Cryptic promoter activity in the DNA sequence corresponding to the pim-1 5′-UTR

Abstract

The serine/threonine kinase pim-1 mRNA contains a long and G/C rich 5′-untranslated region (5′-UTR). Previous work suggested that the pim-1 5′-UTR harbors an internal ribosomal entry site (IRES) allowing for internal initiation of translation. However, several previously reported eukaryotic IRES elements actually contain cryptic promoter activity. To test whether an IRES or a cryptic promoter is present in the pim-1 5′-UTR, the 5′-UTR was re-examined using stringent test procedures. Our results show the presence of strong promoter activity in the DNA sequence corresponding to the pim-1 5′-UTR. Both promoterless dicistronic test and northern blot analysis show transcripts being derived from the cryptic promoter in the pim-1 5′-UTR sequence. This cryptic promoter is active in all cell types tested, including Cos-7, NIH3T3, HEK293, Jurkat and K562 cells. When a dicistronic mRNA containing the pim-1 5′-UTR was translated in vitro or in vivo, no IRES activity could be detected. However, the control IRESs from both human rhinovirus and encephalomyocarditis virus exhibited strong IRES activities. In addition, both the RNase protection assay and the 5′-RACE assay detected endogenous pim-1 transcripts with shorter 5′-UTRs. Our data strongly suggest that the IRES activity reported earlier for the pim-1 5′-UTR sequence is due to cryptic promoter activity. © The Author 2005. Published by Oxford University Press. All rights reserved.