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1.
Selective degradation of mRNA: the role of short-lived proteins in differential destabilization of insulin-induced creatine phosphokinase and myosin heavy chain mRNAs during rat skeletal muscle L6 cell differentiation. 总被引:8,自引:1,他引:7
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This investigation concerns the combined effects of removal and readdition of insulin and inhibition of protein and RNA synthesis on the stability of insulin-induced mRNAs during and after differentiation of rat L6A1 myoblast cells in culture. Addition of insulin accompanying the withdrawal of the mitogenic stimulus of serum to myoblasts caused an 80-fold increase in creatine phosphokinase (CK) activity which was largely accounted for by a similar increase in the amount of CK mRNA. The latter was co-ordinately induced with myosin heavy chain (MHC) mRNA but not malic enzyme (ME) mRNA. Measurements of steady-state levels of mRNA showed that removal of insulin caused CK mRNA, but not MHC mRNA, to be rapidly degraded, the effect being reversed upon readdition of the hormone. Direct measurement of 3H-labeled CK, MHC and beta-actin mRNAs confirmed the selective stabilization and destabilization of CK mRNA by the hormone. Conditions were established for a time-window during which cycloheximide (Cx) produced a virtually total arrest of protein synthesis in myotubes that was reversible upon removal of the inhibitor. Under these conditions, Cx selectively prevented the degradation of CK mRNA in a reversible manner. Actinomycin D (Act D) also arrested the loss of this mRNA. Under the same conditions of mRNA stabilization during de-induction, a superinduction of CK mRNA, but not MHC mRNA, was observed if the two inhibitors were added during induction in the continuous presence of insulin. We conclude that a short-lived protein(s), encoded by a short-lived mRNA(s), selectively regulates the stability of reversibly inducible mRNA. 相似文献
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The formation, distribution and function of ribosomes and microsomal membranes during induced amphibian metamorphosis 总被引:13,自引:10,他引:3
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J. R. Tata 《The Biochemical journal》1967,105(2):783-801
1. A lag period of about 4 days preceded the onset of metamorphosis precociously induced by tri-iodothyronine in tadpoles of the giant American bullfrog (Rana catesbeiana). It was established by the accelerated synthesis or induction of carbamoyl phosphate synthetase and cytochrome oxidase in the liver, serum albumin and adult haemoglobin in the blood, acid phosphatase in the tail, and the increase in the hindleg/tail length ratio. 2. A 4- to 6-fold stimulation, 2 days after the induction of metamorphosis, of the rate of synthesis of rapidly labelled nuclear RNA in liver cells was followed by an increasing amount of RNA appearing in the cytoplasm. Most of the newly formed RNA on induction of metamorphosis was of the ribosomal type. An accelerated turnover at early stages of development preceded a net accumulation of RNA in the cytoplasm, with no change in the amount of DNA per liver. 3. Most hepatic ribosomes of the pre-metamorphic tadpoles were present as 78s monomers and 100s dimers; metamorphosis caused a shift towards larger polysomal aggregates with newly formed ribosomes that were relatively more tightly bound to membranes of the endoplasmic reticulum. 4. The appearance of new polyribosomes in the cytoplasm on induction of metamorphosis was co-ordinated in time with a stimulation of synthesis of phospholipids of the smooth and rough endoplasmic reticulum, followed by a gradual shift in preponderance from the smooth to the rough type of microsomal membranes. 5. Electron- and optical-microscopic examination of intact hepatocytes revealed a striking change in the distribution and nature of ribosomes and microsomal membranes during metamorphosis. 6. Ribosomes prepared from non-metamorphosing and metamorphosing animals were identical in their sedimentation coefficients and in the structural ribosomal proteins. The base composition and sedimentation coefficients of ribosomal RNA were also identical. Induction of metamorphosis also did not alter the incorporation of (32)P into the different phospholipid constituents of microsomal membranes. 7. Nascent (14)C-labelled protein with the highest specific activity was recovered in the ;heavy' rough membrane fraction of microsomes, whereas little (14)C was associated with ;free' polysomes. Protein synthesis in vivo was most markedly stimulated during metamorphosis in the tightly membrane-bound ribosomal fraction after the appearance of new ribosomes. 8. The rate of synthesis of macromolecules in vivo could not be followed beyond 7-8 days after induction because of variable shifts in precursor pools due to regression of larval tissues. 9. The stimulation of RNA and ribosome formation was specifically associated with the process of metamorphosis since no similar response to thyroid hormones occurred in those species (Axolotl and Necturus) in which the hormones failed to induce metamorphosis. 相似文献
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The effects of the tumor promoter phorbol 12-myristate 13-acetate (PMA) on the proliferation, protein kinase C activity (PKC), and c-fos gene expression were examined in cultures of young and senescent (90-95% lifespan completed) WI-38 human diploid fibroblasts. We observed that, following stimulation with medium containing 10% fetal bovine serum (FBS), the translocation of PKC from the cytosol to the particulate compartment was less efficient in senescent WI-38 cells than in young cells. However, when PMA was added to the medium, the intracellular distribution of PKC activity in old cells became nearly identical to that observed in young cells. The inducibility of c-fos mRNA by serum addition, which is a protein kinase C-dependent event [64], was significantly amplified in the presence of PMA. Moreover, the duration of peak c-fos expression, after stimulation by FBS and PMA, increased in senescent cells as compared to young cells. Our results reveal that the normal signal transduction pathway is altered in senescent, slowly proliferating human fibroblasts and that it can be partially restored in the presence of the tumor promoter PMA. 相似文献
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