Figure 2

The regulation of the HSP83 gene expression occurs at the post-transcriptional level. (A) Promastigotes of L. infantum were grown at 26°C, and parallel cultures were incubated at 37°C during 10, 30, and 60 min. Run-on transcripts, labeled with [³²P]UTP, were hybridized to slot blots containing 5 μg of linearized plasmid DNAs. The following plasmids were used: pBLs, pBluescript; rRNA, pRIB (plasmid containing a 590-bp fragment of the L. infantum 24S α rDNA gene [9]); HSP83, pE2 (plasmid containing a fragment of the 3'-UTR of L. infantum HSP83 gene). The values of the hybridization intensity were measured by densitometric analysis of the autoradiographs, and the ratio HSP83/rRNA is shown. Data represent the average of two separate experiments. (B) RNA samples were prepared from promastigotes cultures grown at either 26°C or 37°C for 0 h (lanes 1), 1 h (lanes 2), 2 h (lanes 3) and 4 h (lanes 4) in the presence (CH+) or absence (CH-) of cycloheximide. The drug was added 1 h prior to start the temperature treatment. Lanes C contain RNA samples prepared from parasite cultures before starting the drug treatment. The Northern blots were hybridized with a specific probe for the L. infantum HSP83 3'-UTR.