The University of Western Ontario

Expression of some maize 18 kDa HSPs result from the translation at internal AUG codons
--J. Roger H. Frappier, Robert A. Bouchard, David B. Walden and Burr G. Atkinson

Initiation of translation of eukaryotic mRNA's occurs typically at the first AUG triplet from the 5' end of the message although exceptions have been described (Suzuki et al., Eur. J. Biochem. 207:767-772, 1992). Herein, we report an exception which appears to be common among mRNA transcripts encoding the 18 kDa HSPs in radicles from heat-shocked maize seedlings. We have isolated, sequenced and characterized cDNAs encoding different members of the 18 kDa HSP family and found that the open reading frames of some transcripts contain in-frame, internal AUG codons. Transcription and translation of 18 kDa HSP cDNAs containing internal AUGs in the transcribed RNA appear to synthesize a polypeptide initiating from each AUG codon. Furthermore, an HSP 18 cDNA was expressed in the Invitrogen pTrcHis expression system containing a 33 amino acid leader sequence, resulting in the production of three proteins (corresponding to the internal HSP AUGs). Moreover, translation of hybrid-selected poly(A)+ mRNAs from radicles or plumules of heat-shocked maize seedlings (utilizing oligonucleotide sequences for hybrid selection which are specific for each cDNA) results in the synthesis of different proteins which correspond to the number of AUGs in the hybrid-selected transcript. Both the deduced and observed molecular mass and isoelectric point of each of the proteins synthesized from the AUG most proximal to the 5' end of the transcripts as well as those synthesized from internal AUGs correspond to a member of the 18 kDa HSP family synthesized in vivo. We suggest that the expression of some members of the maize 18 kDa HSP family results from initiation of translation at internal AUG codons. 

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