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Role of 4-HNE metabolism in aging (Drosophila) (R01 grant) Role of glutathione transferases in life span extension of C. elegans Role of 4-HNE metabolism.

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Presentation on theme: "Role of 4-HNE metabolism in aging (Drosophila) (R01 grant) Role of glutathione transferases in life span extension of C. elegans Role of 4-HNE metabolism."— Presentation transcript:

1 Role of 4-HNE metabolism in aging (Drosophila) (R01 grant) Role of glutathione transferases in life span extension of C. elegans Role of 4-HNE metabolism in aging (Drosophila and C. elegans) (Project 2 of P01) Role of glutathione transferases in life span extension of D. melanogaster

2 Lifespan and stress resistance of Caenorhabditis elegans are increased by expression of glutathione transferases capable of metabolizing the lipid peroxidation product 4-hydroxynonenal Ayyadevara, S., Engle, M.R., Singh, S.P., Dandapat, A., Lichti, C.F., Beneš, H., Shmookler Reis, R.J., Liebau, E. and Zimniak, P. (Aging Cell 4: , 2005) Lifespan extension in hypomorphic daf-2 mutants of Caenorhabditis elegans is partially mediated by glutathione transferase CeGSTP2-2 Ayyadevara, S., Dandapat, A., Singh, S.P., Beneš, H., Zimniak, L., Shmookler Reis, R.J. and Zimniak, P. (Aging Cell 4: , 2005) Life span and stress resistance of Caenorhabditis elegans are differentially affected by glutathione transferases metabolizing 4­ hydroxynon-2-enal Ayyadevara, S., Dandapat, A., Singh, S.P., Siegel, E.R., Shmookler Reis, R.J., Zimniak, L. and Zimniak, P. (Mech. Ageing Dev., 2006, in press)

3 Hypothesis: adductscrosslinksmutations aging GST conjugate mGSTA4-4 hGSTA4-4 CeGSTP2-2 DmGSTS1-1 DmGSTD1-1

4 Reactive aldehydes can modify proteins

5 4-HNE conjugation (and the responsible GST) correlate with life span

6 To establish whether 4-HNE-conjugating activity and/or the responsible GSTs only CORRELATE with life span, or whether there is a CAUSE-EFFECT relationship, the levels of 4-HNE-conjugating GSTs were modulated by RNAi knockdown or by transgenic expression.

7 RNAi for CeGSTP2-2 (gst-10 gene product) is effective

8 RNAi against gst-10 but not against gst-8 shortens life span (gst-10 encodes CeGSTP2-2)

9 Expression of mGSTA4-4 or CeGSTP2-2 in transgenic worms

10 Overexpression of mGSTA4-4 and CeGSTP2-2 extends life span

11 Effect of mGSTA4-4 or CeGSTP2-2 on life span

12 Gompertz function

13 Compiled data for 4-HNE-conjugating activity, 4-HNE adducts, and life span (stress resistance shows similar behavior)

14 Effect of daf-2 on CeGSTP2-2 is dependent on daf-16

15 CeGSTP2-2 is localized to a limited set of cells (amphids/phasmids?)

16 Functions of amphids and phasmids David M. Tobin, Cornelia I. Bargmann, Invertebrate Nociception: Behaviors, Neurons and Molecules. J Neurobiol 61: 161–174, Alcedo, J. and Kenyon, C. (2004). Regulation of C. elegans longevity by specific gustatory and olfactory neurons. Neuron 41:

17 Possible mechanism by which CeGSTP2-2 extends life span

18 Do any of C. elegans GSTs other than CeGSTP2-2 affect aging?

19 5 GSTs have 4-HNE-conjugating activity (by RNAi screen) The above 5 GSTs may be responsible for most (or all) of 4-HNE conjugation

20 RNAi appears to be specific Based on sequence analysis (for gst-10) and Western blot (effect of a series of RNAi on gst-10)

21 4-HNE stress resistance: gst-6 and gst-24 are most effective

22 4-HNE-conjugating activity correlates well with 4-HNE stress resistance (10 mM 4-HNE)

23 C. elegans cuticle shields against 4-HNE. Therefore, 10 mM 4-HNE is NOT excessive - in fact, it is equivalent to 10 micromolar 4-HNE in cultured cells

24 4-HNE adducts change with age in C. elegans

25 RNAi targeted to gst-5 and gst-10 shortens life span RNAi targeted to gst-6, gst-8, and gst-24 does not shorten life span

26 Correlation between life span and 4-HNE- conjugating activity is poor: gst-6 and gst-24 gene products have the highest enzymatic activity gst-5 and gst-10 gene products exert the largest effect on life span Possible explanations: 1.4-HNE is not important 2.4-HNE AND localization are important

27 Expression of CeGSTP2-2 is limited to a small number of cells Where is gst-5 localized? gst-6 and gst-24?

28 Unrooted radial phylogenetic tree of C. elegans GSTs

29 D. melanogaster system:  Disruption of the GstS1 gene (encoding a GST responsible for most of 4-HNE-conjugating activity in the fly) leads to a moderate shortening of life span  Transgenic overexpression of the murine mGSTA4-4 (which has high catalytic efficiency for 4-HNE) in some but not all tissues of the fly leads to life span extension  Overall, the D. melanogaster and C. elegans systems confirm a role of 4-HNE in life span determination in two species that diverged approximately one billion years ago. The D. melanogster data are being prepared for publication


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