An alternative non-tyrosine protein kinase product of the c-src gene in chicken skeletal muscle

1990 ◽  
Vol 10 (8) ◽  
pp. 4068-4079
Author(s):  
T Dorai ◽  
L H Wang
Keyword(s):  
Skeletal Muscle ◽  
Kinase Domain ◽  
Specific Protein ◽  
Protein Product ◽  
Upstream Region ◽  
Tyrosine Kinase Domain ◽  
Adult Skeletal Muscle ◽  
Src Gene

While the c-src locus is expressed as a 4.0-kilobase (kb) mRNA coding for pp60c-src in various chicken tissues, including embryonic muscle, it is expressed as a novel 3.0-kb mRNA in adult skeletal muscle. We have analyzed the primary structure of this alternatively transcribed and spliced c-src mRNA. The sequence revealed three open reading frames, with the previously defined c-src exons 1 through 5 or 6 comprising the third, on the 3' untranslated region of this 3-kb mRNA. The exons coding for the tyrosine kinase domain of pp60c-src were excluded. On the 5' side, 2 kb of sequence upstream from the previously defined exon 1 of the c-src gene was included in this mRNA. The start site for the 3-kb mRNA probably lies downstream of that for the 4-kb mRNA. The first reading frame of the 3.0-kb mRNA, called sur (for src upstream region), encoded a 24-kilodalton (kDa) protein product rich in cysteine and proline residues. In vitro analysis indicated that the 24-kDa sur protein was membrane associated. Antibodies to sur protein detected in vivo a 24-kDa muscle-specific protein which was developmentally regulated and corresponded to the switch from the 4-kb to the 3-kb c-src mRNA. A striking kinetic pattern of appearance of sur protein and disappearance of pp60c-src suggests that the expression of these two proteins is inversely related.

scholarly journals An alternative non-tyrosine protein kinase product of the c-src gene in chicken skeletal muscle.

1990 ◽  
Vol 10 (8) ◽  
pp. 4068-4079 ◽  
Author(s):  
T Dorai ◽  
L H Wang
Keyword(s):  
Skeletal Muscle ◽  
Kinase Domain ◽  
Specific Protein ◽  
Protein Product ◽  
Upstream Region ◽  
Tyrosine Kinase Domain ◽  
Adult Skeletal Muscle ◽  
Src Gene

While the c-src locus is expressed as a 4.0-kilobase (kb) mRNA coding for pp60c-src in various chicken tissues, including embryonic muscle, it is expressed as a novel 3.0-kb mRNA in adult skeletal muscle. We have analyzed the primary structure of this alternatively transcribed and spliced c-src mRNA. The sequence revealed three open reading frames, with the previously defined c-src exons 1 through 5 or 6 comprising the third, on the 3' untranslated region of this 3-kb mRNA. The exons coding for the tyrosine kinase domain of pp60c-src were excluded. On the 5' side, 2 kb of sequence upstream from the previously defined exon 1 of the c-src gene was included in this mRNA. The start site for the 3-kb mRNA probably lies downstream of that for the 4-kb mRNA. The first reading frame of the 3.0-kb mRNA, called sur (for src upstream region), encoded a 24-kilodalton (kDa) protein product rich in cysteine and proline residues. In vitro analysis indicated that the 24-kDa sur protein was membrane associated. Antibodies to sur protein detected in vivo a 24-kDa muscle-specific protein which was developmentally regulated and corresponded to the switch from the 4-kb to the 3-kb c-src mRNA. A striking kinetic pattern of appearance of sur protein and disappearance of pp60c-src suggests that the expression of these two proteins is inversely related.

scholarly journals A Nonerythroid Isoform of Protein 4.1R Interacts with Components of the Contractile Apparatus in Skeletal Myofibers

2000 ◽  
Vol 11 (11) ◽  
pp. 3805-3817 ◽  
Author(s):  
Aikaterini Kontrogianni-Konstantopoulos ◽  
Shu-Ching Huang ◽  
Edward J. Benz
Keyword(s):  
Skeletal Muscle ◽  
Protein S ◽  
Binding Assays ◽  
Adult Skeletal Muscle ◽  
Exon 2 ◽  
Alternatively Spliced ◽  
Translation Initiation Codon ◽  
Protein 4.1R

The ∼80-kDa erythroid 4.1R protein is a major component of the erythrocyte cytoskeleton, where it links transmembrane proteins to the underlying spectrin/actin complexes. A diverse collection of 4.1R isoforms has been described in nonerythroid cells, ranging from ∼30 to ∼210 kDa. In the current study, we identified the number and primary structure of 4.1R isoforms expressed in adult skeletal muscle and characterized the localization patterns of 4.1R message and protein. Skeletal muscle 4.1R appears to originate solely from the upstream translation initiation codon (AUG-1) residing in exon 2′. Combinations of alternatively spliced downstream exons generate an array of distinct 4.1R spliceoforms. Two major isoform classes of ∼105/110 and ∼135 kDa are present in muscle homogenates. 4.1R transcripts are distributed in highly ordered signal stripes, whereas 4.1R protein(s) decorate the sarcoplasm in transverse striations that are in register with A-bands. An ∼105/110-kDa 4.1R isoform appears to occur in vivo in a supramolecular complex with major sarcomeric proteins, including myosin, α-actin, and α-tropomyosin. In vitro binding assays showed that 4.1R may interact directly with the aforementioned contractile proteins through its 10-kDa domain. All of these observations suggest a topological model whereby 4.1R may play a scaffolding role by anchoring the actomyosin myofilaments and possibly modulating their displacements during contraction/relaxation.

scholarly journals Molecular Targets of Androgen Signaling that Characterize Skeletal Muscle Recovery and Regeneration

2015 ◽  
Vol 13 (1) ◽  
pp. nrs.13005 ◽  
Author(s):  
James G. MacKrell ◽  
Benjamin C. Yaden ◽  
Heather Bullock ◽  
Keyue Chen ◽  
Pamela Shetler ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Molecular Targets ◽  
Gene Signature ◽  
Muscle Recovery ◽  
In Vivo Models ◽  
Adult Skeletal Muscle ◽  
Injury Model ◽  
Androgen Regulation

The high regenerative capacity of adult skeletal muscle relies on a self-renewing depot of adult stem cells, termed muscle satellite cells (MSCs). Androgens, known mediators of overall body composition and specifically skeletal muscle mass, have been shown to regulate MSCs. The possible overlapping function of androgen regulation of muscle growth and MSC activation has not been carefully investigated with regards to muscle regeneration. Therefore, the aim of this study was to examine coinciding androgen-mediated genetic changes in an in vitro MSC model and clinically relevant in vivo models. A gene signature was established via microarray analysis for androgen-mediated MSC engagement and highlighted several markers including follistatin (FST), IGF-1, C-X-C chemokine receptor 4 (CXCR4), hepatocyte growth factor (HGF) and glucocorticoid receptor (GR/Nr3c1). In an in vivo muscle atrophy model, androgen re-supplementation significantly increased muscle size and expression of IGF-1, FST, and HGF, while significantly decreasing expression of GR. Biphasic gene expression profiles over the 7-day re-supplementation period identifed temporal androgen regulation of molecular targets involved in satellite cell engagement into myogenesis. In a muscle injury model, removal of androgens resulted in delayed muscle recovery and regeneration. Modifications in the androgen signaling gene signature, along with reduced Pax7 and MyoD expression, suggested that limited MSC activation and increased inflammation contributed to the delayed regeneration. However, enhanced MSC activation in the androgen-deplete mouse injury model was driven by an androgen receptor (AR) agonist. These results provide novel in vitro and in vivo evidence describing molecular targets of androgen signaling, while also increasing support for translational use of AR agonists in skeletal muscle recovery and regeneration.

scholarly journals FGF-2–dependent signaling activated in aged human skeletal muscle promotes intramuscular adipogenesis

2021 ◽  
Vol 118 (37) ◽  
pp. e2021013118 ◽  
Author(s):  
Sebastian Mathes ◽  
Alexandra Fahrner ◽  
Umesh Ghoshdastider ◽  
Hannes A. Rüdiger ◽  
Michael Leunig ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Transcriptional Activation ◽  
Human Skeletal Muscle ◽  
Muscle Growth ◽  
Muscle Cells ◽  
Adeno Associated Virus ◽  
Adult Skeletal Muscle

Aged skeletal muscle is markedly affected by fatty muscle infiltration, and strategies to reduce the occurrence of intramuscular adipocytes are urgently needed. Here, we show that fibroblast growth factor-2 (FGF-2) not only stimulates muscle growth but also promotes intramuscular adipogenesis. Using multiple screening assays upstream and downstream of microRNA (miR)-29a signaling, we located the secreted protein and adipogenic inhibitor SPARC to an FGF-2 signaling pathway that is conserved between skeletal muscle cells from mice and humans and that is activated in skeletal muscle of aged mice and humans. FGF-2 induces the miR-29a/SPARC axis through transcriptional activation of FRA-1, which binds and activates an evolutionary conserved AP-1 site element proximal in the miR-29a promoter. Genetic deletions in muscle cells and adeno-associated virus–mediated overexpression of FGF-2 or SPARC in mouse skeletal muscle revealed that this axis regulates differentiation of fibro/adipogenic progenitors in vitro and intramuscular adipose tissue (IMAT) formation in vivo. Skeletal muscle from human donors aged >75 y versus <55 y showed activation of FGF-2–dependent signaling and increased IMAT. Thus, our data highlights a disparate role of FGF-2 in adult skeletal muscle and reveals a pathway to combat fat accumulation in aged human skeletal muscle.

Induction of resistance to the Abelson inhibitor STI571 in human leukemic cells through gene amplification

Blood ◽  
2000 ◽  
Vol 95 (5) ◽  
pp. 1758-1766 ◽  
Author(s):  
Philipp le Coutre ◽  
Elena Tassi ◽  
Marileila Varella-Garcia ◽  
Rossella Barni ◽  
Luca Mologni ◽  
...  
Keyword(s):  
Cell Line ◽  
Gene Amplification ◽  
Marker Chromosome ◽  
Kinase Domain ◽  
Leukemic Cells ◽  
Tyrosine Kinase Domain ◽  
Fish Analysis ◽  
Human Leukemic Cells

The 2-phenylaminopyrimidine derivative STI571 has been shown to selectively inhibit the tyrosine kinase domain of the oncogenicbcr/abl fusion protein. The activity of this inhibitor has been demonstrated so far both in vitro with bcr/abl expressing cells derived from leukemic patients, and in vivo on nude mice inoculated with bcr/abl positive cells. Yet, no information is available on whether leukemic cells can develop resistance to bcr/ablinhibition. The human bcr/abl expressing cell line LAMA84 was cultured with increasing concentrations of STI571. After approximately 6 months of culture, a new cell line was obtained and named LAMA84R. This newly selected cell line showed an IC50 for the STI571 (1.0 μM) 10-fold higher than the IC50 (0.1 μM) of the parental sensitive cell line. Treatment with STI571 was shown to increase both the early and late apoptotic fraction in LAMA84 but not in LAMA84R. The induction of apoptosis in LAMA84 was associated with the activation of caspase 3–like activity, which did not develop in the resistant LAMA84R cell line. LAMA84R cells showed increased levels of bcr/abl protein and mRNA when compared to LAMA84 cells. FISH analysis with BCR- and ABL-specific probes in LAMA84R cells revealed the presence of a marker chromosome containing approximately 13 to 14 copies of the BCR/ABL gene. Thus, overexpression of the Bcr/Abl protein mediated through gene amplification is associated with and probably determines resistance of human leukemic cells to STI571 in vitro.

scholarly journals Thrombospondin-4, an extracellular matrix protein expressed in the developing and adult nervous system promotes neurite outgrowth.

1995 ◽  
Vol 131 (4) ◽  
pp. 1083-1094 ◽  
Author(s):  
S Arber ◽  
P Caroni
Keyword(s):  
Skeletal Muscle ◽  
Extracellular Matrix ◽  
Nervous System ◽  
Neuromuscular Junction ◽  
Neurite Outgrowth ◽  
Adult Skeletal Muscle ◽  
Wide Range ◽  
Thrombospondin 4

Extracellular matrix (ECM) molecules are involved in multiple aspects of cell-to-cell signaling during development and in the adult. In nervous system development, specific recognition processes, e.g., during axonal pathfinding and synaptogenesis involve modulation and signaling by ECM components. Much less is known about their presence and possible roles in the adult nervous system. We now report that thrombospondin-4 (TSP-4), a recently discovered member of the TSP gene family is expressed by neurons, promotes neurite outgrowth, and accumulates at the neuromuscular junction and at certain synapse-rich structures in the adult. To search for muscle genes that may be involved in neuromuscular signaling, we isolated cDNAs induced in adult skeletal muscle by denervation. One of these cDNAs coded for the rat homologue of TSP-4. In skeletal muscle, it was expressed by muscle interstitial cells. The transcript was further detected in heart and in the developing and adult nervous system, where it was expressed by a wide range of neurons. An antiserum to the unique carboxyl-terminal end of the protein allowed to specifically detect TSP-4 in transfected cells in vitro and on cryostat sections in situ. TSP-4 associated with ECM structures in vitro and in vivo. In the adult, it accumulated at the neuromuscular junction and at synapse-rich structures in the cerebellum and retina. To analyze possible activities of TSP-4 towards neurons, we carried out coculture experiments with stably transfected COS cells and motor, sensory, or retina neurons. These experiments revealed that TSP-4 was a preferred substrate for these neurons, and promoted neurite outgrowth. The results establish TSP-4 as a neuronal ECM protein associated with certain synapse-rich structures in the adult. Its activity towards embryonic neurons in vitro and its distribution in vivo suggest that it may be involved in local signaling in the developing and adult nervous system.

scholarly journals A Novel Activating Mutation in the RET Tyrosine Kinase Domain Mediates Neoplastic Transformation

2006 ◽  
Vol 20 (7) ◽  
pp. 1633-1643 ◽  
Author(s):  
Aaron Cranston ◽  
Cristiana Carniti ◽  
Sam Martin ◽  
Piera Mondellini ◽  
Yvette Hooks ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Cell Line ◽  
Glial Cell ◽  
Neurotrophic Factor ◽  
Functional Characterization ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain ◽  
Exogenous Substrate

Abstract We report the finding of a novel missense mutation at codon 833 in the tyrosine kinase of the RET proto-oncogene in a patient with a carcinoma of the thyroid. In vitro experiments demonstrate that the R833C mutation induces transformed foci only when present in the long 3′ splice isoform and, in keeping with a model in which the receptor has to dimerize to be completely activated, glial cell line-derived neurotrophic factor stimulation leads the RETR833C receptor to a higher level of activation. Tyrosine kinase assays show that the RETR833C long isoform has weak intrinsic kinase activity and phosphorylation of an exogenous substrate is not elevated even in the presence of glial cell line-derived neurotrophic factor. Furthermore, the R833C mutation is capable of sustaining the transformed phenotype in vivo but does not confer upon the transformed cells the ability to degrade the basement membrane in a manner analogous to metastasis. Our functional characterization of the R833C substitution suggests that, like the V804M and S891A mutations, this tyrosine kinase mutation confers a weak activating potential upon RET. This is the first report demonstrating that the introduction of an intracellular cysteine can activate RET. However, this does not occur via dimerization in a manner analogous to the extracellular cysteine mutants.

scholarly journals Fibro-adipogenic progenitors of dystrophic mice are insensitive to NOTCH regulation of adipogenesis

2019 ◽  
Vol 2 (3) ◽  
pp. e201900437 ◽  
Author(s):  
Milica Marinkovic ◽  
Claudia Fuoco ◽  
Francesca Sacco ◽  
Andrea Cerquone Perpetuini ◽  
Giulio Giuliani ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Control Mechanism ◽  
Notch Pathway ◽  
Skeletal Muscle Regeneration ◽  
Wild Type ◽  
Adult Skeletal Muscle ◽  
Pathological Conditions ◽  
Dystrophic Mice

Fibro-adipogenic progenitors (FAPs) promote satellite cell differentiation in adult skeletal muscle regeneration. However, in pathological conditions, FAPs are responsible for fibrosis and fatty infiltrations. Here we show that the NOTCH pathway negatively modulates FAP differentiation both in vitro and in vivo. However, FAPs isolated from young dystrophin-deficient mdx mice are insensitive to this control mechanism. An unbiased mass spectrometry–based proteomic analysis of FAPs from muscles of wild-type and mdx mice suggested that the synergistic cooperation between NOTCH and inflammatory signals controls FAP differentiation. Remarkably, we demonstrated that factors released by hematopoietic cells restore the sensitivity to NOTCH adipogenic inhibition in mdx FAPs. These results offer a basis for rationalizing pathological ectopic fat infiltrations in skeletal muscle and may suggest new therapeutic strategies to mitigate the detrimental effects of fat depositions in muscles of dystrophic patients.

Tristetraprolin and LPS-inducible CXC chemokine are rapidly induced in presumptive satellite cells in response to skeletal muscle injury

2002 ◽  
Vol 115 (13) ◽  
pp. 2701-2712 ◽  
Author(s):  
Chetana Sachidanandan ◽  
Ramkumar Sambasivan ◽  
Jyotsna Dhawan
Keyword(s):  
Skeletal Muscle ◽  
Satellite Cells ◽  
Cell Biology ◽  
Muscle Injury ◽  
Rna Binding ◽  
C2c12 Myoblasts ◽  
Adult Skeletal Muscle ◽  
Cxc Chemokine

Myogenic precursor cells known as satellite cells persist in adult skeletal muscle and are responsible for its ability to regenerate after injury. Quiescent satellite cells are activated by signals emanating from damaged muscle. Here we describe the rapid activation of two genes in response to muscle injury; these transcripts encode LPS-inducible CXC chemokine (LIX), a neutrophil chemoattractant, and Tristetraprolin (TTP), an RNA-binding protein implicated in the regulation of cytokine expression. Using a synchronized cell culture model we show that C2C12 myoblasts arrested in G0 exhibit some molecular attributes of satellite cells in vivo: suppression of MyoD and Myf5 expression during G0 and their reactivation in G1. Synchronization also revealed cell cycle dependent expression of CD34, M-cadherin, HGF and PEA3, genes implicated in satellite cell biology. To identify other genes induced in synchronized C2C12 myoblasts we used differential display PCR and isolated LIX and TTP cDNAs. Both LIX and TTP mRNAs are short-lived, encode molecules implicated in inflammation and are transiently induced during growth activation in vitro. Further, LIX and TTP are rapidly induced in response to muscle damage in vivo. TTP expression precedes that of MyoD and is detected 30 minutes after injury. The spatial distribution of LIX and TTP transcripts in injured muscle suggests expression by satellite cells. Our studies suggest that in addition to generating new cells for repair, activated satellite cells may be a source of signaling molecules involved in tissue remodeling during regeneration.

scholarly journals Skeletal muscle fibro-adipogenic progenitors of dystrophic mice are insensitive to NOTCH-dependent regulation of adipogenesis

2017 ◽  
Author(s):  
Milica Marinkovic ◽  
Francesca Sacco ◽  
Filomena Spada ◽  
Lucia Lisa Petrilli ◽  
Claudia Fuoco ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Cell Mass ◽  
Notch Pathway ◽  
Notch Signaling Pathway ◽  
Skeletal Muscle Regeneration ◽  
Mdx Mice ◽  
List Type ◽  
Adult Skeletal Muscle

SummaryFibro adipogenic progenitors (FAPs) promote satellite cell differentiation in adult skeletal muscle regeneration. However, in pathological conditions, FAPs are responsible for fibrosis and fat infiltrations. Here we show that the NOTCH pathway negatively modulates FAP differentiation both in vitro and in vivo. However, FAPs isolated from young dystrophin-deficient mdx mice are insensitive to this control mechanism. Nonetheless, factors released by hematopoietic cells restore the sensitivity to NOTCH adipogenic inhibition. An unbiased mass spectrometry-based proteomic analysis of FAPs from muscles of wild type and mdx mice, revealed that the synergistic cooperation between NOTCH and inflammatory signals controls FAP differentiation. These results offer a basis for rationalizing the pathological outcomes of fat infiltrations in skeletal muscle and may suggest new therapeutic strategies to mitigate the detrimental effects of fatty depositions in muscles of dystrophic patients.HighlightsSingle-cell mass cytometry reveals that wt and mdx FAPs are in different cell states.Activation of the NOTCH signaling pathway negatively regulates adipogenesis of wt but not mdx FAPs.Deep proteomics suggests a mechanism explaining the different sensitivity of mdx- FAPs to NOTCH.TNF-a stimulation restores the anti-adipogenic effect of NOTCH in mdx FAPs.

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