The tumour promoter okadaic acid inhibits reticulocyte-lysate protein synthesis by increasing the net phosphorylation of elongation factor 2.
Okadaic acid, a tumour promoter which potently inhibits protein phosphatases, inhibited translation in the reticulocyte-lysate cell-free system. Inhibition was dose-dependent, with half-maximal effects occurring at 20-40 nM-okadaic acid. Inhibition of translation by okadaic acid resulted in the accumulation of polyribosomes, indicating that it was due to a decrease in the rate of elongation relative to initiation. Okadaic acid (at concentrations which inhibited translation) caused increased phosphorylation of a number of proteins in the lysate. Prominent among these was a protein of Mr 100,000, which has previously been identified as elongation factor 2 (EF-2). EF-2 is a specific substrate for a Ca2+/calmodulin-dependent protein kinase, which phosphorylates EF-2 on threonine residues. The Mr-100,000 band was phosphorylated exclusively on threonine residues, and its degree of 32P labelling was decreased by the Ca2+ chelator EGTA and by the calmodulin antagonist trifluoperazine. These agents attenuated the effects of okadaic acid on EF-2 phosphorylation and translation. When ranges of concentrations of each agent were tested, their effects on EF-2 labelling correlated well with their ability to reverse the okadaic acid-induced inhibition of translation. These findings demonstrate that increased phosphorylation of EF-2 results in an impairment of peptide-chain elongation when natural mRNA is used. The possible physiological role of EF-2 phosphorylation in the control of translation is discussed.
Redpath
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NT
Department of Biochemistry, School of Medical Sciences, University of Bristol, U.K.
Proud
C G
CG
D001798:Blood Proteins / Q000096:biosynthesis*; D002474:Cell-Free System; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000494:pharmacology; D004988:Ethers, Cyclic / Q000494:pharmacology*; D006801:Humans; D007476:Ionophores / Q000494:pharmacology*; D008970:Molecular Weight; D019319:Okadaic Acid; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors / Q000378:metabolism*; D010766:Phosphorylation; D011132:Polyribosomes; D014176:Protein Biosynthesis / Q000187:drug effects; D014268:Trifluoperazine / Q000494:pharmacology
D001798:Blood Proteins; D004988:Ethers, Cyclic; D007476:Ionophores; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors; D019319:Okadaic Acid; D014268:Trifluoperazine; D004533:Egtazic Acid