The role of human sodium dicarboxylate cotransporter in oxidative stress

by Cheung, Kwok-ho

(Uncorrected OCR) Abstract of the thesis entitled

THE ROLE OF HUMAN SODIUM DICARBOXYLATE COTRANSPORTER IN OXIDATIVE STRESS

Submitted by

Alvin, Kwok Ho Cheung

For the degree of Master of philosophy at the University of Hong Kong In August, 2003

Ageing is a genetically controlled process that can be modulated in the

invertebrates by single gene mutations. An insertional mutation of Indy (I'm not dead

yet) gene in Drosophila melanogaster resulted in a 50% increase in their maximal

life-span. Here we show that an increased uptake of Krebs cycle intermediates in

Chinese Hamster Ovary (CHO) cells which have a high endogenous expression of

Sodium Dicarboxylate Cotransporter-l (NaDC-l), a homolog of Indy gene In

eukaryotes, resulted in the increase in oxidative stress.

A respiratory chain inhibitor, rotenone reduced both glucose- and

succinate-induced oxidative stress, indicating that reactive oxygen free radicals

derived from these two substrates are generated from the mitochondrial electron

transport chain. Lithium ion, which is known to be able to reduce the uptake of Krebs

cycle intermediates, only reduced succinate-induced oxidative stress but not

glucose-induced one. This suggests that the increased uptake of Krebs cycle

intermediates lead to the Increase In oxidative stress. This was confirmed by

over-expressing human NaDC-I in EcR (Ecdysone receptor) 293 cells, that has a

very low level of endogenous NaDC-I expression. EcR 293 cells over-expressing

hNaDC-I showed a higher level of succinate-induced oxidative stress. We propose that blocking the uptake activity ofNaDC-1 can reduce oxidative stress. Since Lt is

able to interfere with other sodium-dependent transporters, we set off to screen for

inhibitors specific for hNaDC-I. We identified a molecule called compound X that

reversibly inhibits dicarboxylate uptake by hNaDC-I and reduces succinate-induced

oxidative stress.

Our present studies indicate that an increased uptake of Krebs cycle

intermediates increases oxidative stress, and the lowering of the transport activity of

N aDC-I reduces the oxidative stress. This may be at least one molecular mechanism

involved in the life-span prolonging effect of Indy mutations in Drosophila. Thus,

inhibitors of NaDC-I may be a pharmacological agent to extend the life-span in

mammals.

(304 words)