In vitro segregation of different cell lines with neuronal and glial properties from a stem cell line of rat neurotumor RT4.

Tomozawa, Yasuko; Sueoka, Noboru

doi:10.1073/pnas.75.12.6305

Cited by 37 publications

(17 citation statements)

References 26 publications

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“…1 cific proteins is sequentially lost as the cell further differentiates. For example, the loss of ion channels as cells mature has frequently been documented (see, for example, Hagiwara and Miyazaki, 1977;Spitzer, 1979) and individual CNS cells have been described that share traits of both nerve and glia (DeVitry et al, 1980;Schubert et al, 1974;Tomozawa and Sueoka, 1978;Wilson et al, 1981).…”

Section: Extracellular Proteinsmentioning

confidence: 99%

Protein complexity of central nervous system cell lines

Schubert¹,

Brass²,

Dumas³

1986

J. Neurosci.

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The total extracellular proteins and the most abundant 960 intracellular proteins of clonal CNS nerve and glial cell lines were examined by quantitative 2-dimensional acrylamide gel electrophoresis. While less than 0.2% of the intracellular proteins differ among the 5 nerve and 4 glial cell lines studied, over 65% of the extracellular proteins vary in distribution between the 2 major classes of CNS cells. These data indicate that the phenotypic complexity of nerve and glia populations is similar and that most of the protein complexity is in extracellular molecules.It has been argued on the basis of RNA hybridization studies that the mammalian CNS has a severalfold higher number of unique mRNA sequences than other tissues (Brown and Church, 1972;Chikaraishi et al., 1983; Hahn and Laird, 197 1). Assuming that these mRNAs are translated, there are 2 alternatives that could explain the apparent increase in protein complexity within the brain; they are not mutually exclusive. Each cell within the CNS might make many more species of proteins than cells of other tissues, or there might be a greater number of cell types, each making a few unique species that contribute to total tissue complexity but whose overall protein complexity is similar to cells of different tissues. To distinguish between these alternatives, protein synthesis in a series of clonal CNS nerve and glial cell lines was examined by 2-dimensional gel electrophoresis. Total cellular protein synthesis and the extracellular proteins released into the culture medium were analyzed by computer-assisted methods. It is shown that there is a great deal of variability in the protein species synthesized by cells from the rat CNS and that the variability within the glial population is as great as that between the nerve cells. The majority of this protein complexity is associated with the extracellular proteins, which are more abundant in nerve and glia than in mesodermally derived cells. The total number of cellular proteins synthesized by clonal CNS cell lines is, however, indistinguishable from that of mesodermal cells. It follows that the higher protein complexity in the CNS is due to both the large number of unique phenotypes and the increased number of extracellular proteins relative to other tissues. Materials and MethodsCell lines

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Section: Extracellular Proteinsmentioning

confidence: 99%

Protein complexity of central nervous system cell lines

Schubert¹,

Brass²,

Dumas³

1986

J. Neurosci.

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“…Two others, RT4-B and RT4-D, are derived from a peripheral nerve tumor (23). RT4-B has neuronal attributes (48), whereas RT4-D has gliallike characteristics (23). All four lines were derived from nitrosoethylurea-induced tumors.…”

Section: B Resultsmentioning

confidence: 99%

Cloning of DNA corresponding to rare transcripts of rat brain: evidence of transcriptional and post-transcriptional control and of the existence of nonpolyadenylated transcripts.

Brilliant¹,

Sueoka²,

Chikaraishi³

1984

Mol. Cell. Biol.

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To examine the expression of genes encoding rare transcripts in the rat brain, we have characterized genomic DNA clones corresponding to this class. In brain cells, as in all cell types, rare transcripts constitute the majority of different sequences transcribed. Moreover, when compared with other tissues or cultured cells, brain tissue may be expected to have an even larger set of rare transcripts, some of which could be restricted to subpopulations of neural cells. We have identified seven clones whose transcripts are nonabundant, averaging less than three copies per cell. Clone rgl3 (rat genomic 13) RNA was detected only in the brain, whereas RNA of a second clone, rg4O, was also detected in the brain and in a melanoma. Transcripts of rgl3 were found in cerebellum, cerebral cortex, and regions underlying the cortex, whereas rg4O transcripts were not detected in the cerebellum. Transcripts of both rgl3 and rg4O were found in pelleted polysomal RNA. RNA of another clone, rg34, was found in the brain, liver, and kidney but was found in pelleted polysomal RNA only in the brain, suggesting that its expression may be post-transcriptionally controlled. The remaining four clones represent rare transcripts that are common to the brain, liver, and kidney; rgl8 RNA is restricted to the nucleus, whereas rg3, rg26, and rg36 transcripts are found in the cytoplasm of all three tissues. Transcripts of the brain-specific clone, rgl3, and the commonly expressed clone, rg3, are nonpolyadenylated, presumably belonging to the high-complexity, nonpolyadenylated class of transcripts in the mammalian brain.The mammalian brain, the most complex organ in terms of function and structure, is also the most complex in terms of the diversity of its transcripts. The diversity or complexity of an RNA population can be measured by hybridizing RNA in excess to trace amounts of radioactive, nonrepetitive (unique-sequence) DNA, such that the amount of DNA driven into hybrids at saturation gives a direct measure of the percentage of nonrepetitive DNA transcribed. Complexity measurements have consistently shown that brain RNA in rodents displays a greater complexity than RNA from other tissues (mouse tissue [2,6,15,16,19,26,49]; rat tissue [8,9,13,14,17,25,28,43,47,51]). For example, Chikaraishi et al. (9) have found the nuclear RNA complexity of rat brain (31.2% of unique-sequence DNA coding capacity, assuming asymmetric transcription) to be higher than that of liver (21.8%) or kidney (10.6%).A higher proportion of nuclear RNA complexity (nearly two-thirds) is conserved as cytoplasmic or polysomal RNA in the brain than in the liver or kidney (3, 8)

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“…This result suggests that GFAP gene expression is coordinately coupled with the expression of other glial properties during cell-type conversion. In addition, the RT4-AC and RT4-D sublines were found to significantly express GFAP only at high cell densities and not during logarithmic growth and to express GFAP precociously during morphological differentiation following treatment with 1 mM N6,02'-dibutyryladenosine 3',5'-cyclic monophosphate. These observations closely reflect reports of glial filament expression in astrocyte cultures, suggesting that a common regulatory mechanism is employed by central and peripheral nervous system glia.…”

Section: Introductionmentioning

confidence: 99%

“…Our laboratory has described the isolation and characterization of clonal cell lines from the peripheral nervous system (PNS) tumor RT4 (1)(2)(3)(4)(5)(6)(7)(8). The RT4 tumor was induced by ethylnitrosourea injection of a newborn BDIX rat, a procedure that predominantly gives rise to PNS tumors (1).…”

Section: Introductionmentioning

confidence: 99%

Induction and segregation of glial intermediate filament expression in the RT4 family of peripheral nervous system cell lines.

Freeman¹,

Sueoka²

1987

Proc. Natl. Acad. Sci. U.S.A.

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We have found glial fibrillary acidic protein (GFAP), the major component of astrocyte intermediate fiaments, to be expressed in cell lines of the RT4 peripheral neurotumor family. The RT4 family is a "stem-cell-like" cell line, RT4-AC, that spontaneously undergoes differentiation in culture to three derivative cell types. This process, termed cell-type conversion, results in a segregation among the derivative cell types of parental cell phenotypes that have been described as neuronal-like or glial-like. We have identified a 50-kDa GFAP-immunoreactive cytoskeletal protein and GFAP mRNA in continuous RT4-AC and RT4-D (glial-type derivative) cell lines, but not in two presumptive neuronal-type cell lines. This result suggests that GFAP gene expression is coordinately coupled with the expression of other glial properties during cell-type conversion. In addition, the RT4-AC and RT4-D sublines were found to significantly express GFAP only at high cell densities and not during logarithmic growth and to express GFAP precociously during morphological differentiation following treatment with 1 mM N6,02'-dibutyryladenosine 3',5'-cyclic monophosphate. These observations closely reflect reports of glial filament expression in astrocyte cultures, suggesting that a common regulatory mechanism is employed by central and peripheral nervous system glia.Our laboratory has described the isolation and characterization of clonal cell lines from the peripheral nervous system (PNS) tumor RT4 (1-8). The RT4 tumor was induced by ethylnitrosourea injection of a newborn BDIX rat, a procedure that predominantly gives rise to PNS tumors (1). An unstable cell line, termed RT4-AC, has been described (1, 3) that expresses the glial marker protein S-100 and that also displays voltage-sensitive Na' and K+ channels. The RT4-AC cell line was described as a "stem-cell-type" line because of its ability to repeatedly give rise to three distinct progeny cell types in culture (Fig. 1). These derivative cell types have been described as glial-like (the RT4-D cell type), based on the expression of S-100 and on the loss of the excitable membrane phenotype, or neuronal-like (the RT4-B and RT4-E cell types), based on the loss of S-100 expression and on the retention and enhancement of the excitable membrane phenotype. This differentiation phenomenon has been termed cell-type conversion, and the apparent coordinate segregation of "glial" or "neuronal" properties with conversion has been termed conversion coupling. Clonal RT4 cell lines corresponding to these four cell types have been passaged continuously in culture for over 5 years.Droms and Sueoka (8) have shown that a normally unexpressed high-affinity y-aminobutyric acid uptake mechanism, generally considered to be a glial property in the PNS, can be induced in the RT4-D cell line by a combination treatment with N6,02'-dibutyryladenosine 3',5'-cyclic monophosphate (Bt2cAMP) and testololactone, a synthetic androgen. Under the conditions described, the high-affinity y-aminobutyric acid uptake mecha...

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In vitro segregation of different cell lines with neuronal and glial properties from a stem cell line of rat neurotumor RT4.

Cited by 37 publications

References 26 publications

Protein complexity of central nervous system cell lines

Protein complexity of central nervous system cell lines

Cloning of DNA corresponding to rare transcripts of rat brain: evidence of transcriptional and post-transcriptional control and of the existence of nonpolyadenylated transcripts.

Induction and segregation of glial intermediate filament expression in the RT4 family of peripheral nervous system cell lines.

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