Sequence requirements for upregulated expression of Drosophila hsp70 transgenes during aging.
hsp70 protein and hsp70:lacZ fusion reporters are upregulated during aging and in response to oxidative stress in the thorax of Drosophila. hsp70 expression was increased during aging in each of seven different Drosophila genetic backgrounds tested, 2.6-4.8-fold. DNA sequence requirements were investigated by analysis of nine distinct hsp70:lacZ fusion reporter constructs in multiple independent transgenic lines. hsp70 sequences -194 to +276 supported an average 2.7-fold increase during aging. This increase was reduced or eliminated by deletion or point mutation of the heat shock response elements, consistent with a transcriptional mechanism. Similar sequence requirements were observed for increased expression in response to catalase null mutation as a model of oxidative stress. hsp70 5'UTR sequences were required for efficient basal expression of transgenes, but were not sufficient to confer detectable upregulation during aging. Inclusion of additional hsp70 coding region sequences from +276 to + 1011 created a larger hsp70:lacZ fusion protein and had two effects: dramatic reduction of the overall expression level of the fusion protein, and an additional three to fourfold upregulation during aging. These results suggest that the coding region sequences reduce fusion protein abundance and that this negative effect decreases as a function of age. The data support a model for increased expression of hsp70 transgenes during aging involving both transcriptional and posttranscriptional components.
Wheeler
J C
JC
Department of Biological Sciences, University of Southern California, Los Angeles 90089-1340, USA.
King
V
V
Tower
J
J
D000375:Aging / Q000235:genetics*; D000818:Animals; D030801:Animals, Genetically Modified; D015152:Blotting, Northern; D002374:Catalase / Q000235:genetics; D004331:Drosophila melanogaster / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology*; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007763:Lac Operon; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress / Q000502:physiology; D010957:Plasmids; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000235:genetics; D015533:Transcriptional Activation / Q000187:drug effects / Q000235:genetics; D019076:Transgenes / Q000502:physiology; D001616:beta-Galactosidase / Q000235:genetics
D018840:HSP70 Heat-Shock Proteins; D016877:Oxidants; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D006861:Hydrogen Peroxide; D002374:Catalase; D001616:beta-Galactosidase