Comparative transcriptional profiling identifies takeout as a gene that regulates life span

Abstract

A major challenge in translating the positive effects of dietary restriction (DR) for the improvement of human health is the development of therapeutic mimics. One approach to finding DR mimics is based upon identification of the proximal effectors of DR life span extension. Whole genome profiling of DR in Drosophila shows a large number of changes in gene expression, making it difficult to establish which changes are involved in life span determination as opposed to other unrelated physiological changes. We used comparative whole genome expression profiling to discover genes whose change in expression is shared between DR and two molecular genetic life span extending interventions related to DR, increased dSir2 and decreased Dmp53 activity. We find twenty-one genes shared among the three related life span extending interventions. One of these genes, takeout, thought to be involved in circadian rhythms, feeding behavior and juvenile hormone binding is also increased in four other life span extending conditions: Rpd3, Indy, chico and methuselah. We demonstrate takeout is involved in longevity determination by specifically increasing adult takeout expression and extending life span. These studies demonstrate the power of comparative whole genome transcriptional profiling for identifying specific downstream elements of the DR life span extending pathway.

Figure 1.. Comparison of genes upregulated and downregulated in yw/w ¹¹¹⁸ DR, Canton-S CR and dSir2 overexpressed long-lived flies at Day 10.

(A) Venn diagram comparing the upregulated and downregulated genes for DR flies in a yw/w¹¹¹⁸ and a Canton-S background at age 10 days. (B) Venn diagram comparing upregulated and downregulated genes in DR long-lived flies and dSir2 overexpressing long-lived flies at age 10 Days. DR flies and dSir2 overexpressing flies are in an identical genetic background. The majority of genes expressed in dSir2 overexpression are also expressed in DR (72% upregulated and 61% downregulated). Verification of microarray data using qPCR is in Supplementary Figure 3.

Figure 2.. Comparison of genes upregulated and downregulated in DR, dSir2 overexpression and DN-Dmp53 expressing long-lived flies at Day 10.

(A) Venn diagrams comparing upregulated and downregulated genes in DR, dSir2 overexpression, and DN-Dmp53 in a yw/w¹¹¹⁸ background at age 10 Days. Genes intersecting in all 3 sets are noted in box with arrow. (B) Heatmap comparing the average log2 fold changes for genes significantly altered in the yw/w¹¹¹⁸ DR with the equivalent genes in dSir2 and DN-Dmp53 expressing flies.

Figure 3.

takeout mRNA expression is increased in yw/w¹¹¹⁸ DR, Canton-S DR, w¹¹¹⁸ DR, Sir2 overexpression, DN-Dmp53 expression, Indy, Rpd3, methuselah (mth) and chico. Fold change increase by qPCR of takeout mRNA from 10-Day old flies from these twelve life span extending conditions as compared to their genetically or dietary matched controls.

Figure 4.

Overexpression of takeoutin either of three different adult tissues extends life span of males and females.(A) Expression of takeout in the adult nervous system using the ELAV-Switch neuronal specific GAL4 driver leads to ~25% increase in mean longevity. (B) Flies expressing takeout in the head fat body, S1-32 pericerebral fat body specific GAL4 driver, have ~20% extension of mean life span, while takeout expression in the abdominal fat body, S1-106 abdominal fat body specific GAL4 driver, (C) extends fly life span by ~12-18% (females: left panel; males: right panel; statistical analysis in Table 1 and 2; grey: controls; black: takeout).

Figure 5.. Long-lived takeout overexpressing flies have increased expression of a subset of the upregulated genes found in DR, dSir2 overexpression and DN-Dmp53 expressing flies.

Overexpression of takeout in adult neurons increases the expression of a subset of 9 genes from the 19 upregulated genes shared in yw/w¹¹¹⁸ DR, dSir2 overexpression and DN-Dmp53 expressing long-lived flies at Day 10. Fold change increase by qPCR of each of the noted genes using mRNA extracted from 10-Day old flies induced to express takeout (Elav-GeneSwitch-UAS-to) as compared to genetically identical controls Elav-GeneSwitch; UAS-to flies without fed diluent.