Methods for using soy peptides to inhibit H3 acetylation, reduce expression of HMG CoA reductase, and increase LDL receptor and Sp1 expression in a mammal

#This application is related to U.S. application Ser. No. 11/532,526, filed concurrently on Sep. 16, 2006, the contents of which are incorporated by reference herein in its entirety.

#1. Field of the Invention

#This disclosure relates generally to a class of peptides that provide mammals with a variety of health related benefits. More specifically, the present disclosure related to using soy peptides to inhibit H3 acetylation, reduce expression of HMG CoA reductase and increase LDL receptor and Sp1 expression in a mammal.

#The above-mentioned features and objects of the present disclosure will become more apparent with reference to the following description taken in conjunction with the accompanying drawings wherein like reference numerals denote like elements and in which:

#FIG. 1 shows the 2S albumin protein encoded by Gm2S 1 cDNA (SEQ ID NO 1). Arrows indicate endoproteolytic sites that give rise to small subunit (lunasin) (SEQ ID NO 2) and the large subunit (methionine rich protein). Important regions in both subunits are indicated.

#FIG. 2 shows the results of experiments measuring HMG CoA reductase expression when cholesterol free media is treated with lunasin.

#Lunasin (a.k.a. lunastatin) is a recently discovered bioactive component in soy with a novel chromatin-binding property and epigenetic effects on gene expression (17, 18). The lunasin soy peptide is heat stable, water soluble and found in significant amounts in select soy protein preparations (19). Studies show that it can get inside mammalian epithelial cells through its RGD cell adhesion motif, bind preferentially to deacetylated histones and inhibit histone H3 and H4 acetylation (20). There is growing evidence that cellular transformation, responses to hormones and dietary and environmental effects involve epigenetic changes in gene expression, which are modulated by the reversible processes of DNA methylation-demethylation and histone acetylation-deacetylation (21, 22). Lunasin is the first natural substance to be identified as a histone acetylase inhibitor, although it does not directly affect the histone acetylase enzyme. It inhibits H3 and H4 acetylation by binding to specific deacetylated lysine residues in the N-terminal tail of histones H3 and H4, making them unavailable as substrates for histone acetylation. The elucidation of the mechanism of action makes lunasin an important molecule for research studies to understand the emerging role of epigenetics and chromatin modifications in important biological processes.

#The study on the effect of lunasin on prostate carcinogenesis at the University of California at Davis revealed the effects of lunasin on histone H4 modifications and the up regulation of chemopreventive genes, (23). However, until now, the specific effect of lunasin binding to deacetylated H3 N-terminal tail and the inhibition of H3 histone acetylation in biological systems had not yet been investigated. To determine the specific biological effect of lunasin binding to deacetylated histone H3 and inhibition of acetylation, the induction of genes involved in cholesterol biosynthesis by the sterol regulatory element binding proteins (SREBP) was chosen as a biological model. This biological model was chosen because activation of SREBPs by sterol depletion results in the increased acetylation of histone H3 but not histone H4, by the histone acetylase enzyme PCAF, in chromatin proximal to the promoters of HMG CoA reductase and the LDL receptor genes (24) and SREBP activation results in the increased recruitment of co-regulatory factors, CREB to the promoter of HMG CoA reductase gene, and Sp1 to the promoter of LDL receptor gene (24).

#Our studies on in vitro histone acetylase (HAT) assays show that lunasin significantly inhibits histone H3 acetylation (specifically lysine 14 in H3 N-terminal tail) by the histone acetylase enzyme, PCAF. Cell culture experiments using HepG2 liver cells show that synthetic lunasin can significantly reduce HMG CoA reductase expression and increase LDL receptor gene expression in cholesterol-free media similar to the effects of statin (cholesterol-lowering) drugs. Our studies have also shown that the increase in LDL receptor expression coincides with the increase in Sp1 expression in cholesterol-free media. Based on these studies, a molecular mechanism of action is proposed wherein synthetic lunasin reduces total and LDL cholesterol levels by binding to deacetylated histone H3 and inhibiting histone H3 acetylation by PCAF (through its association with the CREB-binding protein), thereby reducing SREBP activation of the HMG CoA reductase gene resulting in lower endogenous cholesterol biosynthesis, and by increasing the expression of the Sp1 co-activator in sterol-free media and upon SREBP activation, higher amount of membrane bound LDL receptors is expressed leading to significant reduction of plasma LDL cholesterol levels (25).