Protein Synthesis in Isolated Nuclei From Adult Rat Brain

Abstract— The subcellular distribution of cytidine 5′‐monophospho‐N‐acetylneuraminic acid synthetase, which is a key enzyme involved in the biosynthesis of sialo‐glycoproteins and gangliosides, was studied in the frontal grey cortex and the corpus callosum of the calf brain. It appeared that the enzyme was highly concentrated in nuclear fractions, which were shown to be relatively pure as evaluated from morphological, enzymic and chemical data. The possibility that the nuclear localization of the enzyme was due to an artifact, as the result of adsorption onto the nuclear membrane or absorption into the nucleus during the homogenization of the tissue, was ruled out completely and so it appeared that cytidine 5′‐monophospho‐N‐acetylneuraminic acid synthetase, at least for the greater part, is a true nuclear enzyme. The subnuclear localization of the enzyme was shown to be in the nuclear sap. Both neuronal and glial cell nuclei contained the enzyme, which makes it very likely that biosynthesis of sialo‐glycoproteins and/or gangliosides occurs in neurons as well as in glial cells. The significance of the nuclear localization of the enzyme for the biosynthesis of sialic acid containing macromolecules and the possible regulatory role played by the nucleus in this process are discussed.

Section: Fractionation Of the Nuclei With Detergent And Salt Solutionssupporting

confidence: 91%

Section: Identification and Purity Of The Nucleimentioning

confidence: 99%

THE SUBCELLULAR LOCALIZATION OF CYTIDINE 5′‐MONOPHOSPHO‐N‐ACETYLNEURAMINIC ACID SYNTHETASE IN CALF BRAIN

Eijnden

1973

“…The overall DNA recovery, without the use of detergents, was 11 per cent and the cross-contamination of one nuclear type with another was 5-10 per cent. BURDMAN and JOURNEY (1969) fractionated a mixed population of nuclei from rat brain on a similar gradient and found that neuronal nuclei sedimented above 2.4 M-sucrose and astrocytic nuclei above 2.6 M-sucrose, a population of glial nuclei pelleting through 2.6 M-sucrose. The cross-contamination between the nuclear populations was 20 per cent.…”

Section: Discussionmentioning

confidence: 99%

Studies on Rna Synthesis in Two Populations of Nuclei From the Mammalian Cerebral Cortex

Thompson

1973

182

A method is presented for the rapid separation of cell nuclei from the rabbit cerebral cortex into two populations. The first of these consists largely of nuclei with the morphological characteristics of neuronal nuclei, the second almost entirely of nuclei with the morphological characteristics of glial cell nuclei. From studies based upon sensitivity to the toxin α‐amanitin, the ratio of incorporation of different bases, ionic requirements and differential sensitivity to actinomycin D, it is concluded that under both the classical low and high salt conditions described by other workers, two enzymes are active in RNA synthesis. The presence of a third enzyme of low activity cannot be excluded. No qualitative difference in the number of enzymes involved in RNA synthesis in neuronal and glial cell nuclei has been found, but there are quantitative differences in activity between the two nuclear populations.

“…IN ADDITION to the histones, acidic proteins from cell nuclei of liver and thymus are implicated as regulators in the expression of genetic information (WANG, 1967~;WANG, 1968;PLATZ, KISH and KLEINSMITH, 1970). These acidic proteins are of equal if not greater importance than histones in this role for three reasons: First, acidic proteins are metabolically active (DALY, ALLFREY and MIRSKY, 1952;BURDMAN, HAGLID and DRAVID, 1970); secondly, they reverse the inhibition by histones of DNAdependent RNA synthesis (WANG, 1968) and finally, they are less abundant in genetically repressed heterochromatin than in active chromatin (FRENSTER, 1965). Several investigators have reported on the properties of these acidic nuclear proteins from liver and thymus tissues (WANG, 1967b;JOHNS and FORRESTER, 1969); however, there have been relatively few studies (BURDMAN et al, 1970) on brain tissue.…”

mentioning

confidence: 99%

“…The nuclei had little visible cytoplasmic contamination and they retained their morphological characteristics. Two groups of acidic proteins were extracted from the purified cell nuclei according to procedure B of BURDMAN et al (1970) with one exception-the pellet obtained after the 27,OOOg 30-min centrifugation was extracted with 1 M-NaCl instead of 2 M-NaCl. Our Fraction I corresponded to the supernatant fluid obtained after the 20,000 g 20-min centrifugation.…”

mentioning

confidence: 99%

CHARACTERIZATION OF ACIDIC PROTEINS IN CELL ACRYLAMIDE GEL ELECTROPHORESIS NUCLEI FROM RAT BRAIN BY HIGH‐RESOLUTION¹

Davis

Copenhaver

Carver

1972

Abstract— Acidic proteins from cell nuclei of rat brain were evaluated by gel electrophoresis. By the use of a gel containing a high ratio of bis‐acrylamide to acrylamide, 31 bands were demonstrated for this protein fraction. Gels without the high concentration of copolymer lacked the resolving power to separate these proteins. Because the number of bands present in the acidic protein fraction is greater than the number of histones present in cell nuclei, the suggestion that the acidic proteins play a role in genetic regulation seems reasonable.