<title>ABSTRACT</title>In gene-trap screening of plant genomes, promoterless reporter constructs are often expressed without trapping of annotated gene promoters. The molecular basis of this phenomenon, which has been interpreted as the trapping of cryptic promoters, is poorly understood. In this study, using <italic>Arabidopsis</italic> gene-trap lines in which a firefly luciferase (<italic>LUC</italic>) open reading frame (ORF) was expressed from intergenic regions, we found that cryptic promoter activation occurs by at least two different mechanisms: one is the capturing of pre-existing promoter-like chromatin marked by H3K4me3 and H2A.Z, and the other is the entirely new formation of promoter chromatin near the 5’ end of the inserted <italic>LUC</italic> ORF. To discriminate between these, we denoted the former mechanism as “cryptic promoter capturing”, and the latter one as “promoter <italic>de novo</italic> origination”. The latter finding raises a question as to how inserted <italic>LUC</italic> ORF sequence is involved in this phenomenon. To examine this, we performed a model experiment with chimeric <italic>LUC</italic> genes in transgenic plants. Using <italic>Arabidopsis psaH1</italic> promoter–<italic>LUC</italic> constructs, we found that the functional core promoter region, where transcription start sites (TSS) occur, cannot simply be determined by the upstream nor core promoter sequences; rather, its positioning proximal to the inserted <italic>LUC</italic> ORF sequence was more critical. This result suggests that the insertion of the <italic>LUC</italic> ORF sequence alters the local distribution of the TSS in the plant genome. The possible impact of the two types of cryptic promoter activation mechanisms on plant genome evolution and endosymbiotic gene transfer is discussed.