Details

Autor(en) / Beteiligte

• Chen, Jia
• Zhao, Xiao-Nan
• Yang, Li
• Hu, Guang-Jing
• Lu, Ming
• Xiong, Ying
• Yang, Xin-Ying
• Chang, Catherine C Y
• Song, Bao-Liang
• Chang, Ta-Yuan
• Li, Bo-Liang

Titel

RNA secondary structures located in the interchromosomal region of human ACAT1 chimeric mRNA are required to produce the 56-kDa isoform

Ist Teil von

• Cell research, 2008-09, Vol.18 (9), p.921-936

Ort / Verlag

England: Nature Publishing Group

Erscheinungsjahr

2008

Link zum Volltext

Quelle

MEDLINE

Beschreibungen/Notizen

• We have previously reported that the human ACAT1 gene produces a chimeric mRNA through the interchromosomal processing of two discontinuous RNAs transcribed from chromosomes 1 and 7. The chimeric mRNA uses AUG1397-1399 and GGC1274-1276 as translation initiation codons to produce normal 50-kDa ACAT1 and a novel enzymatically active 56-kDa isoform, respectively, with the latter being authentically present in human cells, including human monocyte- derived macrophages. In this work, we report that RNA secondary structures located in the vicinity of the GGC1274-1276 codon are required for production of the 56-kDa isoform. The effects of the three predicted stem-loops （nt 1255-1268, 1286-1342 and 1355-1384） were tested individually by transfecting expression plasmids into cells that contained the wild-type, deleted or mutant stem-loop sequences linked to a partial ACAT1 AUG open reading frame （ORF） or to the ORFs of other genes. The expression patterns were monitored by western blot analyses. We found that the upstream stem-loop1255-1268 from chromosome 7 and downstream stem-loop1286-1342 from chromosome 1 were needed for production of the 56-kDa isoform, whereas the last stem-loop135s-1384 from chromosome 1 was dispensable. The results of experi- ments using both monocistronic and bicistronic vectors with a stable hairpin showed that translation initiation from the GGC1274-1276 codon was mediated by an internal ribosome entry site （IRES）. Further experiments revealed that translation initiation from the GGC1274-1276 codon requires the upstream AU-constituted RNA secondary structure and the downstream GC-rich structure. This mechanistic work provides further support for the biological significance of the chimeric nature of the human ACAT1 transcript.