Elsevier

Experimental Gerontology

Volume 41, Issue 10, October 2006, Pages 910-921
Experimental Gerontology

The search for DAF-16/FOXO transcriptional targets: Approaches and discoveries

https://doi.org/10.1016/j.exger.2006.06.040Get rights and content

Abstract

The insulin/IGF-1 receptor (IIR)/FOXO pathway is remarkably conserved in worms, flies, and mammals, and downregulation of signaling in this pathway has been shown to extend lifespan in all of these animals. FOXO-mediated transcription is required for the long lifespan of IIR mutants; thus, there is great interest in identifying FOXO target genes, as they may carry out the biochemical activities that extend longevity. A number of approaches have been used to identify the transcriptional targets of FOXO. Thus far, the best data available on the components downstream of this pathway are from experiments involving the Caenorhabditis elegans FOXO transcription factor, DAF-16; some of these targets have been tested for their contributions to longevity, dauer formation, and fat storage. Here, I examine and compare the approaches used to identify DAF-16/FOXO targets, review the genes regulated by DAF-16, and discuss the processes that may be at work to extend lifespan in IIR mutants. Rather than upregulating every possible beneficial gene, DAF-16 appears to selectively upregulate genes that contribute to specific protective mechanisms, while simultaneously downregulating potentially deleterious genes. In addition to genes that carry out expected roles in stress protection, many previously unknown targets have been identified in these studies, suggesting that some mechanisms of lifespan extension still await discovery. These mechanisms may act cooperatively or cumulatively to increase longevity, and are likely to be at least partially conserved in higher organisms.

Introduction

The Caenorhabditis elegans FOXO transcription factor, DAF-16, is controlled by the activity of the DAF-2 insulin receptor, repressing DAF-16 activity through phosphorylation and cytoplasmic retention (Lin et al., 2001, Henderson and Johnson, 2001). In the absence of DAF-2/insulin receptor signaling, DAF-16/FOXO moves into the nucleus and regulates transcription of its targets. The identification of these target genes has been the focus of much attention because they may mediate the various phenotypes of daf-2 mutants, including longevity determination (Kenyon et al., 1993), fat metabolism (Lee et al., 2003, Ashrafi et al., 2003), and formation of the diapause state known as dauer (Riddle and Albert, 1997). The genes making up the DAF-16-regulated transcriptome are likely to carry out the biochemical activities necessary for these phenotypes. The roles that DAF-16/FOXO performs in C. elegans may be broadly conserved, as its homologs in flies (dFOXO) and mammalian cells (FOXO3a, FOXO1) are also critical for lifespan (Hwangbo et al., 2004) and cell survival (Brunet et al., 2004). Elimination of the insulin receptor specifically from adipose tissue (the Fat Insulin Receptor KnockOut, or FIRKO, mouse) results in increased lifespan (Bluher et al., 2003), highlighting the importance of the longevity-determining genes regulated by FOXOs in mammals. The identification of FOXO target genes in all of these organisms will help unravel the mechanisms underlying longevity and cell survival. Once the roles of individual genes are elucidated, some may serve as targets for clinical intervention, both for normal aging and for age-related disease.

Section snippets

Approaches used to identify DAF-16 targets

Of all the FOXO transcription factors, most is known about the targets of the C. elegans homolog, DAF-16. Identifying DAF-16 targets through classical genetics is challenging: an EMS screen for suppressors of the dauer-constitutive phenotype of daf-2 in the presence of a daf-16 transgene yielded many alleles of daf-18 (the PTEN phosphatase upstream of daf-16) rather than genes downstream of daf-16 (Lin et al., 2001). This is perhaps less surprising in light of later evidence that the functions

Why might the DAF-16 target sets vary?

It is fair to ask at this point how much overlap exists between the target sets produced by these studies, and if there are differences, what are the sources of this variation? In fact, many of the significantly changed genes have been identified in multiple studies (Murphy et al., 2003, McElwee et al., 2003, McElwee et al., 2004, Halaschek-Wiener et al., 2005), so it is likely that most of the approaches have resulted in valuable data. However, both biological factors and technical

DAF-16 target genes: clues to underlying biological mechanisms

Now that several approaches have identified DAF-16 targets, we can begin to think about how they contribute to daf-16-mediated phenotypes. Many of the DAF-16 targets revealed by genome-wide approaches were expected, as they had already been studied in candidate approaches. Additionally, many are shared with the dauer transcriptome (McElwee et al., 2004, Halaschek-Wiener et al., 2005, Wang and Kim, 2003); this is not surprising, given that daf-2 mutants form dauers constitutively at high

Conclusions

What has been learned by amassing this collection of genes regulated by daf-2 and daf-16? While many of the targets confirm previous hypotheses (protection from a variety of stresses, overlap with dauer genes), many of the biological functions of FOXO/DAF-16 targets remain undiscovered. The vast majority of the genes have not yet been tested for their contributions to daf-2 phenotypes; until those genes are analyzed, it seems premature to claim that the mechanisms by which insulin receptor

Acknowledgements

The author thanks Zemer Gitai, Wendy Shaw, Shijing Luo, and Max Jan for useful comments. Due to space limitations, many relevant studies could not be addressed in this review, and the author apologizes for these omissions.

References (66)

  • S. Ramaswamy

    A novel mechanism of gene regulation and tumor suppression by the transcription factor FKHR

    Cancer Cell

    (2002)
  • S. Rea et al.

    A metabolic model for life span determination in Caenorhabditis elegans

    Dev. Cell

    (2003)
  • J. van Helden et al.

    Extracting regulatory sites from the upstream region of yeast genes by computational analysis of oligonucleotide frequencies

    J. Mol. Biol.

    (1998)
  • S. Wolff

    SMK-1, an essential regulator of DAF-16-mediated longevity

    Cell

    (2006)
  • H. Yu et al.

    DAF-16-dependent and independent expression targets of DAF-2 insulin receptor-like pathway in Caenorhabditis elegans include FKBPs

    J. Mol. Biol.

    (2001)
  • K. Ashrafi

    Genome-wide RNAi analysis of Caenorhabditis elegans fat regulatory genes

    Nature

    (2003)
  • D. Barsyte et al.

    Longevity and heavy metal resistance in daf-2 and age-1 long-lived mutants of Caenorhabditis elegans

    FASEB J.

    (2001)
  • M. Bluher et al.

    Extended longevity in mice lacking the insulin receptor in adipose tissue

    Science

    (2003)
  • A. Brunet

    Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase

    Science

    (2004)
  • H.J. Bussemaker et al.

    Building a dictionary for genomes: identification of presumptive regulatory sites by statistical analysis

    Proc. Natl. Acad. Sci. USA

    (2000)
  • C.e.S. Consortium

    Genome sequence of the nematode C. elegans: a platform for investigating biology

    Science

    (1998)
  • A. Dillin et al.

    Timing requirements for insulin/IGF-1 signaling in C. elegans

    Science

    (2002)
  • M.B. Eisen

    Cluster analysis and display of genome-wide expression patterns

    Proc. Natl. Acad. Sci. USA

    (1998)
  • M.A. Essers

    Functional interaction between beta-catenin and FOXO in oxidative stress signaling

    Science

    (2005)
  • A.G. Fraser

    Functional genomic analysis of C. elegans chromosome I by systematic RNA interference

    Nature

    (2000)
  • T. Furuyama

    Identification of the differential distribution patterns of mRNAs and consensus binding sequences for mouse DAF-16 homologues

    Biochem. J.

    (2000)
  • T.R. Golden et al.

    Microarray analysis of gene expression with age in individual nematodes

    Aging Cell

    (2004)
  • E.L. Greer et al.

    FOXO transcription factors at the interface between longevity and tumor suppression

    Oncogene

    (2005)
  • J. Halaschek-Wiener

    Analysis of long-lived C. elegans daf-2 mutants using serial analysis of gene expression

    Genome. Res.

    (2005)
  • B. Hamilton

    A systematic RNAi screen for longevity genes in C. elegans

    Genes Dev.

    (2005)
  • M. Hansen

    New genes tied to endocrine, metabolic, and dietary regulation of lifespan from a Caenorhabditis elegans genomic RNAi screen

    PLoS Genet.

    (2005)
  • Y. Honda et al.

    The daf-2 gene network for longevity regulates oxidative stress resistance and Mn-superoxide dismutase gene expression in Caenorhabditis elegans

    FASEB J.

    (1999)
  • A.L. Hsu et al.

    Regulation of aging and age-related disease by DAF-16 and heat-shock factor

    Science

    (2003)
  • Cited by (0)

    View full text