Pitfalls of the CAT reporter gene for analyzing translational regulation in Leishmania.
Heterologous reporter genes are widely used for the characterization of gene expression in many organisms. Particularly, constructs bearing reporter genes have greatly contributed to our understanding of gene regulation in kinetoplastids. In some specific circumstances, however, such heterologous reporter has a risk of resulting in irrelevant observations and conclusions, which are primarily due to the introduction of foreign sequence elements. This communication describes our recent experience using the chloramphenicol acetyltransferase (CAT) gene as a reporter for analysis of the translational regulation of HSP70 genes in Leishmania infantum. We show that chimeric mRNAs consisting of the CAT open reading frame (ORF) and the untranslated regions (UTRs) from HSP70-II genes behave differently as endogenous HSP70-II mRNAs and that this difference is due to the presence of CAT sequences. Thus, the main purpose of this communication is to alert researchers working in gene regulation to be cautious when interpreting results based on heterologous reporter genes.
Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid, 28049, Madrid, Spain.
Cristina
C
Folgueira
Jose M
JM
Requena
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis* / Q000235:genetics; D005786:Gene Expression Regulation / Q000502:physiology*; D005798:Genes, Bacterial; D017930:Genes, Reporter; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D018314:Leishmania infantum / Q000235:genetics / Q000502:physiology*; D008967:Molecular Biology / Q000379:methods*; D014176:Protein Biosynthesis / Q000502:physiology*; D015800:Protozoan Proteins / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D016053:RNA, Protozoan / Q000378:metabolism
D018840:HSP70 Heat-Shock Proteins; D015800:Protozoan Proteins; D012333:RNA, Messenger; D016053:RNA, Protozoan; D015500:Chloramphenicol O-Acetyltransferase