Expression of a single transfected cDNA converts fibroblasts to myoblasts

doi:10.1016/0092-8674(87)90585-X

Cell

Volume 51, Issue 6, 24 December 1987, Pages 987-1000

https://doi.org/10.1016/0092-8674(87)90585-X Get rights and content

Abstract

5-azacytidine treatment of mouse $C3H10T1 2$ embryonic fibroblasts converts them to myoblasts at a frequency suggesting alteration of one or only a few closely linked regulatory loci. Assuming such loci to be differentially expressed as poly(A)⁺ RNA in proliferating myoblasts, we prepared proliferating myoblast-specific, subtracted cDNA probes to screen a myocyte cDNA library. Based on a number of criteria, three cDNAs were selected and characterized. We show that expression of one of these cDNAs transfected into $C3H10T1 2$ fibroblasts, where it is not normally expressed, is sufficient to convert them to stable myoblasts. Myogenesis also occurs, but to a lesser extent, when this cDNA is expressed in a number of other cell lines. The major open reading frame encoded by this cDNA contains a short protein segment similar to a sequence present in the myc protein family.

References (60)

H.M. Blau et al.
Cytoplasmic activation of human nuclear genes in stable heterocaryons
Cell
(1983)
A.B. Chapman et al.
Analysis of gene expression during differentiation of adipogenic cells in culture and hormonal control of the developmental program
J. Biol. Chem.
(1984)
H. Green et al.
Sublines of mouse 3T3 cells that accumulate lipid
Cell
(1974)
H. Green et al.
Spontaneous heritable changes leading to increased adipose conversion in 3T3 cells
Cell
(1976)
S. Henikoff
Unidirectional digestion with exonuclease III in DNA sequence analysis
Meth. Enzymol.
(1987)
P.A. Jones et al.
Cellular differentiation, cytidine analogs and DNA methylation
Cell
(1980)
K. Kelly et al.
Cell-specific regulation of the c-myc gene by lymphocyte mitogens and platelet-derived growth factor
Cell
(1983)
S.F. Konieczny et al.
5-azacytidine induction of stable mesodermal stem cell lineages from $10T1 2$ cells: evidence for regulatory genes controlling determination
Cell
(1984)
A.B. Lassar et al.
Transfection of a DNA locus that mediates the conversion of $10T1 2$ fibroblasts to myoblasts
Cell
(1986)
J.B. Lawrence et al.
Extinction of muscle-specific properties in somatic cell heterokaryons
Dev. Biol.
(1984)

M. Linial

Creation of a processed pseudogene by retroviral infection

Cell

(1987)

T.A. Linkhart et al.

Myogenic differentiation in permanent clonal mouse myoblast cell lines: regulation by macromolecular growth factors in the culture medium

Dev. Biol.

(1981)

R. Sen et al.

Inducibility of κ immunoglobulin enhancer-binding protein NF-κB by a posttranslational mechanism

Cell

(1986)

S.M. Taylor et al.

Multiple new phenotypes induced in $10T1 2$ and 3T3 cells treated with 5-azacytidine

Cell

(1979)

R. Villares et al.

The achaete-scute gene complex of D. melanogaster: conserved domains in a subset of genes required for neurogenesis and their homology to myc

Cell

(1987)

W.E. Wright et al.

The suppression of myogenic function in heterokaryons formed by fusing chick myocytes to diploid rat fibroblasts

Cell Differ.

(1983)

K. Alitalo et al.

Nucleotide sequence of the v-myc oncogene of avian retrovirus MC29

D. Bader et al.

Immunochemical analysis of myosin heavy chain during avian myogenesis in vivo and in vitro

J. Cell Biol.

(1982)

J.M. Berg

Potential metal-binding domains in nucleic acid binding proteins

Science

(1986)

O. Bernard et al.

Sequence of the murine and human cellular myc oncogenes and two modes of myc transcription resulting from chromosome translocation in B lymphoid tumours

EMBO J.

(1983)

H.M. Blau et al.

Plasticity of the differentiated state

Science

(1985)

C.H. Clegg et al.

Growth factor control of skeletal muscle differentiation: commitment to terminal differentiation occurs in G1 phase and is repressed by fibroblast growth factor

J. Cell Biol.

(1987)

K.F. Conklin et al.

Varied interactions between proviruses and adjacent host chromatin

Mol. Cell. Biol.

(1986)

J.A. Coppola et al.

Constitutive c-myc oncogene expression blocks mouse erythroleukaemia cell differentiation but not commitment

Nature

(1986)

M.M. Davis et al.

Cell-type-specific cDNA probes and the murine I region: the localization and orientation of A^d_a

R.A. DePinho et al.

Structure and expression of the murine N-myc gene

S.R. Dienstman et al.

Myogenesis: a cell lineage interpretation

E. Dmitrovsky et al.

Expression of a transfected human c-myc oncogene inhibits differentiation of a mouse erythroleukaemia cell line

Nature

(1986)

C. Dony et al.

Post-transcriptional control of myc and p53 expression during differentiation of the embryonal carcinoma cell line F9

T. Endo et al.

Transcriptional and posttranscriptional control of c-myc during myogenesis: its mRNA remains inducible in differentiated cells and does not suppress the differentiated phenotype

Mol. Cell. Biol.

(1986)

Cited by (2826)

Targeting long non-coding RNAs in cancer therapy using CRISPR-Cas9 technology: A novel paradigm for precision oncology
2024, Journal of Biotechnology
Cancer is the second leading cause of death worldwide, despite recent advances in its identification and management. To improve cancer patient diagnosis and care, it is necessary to identify new biomarkers and molecular targets. In recent years, long non-coding RNAs (lncRNAs) have surfaced as important contributors to various cellular activities, with growing proof indicating their substantial role in the genesis, development, and spread of cancer. Their unique expression profiles within specific tissues and their wide-ranging functionalities make lncRNAs excellent candidates for potential therapeutic intervention in cancer management. They are implicated in multiple hallmarks of cancer, such as uncontrolled proliferation, angiogenesis, and immune evasion. This review article explores the innovative application of CRISPR-Cas9 technology in targeting lncRNAs as a cancer therapeutic strategy. The CRISPR-Cas9 system has been widely applied in functional genomics, gene therapy, and cancer research, offering a versatile platform for lncRNA targeting. CRISPR-Cas9-mediated targeting of lncRNAs can be achieved through CRISPR interference, activation or the complete knockout of lncRNA loci. Combining CRISPR-Cas9 technology with high-throughput functional genomics makes it possible to identify lncRNAs critical for the survival of specific cancer subtypes, opening the door for tailored treatments and personalised cancer therapies. CRISPR-Cas9-mediated lncRNA targeting with other cutting-edge cancer therapies, such as immunotherapy and targeted molecular therapeutics can be used to overcome the drug resistance in cancer. The synergy of lncRNA research and CRISPR-Cas9 technology presents immense potential for individualized cancer treatment, offering renewed hope in the battle against this disease.
3D organization of enhancers in MuSCs
2024, Current Topics in Developmental Biology
Skeletal muscle stem cells (MuSCs), also known as satellite cells, are essential for muscle growth and injury induced regeneration. In healthy adult muscle, MuSCs remain in a quiescent state located in a specialized niche beneath the basal lamina. Upon injury, these dormant MuSCs can quickly activate to re-enter the cell cycle and differentiate into new myofibers, while a subset undergoes self-renewal and returns to quiescence to restore the stem cell pool. The myogenic lineage progression is intricately controlled by complex intrinsic and extrinsic cues and coupled with dynamic transcriptional programs. In transcriptional regulation, enhancers are key regulatory elements controlling spatiotemporal gene expression through physical contacting promoters of target genes. The three-dimensional (3D) chromatin architecture is known to orchestrate the establishment of proper enhancer-promoter interactions throughout development and aging. However, studies dissecting the 3D organization of enhancers in MuSCs are just emerging. Here, we provide an overview of the general properties of enhancers and newly developed methods for assessing their activity. In particular, we summarize recent discoveries regarding the 3D rewiring of enhancers during MuSC specification, lineage progression as well as aging.
Chromatin organization of muscle stem cell
2024, Current Topics in Developmental Biology
The proper functioning of skeletal muscles is essential throughout life. A crucial crosstalk between the environment and several cellular mechanisms allows striated muscles to perform successfully. Notably, the skeletal muscle tissue reacts to an injury producing a completely functioning tissue. The muscle’s robust regenerative capacity relies on the fine coordination between muscle stem cells (MuSCs or “satellite cells”) and their specific microenvironment that dictates stem cells’ activation, differentiation, and self-renewal. Critical for the muscle stem cell pool is a fine regulation of chromatin organization and gene expression. Acquiring a lineage-specific 3D genome architecture constitutes a crucial modulator of muscle stem cell function during development, in the adult stage, in physiological and pathological conditions. The context-dependent relationship between genome structure, such as accessibility and chromatin compartmentalization, and their functional effects will be analysed considering the improved 3D epigenome knowledge, underlining the intimate liaison between environmental encounters and epigenetics.
Fibroblast Reprogramming in Cardiac Repair
2024, JACC: Basic to Translational Science
Cardiovascular disease is one of the major causes of death worldwide. Limited proliferative capacity of adult mammalian cardiomyocytes has prompted researchers to exploit regenerative therapy after myocardial injury, such as myocardial infarction, to attenuate heart dysfunction caused by such injury. Direct cardiac reprogramming is a recently emerged promising approach to repair damaged myocardium by directly converting resident cardiac fibroblasts into cardiomyocyte-like cells. The achievement of in vivo direct reprogramming of fibroblasts has been shown, by multiple laboratories independently, to improve cardiac function and mitigate fibrosis post–myocardial infarction, which holds great potential for clinical application. There have been numerous pieces of valuable work in both basic and translational research to enhance our understanding and continued refinement of direct cardiac reprogramming in recent years. However, there remain many challenges to overcome before we can truly take advantage of this technique to treat patients with ischemic cardiac diseases. Here, we review recent progress of fibroblast reprogramming in cardiac repair, including the optimization of several reprogramming strategies, mechanistic exploration, and translational efforts, and we make recommendations for future research to further understand and translate direct cardiac reprogramming from bench to bedside. Challenges relating to these efforts will also be discussed.
The novel RNA-RNA activation of H19 on MyoD transcripts promoting myogenic differentiation of goat muscle satellite cells
2023, International Journal of Biological Macromolecules
The elaborate interplay of coding and noncoding factors governs muscle growth and development. Here, we reported a mutual activation between long noncoding RNA (lncRNA) H19 and MyoD (myogenic determination gene number 1) in the muscle process. We successfully cloned the two isoforms of goat H19, which were significantly enriched and positively correlated with MyoD transcripts in skeletal muscles or differentiating muscle satellite cells (MuSCs). To systematically screen genes altered by H19, we performed RNA-seq using cDNA libraries of differentiating H19-deficiency MuSCs and consequently anchored MyoD as the critical genes in mediating H19 function. Intriguingly, some transcripts of MyoD and H19 overlapped in the cytoplasm, which was dramatically damaged when the core complementary nucleotides were mutated. Meanwhile, MyoD RNA successfully pulled down H19 in MS2-RIP experiments. Furthermore, HuR could bind both H19 and MyoD transcripts, while H19 or its truncated mutants successfully stabilized MyoD mRNA, with or without HuR deficiency. In turn, novel functional MyoD protein-binding sites were identified in the promoter and exons of the H19 gene. Our results suggest that MyoD activates H19 transcriptionally, and RNA-RNA hybridization is critical for H19-promoted MyoD expression, which extends our knowledge of the hierarchy of regulatory networks in muscle growth.
Insights and applications of direct neuronal reprogramming
2023, Current Opinion in Genetics and Development
Direct neuronal reprogramming converts somatic cells of a defined lineage into induced neuronal cells without going through a pluripotent intermediate. This approach not only provides access to the otherwise largely inaccessible cells of the brain for neuronal disease modeling, but also holds great promise for ultimately enabling neuronal cell replacement without the use of transplantation. To improve efficiency and specificity of direct neuronal reprogramming, much of the current efforts aim to understand the mechanisms that safeguard cell identities and how the reprogramming cells overcome the barriers resisting fate changes. Here, we review recent discoveries into the mechanisms by which the donor cell program is silenced, and new cell identities are established. We also discuss advancements that have been made toward fine-tuning the output of these reprogramming systems to generate specific types of neuronal cells. Finally, we highlight the benefit of using direct neuronal reprogramming to study age-related disorders and the potential of in vivo direct reprogramming in regenerative medicine.

View all citing articles on Scopus

View full text

Cell

ArticleExpression of a single transfected cDNA converts fibroblasts to myoblasts

Abstract

Cell

J. Biol. Chem.

Cell

Cell

Meth. Enzymol.

Cell

Cell

Cell

Cell

Dev. Biol.

Cell

Dev. Biol.

Cell

Cell

Cell

Cell Differ.

Nucleotide sequence of the v-myc oncogene of avian retrovirus MC29

Immunochemical analysis of myosin heavy chain during avian myogenesis in vivo and in vitro

J. Cell Biol.

Potential metal-binding domains in nucleic acid binding proteins

Science

Sequence of the murine and human cellular myc oncogenes and two modes of myc transcription resulting from chromosome translocation in B lymphoid tumours

EMBO J.

Plasticity of the differentiated state

Science

Growth factor control of skeletal muscle differentiation: commitment to terminal differentiation occurs in G1 phase and is repressed by fibroblast growth factor

J. Cell Biol.

Varied interactions between proviruses and adjacent host chromatin

Mol. Cell. Biol.

Constitutive c-myc oncogene expression blocks mouse erythroleukaemia cell differentiation but not commitment

Nature

Cell-type-specific cDNA probes and the murine I region: the localization and orientation of Ada

Structure and expression of the murine N-myc gene

Myogenesis: a cell lineage interpretation

Expression of a transfected human c-myc oncogene inhibits differentiation of a mouse erythroleukaemia cell line

Nature

Post-transcriptional control of myc and p53 expression during differentiation of the embryonal carcinoma cell line F9

Transcriptional and posttranscriptional control of c-myc during myogenesis: its mRNA remains inducible in differentiated cells and does not suppress the differentiated phenotype

Mol. Cell. Biol.

Article
Expression of a single transfected cDNA converts fibroblasts to myoblasts

Cell-type-specific cDNA probes and the murine I region: the localization and orientation of A^d_a