basic helix-loop-helix (bHLH) transcription factors. Two bHLH genes, dHAND and eHAND, have now been isolated that are expressed in the bilateral heart primordia and subsequently throughout the primitive tubular heart and its derivatives during chick and mouse embryogenesis. Incubation of stage 8 chick embryos with dHAND and eHAND antisense oligonucleotides revealed that either oligonucleotide alone had no effect on embryonic development, whereas together they arrested development at the looping heart tube stage. Thus, dHAND and eHAND may play redundant roles in the regulation of the morphogenetic events of vertebrate heart development.

Actins↗
PubMed1995

Inactivation of the myogenic bHLH gene MRF4 results in up-regulation of myogenin and rib anomalies.

The myogenic basic helix-loop-helix (bHLH) proteins MyoD, myf5, myogenin, and MRF4 can initiate myogenesis when expressed in nonmuscle cells. During embryogenesis, each of the myogenic bHLH genes is expressed in a unique temporospatial pattern within the skeletal muscle lineage, suggesting that they play distinct roles in muscle development. Gene targeting has shown that MyoD and myf5 play partially redundant roles in the genesis of myoblasts, whereas myogenin is required for terminal differentiation. MRF4 is expressed transiently in the somite myotome during embryogenesis and then becomes up-regulated during late fetal development to eventually become the predominant myogenic bHLH factor expressed in adult skeletal muscle. On the basis of its expression pattern, it has been proposed that MRF4 may regulate skeletal muscle maturation and aspects of adult myogenesis. To determine the function of MRF4, we generated mice carrying a homozygous germ-line mutation in the MRF4 gene. These mice showed only a subtle reduction in expression of a subset of muscle-specific genes but showed a dramatic increase in expression of myogenin, suggesting that it may compensate for the absence of MRF4 and demonstrating that MRF4 is required for the down-regulation of myogenin expression that normally occurs in postnatal skeletal muscle. Paradoxically, MRF4-null mice exhibited multiple rib anomalies, including extensive bifurcations, fusions, and supernumerary processes. These results demonstrate an unanticipated regulatory relationship between myogenin and MRF4 and suggest that MRF4 influences rib outgrowth through an indirect mechanism.

Animals↗
PubMed1995

The paired-like homeo box gene MHox is required for early events of skeletogenesis in multiple lineages.

Formation of cartilage and bone involves sequential processes in which undifferentiated mesenchyme aggregates into primordial condensations that subsequently grow and differentiate, eventually forming the adult skeleton. Although much has been learned about the structural molecules that compose cartilage and bone, little is known about the nuclear factors that regulate chondrogenesis and osteogenesis. MHox is a homeo box-containing gene that is expressed in the mesenchyme of facial, limb, and vertebral skeletal precursors during mouse embryogenesis. MHox expression has been shown to require epithelial-derived signals, suggesting that MHox may regulate the epithelial-mesenchymal interactions required for skeletal organogenesis. To determine the functions of MHox, we generated a loss-of-function mutation in the MHox gene. Mice homozygous for a mutant MHox allele die soon after birth and exhibit defects of skeletogenesis, involving the loss or malformation of craniofacial, limb, and vertebral skeletal structures. The affected skeletal elements are derived from the cranial neural crest, as well as somitic and lateral mesoderm. Analysis of the mutant phenotype during ontogeny demonstrated a defect in the formation and growth of chondrogenic and osteogenic precursors. These findings provide evidence that MHox regulates the formation of preskeletal condensations from undifferentiated mesenchyme.

Abnormalities, Multiple↗
PubMed1995

The mouse MRF4 promoter is trans-activated directly and indirectly by muscle-specific transcription factors.

MRF4 is a member of the basic helix-loop-helix (bHLH) family of muscle-specific transcription factors, which also includes MyoD, myogenin, and myf5. The myocyte enhancer binding factor 2 (MEF2) proteins also serve as important muscle-specific transcription factors. In addition to activating the expression of many muscle-specific structural genes, various members of these two classes of proteins activate their own expression and the expression of each other in a complex transcriptional network that results in the establishment and maintenance of the muscle phenotype. To begin to determine how the expression of MRF4 is regulated by other muscle-specific transcription factors, we have isolated a region of the MRF4 gene that confers muscle-specific expression and have analyzed this promoter region for cis-acting elements involved in trans-activation by the myogenic bHLH and MEF2 transcription factors. Here, we show that in 10T1/2 fibroblasts the MRF4 promoter is trans-activated by myogenin, MyoD, myf5, and by the MEF2 factors, but that MRF4 does not activate expression of its own promoter. Myogenin activated the MRF4 promoter directly by an E box-dependent mechanism, while MEF2 factors activated the promoter through an indirect pathway. The E box-dependent regulation of the MRF4 promoter is in contrast to the regulation of the myogenin and MyoD promoters and may represent a mechanism for the differential expression of these factors during myogenesis.

Animals↗
PubMed1995

p53-independent expression of p21Cip1 in muscle and other terminally differentiating cells.

Terminal differentiation is coupled to withdrawal from the cell cycle. The cyclin-dependent kinase inhibitor (CKI) p21Cip1 is transcriptionally regulated by p53 and can induce growth arrest. CKIs are therefore potential mediators of developmental control of cell proliferation. The expression pattern of mouse p21 correlated with terminal differentiation of multiple cell lineages including skeletal muscle, cartilage, skin, and nasal epithelium in a p53-independent manner. Although the muscle-specific transcription factor MyoD is sufficient to activate p21 expression in 10T1/2 cells, p21 was expressed in myogenic cells of mice lacking the genes encoding MyoD and myogenin, demonstrating that p21 expression does not require these transcription factors. The p21 protein may function during development as an inducible growth inhibitor that contributes to cell cycle exit and differentiation.

Animals↗
PubMed1995

Requirement of MADS domain transcription factor D-MEF2 for muscle formation in Drosophila.

Members of the myocyte enhancer binding factor-2 (MEF2) family of MADS (MCM1, agamous, deficiens, and serum response factor) box transcription factors are expressed in the skeletal, cardiac, and smooth muscle lineages of vertebrate and Drosophila embryos. These factors bind an adenine-thymidine-rich DNA sequence associated with muscle-specific genes. The function of MEF2 was determined by generating a loss-of-function of the single mef2 gene in Drosophila (D-mef2). In loss-of-function embryos, somatic, cardiac, and visceral muscle cells did not differentiate, but myoblasts were normally specified and positioned. These results demonstrate that different muscle cell types share a common myogenic differentiation program controlled by MEF2.

Animals↗
PubMed1995

Activation of the myogenin promoter during mouse embryogenesis in the absence of positive autoregulation.

Myogenin, a member of the MyoD family of helix-loop-helix proteins, can induce myogenesis in a wide range of cell types. In addition to activating muscle structural genes, members of the MyoD family can autoactivate their own and cross-activate one another's expression in transfected cells. This has led to the hypothesis that autoregulatory loops among these factors provide a mechanism for amplifying and maintaining the muscle-specific gene expression program in vivo. Here, we make use of myogenin-null mice to directly test this hypothesis. To investigate whether the myogenin protein autoregulates the myogenin gene during embryogenesis, we introduced a myogenin-lacZ transgene into mice harboring a null mutation at the myogenin locus. Despite a severe deficiency of skeletal muscle in myogenin-null neonates, the myogenin-lacZ transgene was expressed normally in myogenic cells throughout embryogenesis. These results show that myogenin is not required for regulation of the myogenin gene and argue against the existence of a myogenin autoregulatory loop in the embryo.

Animals↗
PubMed1995

Myogenin's functions do not overlap with those of MyoD or Myf-5 during mouse embryogenesis.

The four myogenic basic helix-loop-helix proteins, MyoD, myogenin, Myf-5, and MRF4, can each activate skeletal muscle differentiation when introduced into nonmuscle cells. During embryogenesis, each of these genes is expressed in a unique but overlapping pattern in skeletal muscle precursors and their descendants. Gene knockout experiments have shown that MyoD and Myf-5 play seemingly redundant roles in the generation of myoblasts. However, the role of either of these genes during differentiation in vivo has not been determined. In contrast, a myogenin-null mutation blocks differentiation and results in a dramatic decrease in muscle fiber formation, yet the role of myogenin in the generation or maintenance of myoblast populations is not known. Because myogenin possesses the same myogenic activity as MyoD and Myf-5 in vitro and the expression patterns of these three genes overlap in vivo, we sought to determine if myogenin shares certain functions with either MyoD or Myf-5 in vivo. We therefore generated mice with double homozygous null mutations in the genes encoding MyoD and myogenin or Myf-5 and myogenin. These mice showed embryonic and perinatal phenotypes characteristic of the combined defects observed in mice mutant for each gene alone. As shown by histological analysis and expression of muscle-specific genes, the numbers of undifferentiated myoblasts and residual myofibers were comparable between myogenin-mutant homozygotes and the double-mutant homozygotes. Myoblasts isolated from neonates of the combined mutant genotypes underwent myogenesis in tissue culture, indicating that no more than two of the four myogenic factors are required to support muscle differentiation. These results demonstrate that the functions of myogenin do not overlap with those of MyoD or Myf-5 and support the view that myogenin acts in a genetic pathway downstream of MyoD and Myf-5.

Animals↗
PubMed1995

Dermo-1: a novel twist-related bHLH protein expressed in the developing dermis.

Transcription factors belonging to the basic helix-loop-helix (bHLH) family have been shown to control differentiation of a variety of cell types. Tissue-specific bHLH proteins dimerize preferentially with ubiquitous bHLH proteins to form heterodimers that bind the E-box consensus sequence (CANNTG) in the control regions of target genes. Using the yeast two-hybrid system to screen for tissue-specific bHLH proteins, which dimerize with the ubiquitous bHLH protein E12, we cloned a novel bHLH protein, named Dermo-1. Within its bHLH region, Dermo-1 shares extensive homology with members of the twist family of bHLH proteins, which are expressed in embryonic mesoderm. During mouse embryogenesis, Dermo-1 showed an expression pattern similar to, but distinct from, that of mouse twist. Dermo-1 was expressed at a low level in the sclerotome and dermatome of the somites, and in the limb buds at Day 10.5 post coitum (p.c.), and accumulated predominantly in the dermatome, prevertebrae, and the derivatives of the branchial arches by Day 13.5 p.c. As differentiation of prechondrial cells proceeded, Dermo-1 expression became restricted to the perichondrium. Expression of Dermo-1 increased continuously in the dermis through Day 17.5 p.c. and was also detected in the dermis of neonates, but became downregulated in adult tissues. The Dermo-1 protein bound the E-box consensus sequence in the presence of E12, but transcriptional activity was not detectable. Instead, Dermo-1 repressed transcriptional activity of myogenic bHLH proteins. The expression pattern of Dermo-1 suggests that it functions as a regulator of gene expression in a subset of mesenchymal cell lineages including developing dermis.

Amino Acid Sequence↗
PubMed1995

Paraxis: a basic helix-loop-helix protein expressed in paraxial mesoderm and developing somites.

During vertebrate embryogenesis, cells from the paraxial mesoderm coalesce in a rostral-to-caudal progression to form the somites. Subsequent compartmentalization of the somites yields the sclerotome, myotome, and dermatome, which give rise to the axial skeleton, axial musculature, and dermis, respectively. Recently, we cloned a novel basic helix-loop-helix (bHLH) protein, called scleraxis, which is expressed in the sclerotome, in mesenchymal precursors of bone and cartilage, and in connective tissues. Here we report the cloning of a bHLH protein, called paraxis, which is nearly identical to scleraxis within the bHLH region but diverges in its amino and carboxyl termini. During mouse embryogenesis, paraxis transcripts are first detected at about Day 7.5 postcoitum within primitive mesoderm lying posterior to the head and heart primordia. Subsequently, paraxis expression progresses caudally through the paraxial mesoderm, immediately preceding somite formation. Paraxis is expressed at high levels in newly formed somites before the first detectable expression of the myogenic bHLH genes, and as the somite becomes compartmentalized, paraxis becomes downregulated in the myotome. Paraxis and scleraxis are coexpressed in the sclerotome, but paraxis expression declines soon after sclerotome formation, whereas scleroaxis expression increases in the sclerotome and its derivatives. The sequential expression of paraxis and scleraxis in the paraxial mesoderm and somites suggests that these bHLH proteins may comprise part of a regulatory pathway involved in patterning of the paraxial mesoderm and in the establishment of somitic cell lineages.

Amino Acid Sequence↗
PubMed1995

Expression of the novel basic helix-loop-helix gene eHAND in neural crest derivatives and extraembryonic membranes during mouse development.

We employed the yeast two-hybrid technique to screen a mouse embryo cDNA library for novel tissue-specific Class B basic helix-loop-helix (bHLH) transcription factors, which heterodimerize with the ubiquitously expressed Class A bHLH protein E12. From this screen, we cloned a novel bHLH protein, which we named eHAND. Its low sequence identity with other bHLH family members and unique expression pattern during development suggest that eHAND defines a new subclass of Class B bHLH proteins. eHAND was expressed at high levels in trophoblast cells and extraembryonic membranes throughout development. The first site of eHAND expression in embryos was the heart, where it was expressed at high levels between 8.5 and 10.5 days post coitum (d.p.c.), after which transcript levels declined abruptly. By 13.5 d.p.c., eHAND expression in the heart was localized to regions of valve formation. Expression in other regions of the embryo was confined to tissues with a substantial neural crest component. eHAND was expressed in the first branchial arch and its derivatives, in the sympathoadrenal lineage, and in the enteric systems. The expression pattern of eHAND during development is distinct from that of other bHLH genes and suggests that it has a role in formation of extraembryonic tissues, heart, and neural crest derivatives.

Amino Acid Sequence↗
PubMed1995

A series of mutations in the D-MEF2 transcription factor reveal multiple functions in larval and adult myogenesis in Drosophila.

The D-mef2 gene encodes a MADS domain transcription factor expressed in differentiated muscles and their precursors in the Drosophila embryo. Embryos deficient for D-MEF2 protein due to a deletion of upstream transcriptional control sequences fail to form muscle, suggesting that the gene is required for muscle cell differentiation. To directly demonstrate a role for D-mef2 in embryonic myogenesis, we isolated gene mutants containing EMS-induced point mutations, characterized the effects of these mutations on D-MEF2 protein stability and nuclear localization, and analyzed the resulting muscle phenotypes. Our results show that in the somatic muscle lineage, D-mef2 is required for both the formation and patterning of body wall muscle. In the absence of somatic myogenesis, there is extensive apoptosis among the myoblast cell population. In contrast, in the cardiac muscle lineage, morphogenesis of the dorsal vessel occurs normally but the three myosin subunit genes are not expressed. Mutant embryos also exhibit an abnormal midgut morphology, which correlates with the absence of alpha PS2 integrin gene expression and muscle-specific enhancer function, suggesting that D-mef2 regulates the inflated locus which encodes this integrin subunit. D-MEF2 is also expressed in adepithelial cells and rare D-mef2 transheterozygous mutant adults fail to fly, consistent with defects observed in the indirect flight muscles. These results demonstrate that the D-mef2 gene has multiple functions in myogenesis and tissue morphogenesis during Drosophila development.

Animals↗
PubMed1995

PICK1: a perinuclear binding protein and substrate for protein kinase C isolated by the yeast two-hybrid system.

Protein kinase C (PKC) plays a central role in the control of proliferation and differentiation of a wide range of cell types by mediating the signal transduction response to hormones and growth factors. Upon activation by diacylglycerol, PKC translocates to different subcellular sites where it phosphorylates numerous proteins, most of which are unidentified. We used the yeast two-hybrid system to identify proteins that interact with activated PKC alpha. Using the catalytic region of PKC fused to the DNA binding domain of yeast GAL4 as "bait" to screen a mouse T cell cDNA library in which cDNA was fused to the GAL4 activation domain, we cloned several novel proteins that interact with C-kinase (PICKs). One of these proteins, designated PICK1, interacts specifically with the catalytic domain of PKC and is an efficient substrate for phosphorylation by PKC in vitro and in vivo. PICK1 is localized to the perinuclear region and is phosphorylated in response to PKC activation. PICK1 and other PICKs may play important roles in mediating the actions of PKC.

Amino Acid Sequence↗