Dietary restriction, i.e. the restriction of total calories, is the only effective method for increasing the maximum survival of mammals, and it is believed that the increase in survival is due to an alteration in the aging process(es). Therefore, dietary restriction is a powerful tool for studying the biolobical mechanism underlying the aging process and the impact of nutrition on the aging process. Several years ago, the applicant proposed that dietary restriction was acting at the level of gene expresssion. During the past three years, the applicant's laboratory has shown that dietary restriction alters the age-related change in the levels of a variety of mRNA species in several tissues from rat, and that the alteration in mRNA levels is paralelled by an alteration in nuclear transcription. Thus, he has shown that dietary restriction does indeed alter gene expression and that this alteration occurs primarily at the level of transcription. The purpose of the research described in this proposal is to elucidate the molecular mechanism through which dietary restriction alters the transcription of genes. Specifically the applicant will test the following hypothesis: the changes in transcription that arise from dietary restriction are due to changes in the levels/activities of specific transcription factors that regulate the transcription of specific genes. This hypothesis will be tested by studying the expression of the gene for a heat shock protein, hsp 70, in hepatocytes or lymphocytes isolated from male Fisher F344 rats fed ad libitum or a calorie-restricted diet (60% of diet consumed by rats fed ad libitum). The specific objectives of the research described in this proposal are as follows: (1) To characterize the effect of aging and dietary restriction on the induction of hsp70 expression (synthesis, mRNA levels, and transcription) by heat shock and a heavy metal, and to characterize the expression of hsp70 by individual cells from young and old rats fed ad libitum and the restricted-diet by in situ hybridization; (2) to characterize the effect of aging and dietary restriction on the initiation of hsp70 transcription by the in vitro and in vivo transcription of hsp70 promoter-reporter templates; and (3) to characterize the effect of aging and dietary restriction on transcription factors that regulate hsp70 expression. The activation (DNA binding) and expression (i.e., protein and mRNA levels) of heat shock transcription factor as well as CAAT-box transcription factor and Sp1 will be determined in young and old rats on the two diets.
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