Actin polymerization and synthesis in cultured neurones.
The protein content of sympathetic neurones explanted from 10-11-day old chick embryos into culture medium containing nerve growth factor (NGF) increases steadily from about 100 to about 400 pg/cell in 7 days. Actin remains at close to 5% of the total protein during this period, but the proportion of unpolymerized actin falls. As measured by the inhibition of DNase I activity, rounded neurones without neurites contain 70 +/- 7% of their total actin in monomeric form, whereas cells in mature, neurite-bearing cultures contain 39 +/- 7%. When allowance is made for the increase in size of the neuronal cell bodies, the actin present in the neurites ('axons') alone is found to be almost entirely in filamentous form. Cultures exposed to radioactive leucine rapidly incorporate radioactivity into both sedimentable and non-sedimentable forms of actin. Actin-specific activities in the two fractions--estimated after isolation of the actin on small DNase I--Sepharose affinity columns--are similar after labelling for less than 1 h. Direct incorporation of newly-synthesized actin into filaments is suggested from these results. Pulse-chase experiments show that non-sedimentable protein in cultured sympathetic neurones turns over more rapidly than sedimentable protein. However, this is not true for actin, which shows a similar specific activity in sedimentable and non-sedimentable forms--even after 6 days of cold chase. This anomalous behaviour is simply explained by an exchange of actin molecules between filamentous and non-filamentous forms. Control experiments indicate that exchange does not occur to this degree during preparation of subcellular fractions. It is consequently attributed to exchange processes in the living cell.
D000199:Actins / Q000378:metabolism*; D000818:Animals; D001369:Axons / Q000648:ultrastructure; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D002642:Chick Embryo; D003850:Deoxyribonuclease I; D004706:Endodeoxyribonucleases / Q000037:antagonists & inhibitors; D005728:Ganglia, Sympathetic / Q000166:cytology*; D007930:Leucine / Q000378:metabolism; D046911:Macromolecular Substances; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D011108:Polymers; D011506:Proteins / Q000378:metabolism
D000199:Actins; D046911:Macromolecular Substances; D011108:Polymers; D011506:Proteins; D004706:Endodeoxyribonucleases; D003850:Deoxyribonuclease I; D007930:Leucine