Inactivation of mRNA cap-binding protein complex in Drosophila melanogaster embryos under heat shock

J.M Zapata,F.G Maroto,J.M Sierra

doi:10.1016/s0021-9258(18)98508-x

J.M Zapata, F.G Maroto + Show 1 more

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https://doi.org/10.1016/s0021-9258(18)98508-x

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Abstract

We have studied the role of Drosophila 35-kDa cap-binding protein (CBP) and CBP complex in the mechanism of messenger RNA discrimination established in heat-shocked Drosophila embryos. Drosophila 35-kDa CBP is functionally equivalent to the mammalian eucaryotic initiation factor (eIF)-4E and CBP complex, which includes eIF-4E, might be the counterpart of mammalian eIF-4F. By using anti-eIF-4E antibodies, we found that although translation of the bulk of normal messengers in Drosophila lysates was very dependent on eIF-4E, the mRNAs for the heat shock proteins (hsps) (particularly hsp70 mRNA and with the exception of hsp83 mRNA) were translated almost independently of this factor, suggesting that they may have unstructured leaders. Accordingly, hsp70 mRNA and, to a lesser extent, the mRNAs for the small hsps were found to be more resistant to inhibition by K+ than normal and hsp83 mRNAs. Moreover, Drosophila CBP complex was able to rescue partial but specifically the synthesis of normal proteins when added to a lysate from heat-shocked embryos. However, no significant effect was obtained by Drosophila eIF-4E or eIF-2. Consistent with these results, we found a great decrease in the amount of the CBP complex purified from heat-shocked embryos as compared with normal ones, whereas the amounts of free eIF-4E purified from either source were similar. Together, the above results suggest that some modification leading to the disruption of Drosophila CBP complex may account, at least to some extent, for the mRNA discrimination established in heat-shocked Drosophila embryos.

Highlights

Thepreferentialtranslation of theheat shock mRNAs independently of this factor, suggesting that themyay during heat shock appears to be due both to the occurrence have unstructured leadersA. ccordingly, hsp70 mRNA of a specific structural featurevery close to the 5’-enodf their and, to a lesser extent, themRNAs for the small hsps leader sequences [6,7,8], as well as to a change in some comwere found to be more resistant to inhibition by K+ ponent(s) of the translational machinerywhich has not been than normal and hspm83RNAs
Additional experimentsappear to be required to support this hypothesis further as it was mainly based on the observation that, in contrasttothe normal messengers, translation of the heat shock mRNAs, hsp70 mRNA, was resistant toinhibition by cap analogs
(lanes 1-4), we found that theaddition of cap-binding proteins (CBPs) complex to a normal lysate supplemented withglobin mRNA did not result in theincrease of any of the polypeptides synthesized by these lysates including globin (Fig. 5C,lanes 5-8)

Summary

Introduction

Thepreferentialtranslation of theheat shock mRNAs independently of this factor, suggesting that themyay during heat shock appears to be due both to the occurrence have unstructured leadersA. ccordingly, hsp mRNA of a specific structural featurevery close to the 5’-enodf their and, to a lesser extent, themRNAs for the small hsps leader sequences [6,7,8], as well as to a change in some comwere found to be more resistant to inhibition by K+ ponent(s) of the translational machinerywhich has not been than normal and hspm83RNAs. CBP complex was able to rescpuaertial but between the phosphorylation state of the ribosomal protein the synthesis of normal proteinswhen added taolysate S6 with the levels of normalproteinsynthesisduringthe from heat-shocked embryos. Heat-shocked embryos as compared with normal ones, Sanders et al [14] suggested that the regulatory whereastheamounts of freeeIF-4Epurifiedfrom factors are present in the post-ribosomal supernatant fraction. The above resulRtsecent results from our laboratory [9] suggested that in suggest that some modification leading to the disruption of Drosophila CBP complex may account, at least to some extent, for the mRNA discrimination established in heat-shockedDrosophila embryos

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