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PubMed1995

Transcriptional activation by Myc is under negative control by the transcription factor AP-2.

The Myc protein binds to and transactivates the expression of genes via E-box elements containing a central CAC(G/A)TG sequence. The transcriptional activation function of Myc is required for its ability to induce cell cycle progression, cellular transformation and apoptosis. Here we show that transactivation by Myc is under negative control by the transcription factor AP-2. AP-2 inhibits transactivation by Myc via two distinct mechanisms. First, high affinity binding sites for AP-2 overlap Myc-response elements in two bona fide target genes of Myc, prothymosin-alpha and ornithine decarboxylase. On these sites, AP-2 competes for binding of either Myc/Max heterodimers or Max/Max homodimers. The second mechanism involves a specific interaction between C-terminal domains of AP-2 and the BR/HLH/LZ domain of Myc, but not Max or Mad. Binding of AP-2 to Myc does not preclude association of Myc with Max, but impairs DNA binding of the Myc/Max complex and inhibits transactivation by Myc even in the absence of an overlapping AP-2 binding site. Taken together, our data suggest that AP-2 acts as a negative regulator of transactivation by Myc.

Animals↗

PubMed1996

Prostaglandin E2 inhibits the nuclear transcription of the human interleukin 2, but not the Il-4, gene in human T cells by targeting transcription factors AP-1 and NF-AT.

Prostaglandin E2 (PGE2) release from activated macrophages and/or stimulation of T cells is associated with cAMP formation and activation of protein kinase A (PKA). cAMP inhibits Th1- but not Th2-cytokine production and may influence the nature of the immune response to a given antigen. Using DNA transfection and electrophoretic mobility shift assays (EMSA), we have examined the mechanisms for the transcriptional regulation of human IL-2 and IL-4 genes by PGE2. Stimulation of Jurkat cells with ionomycin and PMA in the presence of PGE2 inhibited the IL-2- but not the IL-4-promoter activity. In EMSAs, nuclear extracts from primary human T cells stimulated with ionomycin and phorbol esters in the presence of PGE2 demonstrated decreased binding at the AP-1 and NF-AT sites of the human IL-2 promoter; binding to the OCT-1 and NF-kappa B sites was not affected. These results suggest that cAMP regulates IL-2 production in human T cells by a transcriptional mechanism which involves discrete transactivating pathways for IL-2-promoter activation.

Calcium↗

PubMed1995

Recycling of the general transcription factors during RNA polymerase II transcription.

We have analyzed the fate of the RNA polymerase II (RNAPII) general transcription factors during the transition from initiation to elongation using multiple approaches. We demonstrate that all of the basal factors coexist in mature initiation complexes but that following nucleotide addition, this complex becomes disrupted. During this transition, TFIID remains promoter-bound whereas TFIIB, TFIIE, TFIIF, and TFIIH are released. Upon release, TFIIB reassociates with TFIID, reforming the RNAPII docking site, the DB complex. TFIIE is released before formation of the tenth phosphodiester bond. This precedes TFIIH release, which occurrs after the transcription complex reaches +30. TFIIF is unique in that it is the only basal factor detected in the RNAPII elongation complex. Following its release from the initiation complex, TFIIF has the ability to reassociate with a stalled RNAPII.

Adenoviridae↗

PubMed2003

RNA leaching of transcription factors disrupts transcription in myotonic dystrophy.

Myotonic dystrophy type 1 (DM1) is caused by a CUGn expansion (n approximately 50 to 5000) in the 3' untranslated region of the mRNA of the DM protein kinase gene. We show that mutant RNA binds and sequesters transcription factors (TFs), with up to 90% depletion of selected TFs from active chromatin. Diverse genes are consequently reduced in expression, including the ion transporter CIC-1, which has been implicated in myotonia. When TF specificity protein 1 (Sp1) was overexpressed in DM1-affected cells, low levels of messenger RNA for CIC-1 were restored to normal. Transcription factor leaching from chromatin by mutant RNA provides a potentially unifying pathomechanistic explanation for this disease.

Binding Sites↗

PubMed2000

Transcriptional control of SPARC by v-Jun and other members of the AP1 family of transcription factors.

Transformation of chick embryo fibroblasts by the v-Jun oncoprotein correlates with a down-regulation of the extracellular matrix protein SPARC and repression of the corresponding mRNA. Alteration in SPARC expression has been repeatedly reported in human cancers of various origin, and is thought to contribute to the remodeling of the extracellular matrix during neoplastic progression. Transcriptional control of SPARC is poorly understood. We show here that (i) v-Jun-mediated repression of the endogenous SPARC gene is enhanced by Fra2 but alleviated by ATF2, Fra2 and ATF2 being the two major partners of v-Jun in the transformed cells; (ii) high basal activity as well as repression by v-Jun and modulation by Fra2 and ATF2 is restricted to a small proximal fragment (-124/+16) of the chicken SPARC promoter; (iii) the activity of this minimal promoter is modulated by all the AP1 family members known in chickens (c-Jun and JunD; c-Fos and Fra2; ATF2; c-Maf, MafA, and MafB). Taken together these data demonstrate that, at least in avian primary cells, SPARC expression is under the control of the AP1 transcription factor. Further studies with the minimal (-124/+16) promoter fragment are needed to understand how this control takes place at the molecular level.

Activating Transcription Factor 2↗

PubMed2004

Co-operative interactions between NFAT (nuclear factor of activated T cells) c1 and the zinc finger transcription factors Sp1/Sp3 and Egr-1 regulate MT1-MMP (membrane type 1 matrix metalloproteinase) transcription by glomerular mesangial cells.

The transition of normally quiescent glomerular MCs (mesangial cells) to a highly proliferative phenotype with characteristics of myofibroblasts is a process commonly observed in inflammatory diseases affecting the renal glomerulus, the ultimate result of which is glomerulosclerosis. Generation of proteolytically active MMP (matrix metalloproteinase)-2 by the membrane-associated membrane type 1 (MT1)-MMP is responsible for the transition of mesangial cells to the myofibroblast phenotype [Turck, Pollock, Lee, Marti and Lovett (1996) J. Biol. Chem. 271, 15074-15083]. In the present study, we show that the expression of MT1-MMP within the context of MCs is mediated by three discrete cis -acting elements: a proximal non-canonical Sp1 site that preferentially binds Sp1; an overlapping Sp1/Egr-1-binding site that preferentially binds Egr-1; and a more distal binding site for the NFAT (nuclear factor of activated T cells) that binds the NFAT c1 isoform present in MC nuclear extracts. Transfection with an NFAT c1 expression plasmid, or activation of calcineurin with a calcium ionophore, yielded major increases in NFAT c1 nuclear DNA-binding activity, MT1-MMP transcription and protein synthesis, which were additive with the lower levels of transactivation provided by the proximal Sp1 and the overlapping Sp1/Egr-1 sites. Specific binding of NFAT c1 to the MT1-MMP promoter was confirmed by chromatin immunoprecipitation studies, while MT1-MMP expression was suppressed by treatment with the calcineurin inhibitor, cyclosporin A. These studies are the first demonstration that a specific NFAT isoform enhances transcription of an MMP (MT1-MMP) that plays a major role in the proteolytic events that are a dominant feature of acute glomerular inflammation. Suppression of MT1-MMP by commonly used calcineurin inhibitors may play a role in the development of renal fibrosis following renal transplantation.

Animals↗

PubMed2000

Cell-specific transcription of leukotriene C(4) synthase involves a Kruppel-like transcription factor and Sp1.

Leukotriene C(4) synthase (LTC(4)S) is responsible for the biosynthesis of cysteinyl leukotrienes that participate in allergic and asthmatic inflammation. We analyzed 2.1 kilobases of the 5'-flanking region of the human LTC(4)S gene, which contains three DNase I hypersensitivity sites, for its transcriptional activity when fused to a promoterless and enhancerless luciferase gene. Deletion analysis revealed a nonspecific basal promoter region between nucleotides -122 and -56 upstream of the translation start site which contains a consensus Sp1 binding site and a putative initiator element (Inr) and cell-specific enhancer regions further upstream. A single mutation of either the Sp1 binding site between nucleotides -120 and -115 or the Inr (CAGAC) between nucleotides -66 and -62 reduced the expression of the reporter gene by approximately 60%, whereas double mutations decreased the expression by approximately 80%. The incubation of nuclear extracts from THP-1 and K562 cells with a (32)P-labeled oligonucleotide containing the Sp1 site or the Inr sequence gave gel-shifted complexes that were blocked by their respective cold oligonucleotides, and antisera specific for Sp1 and Sp3 provided supershifts for the former. Linker-scanning mutations of a cell-specific regulatory region revealed that mutations from nucleotides -165 to -125 reduced reporter activity. This region contains a tandem CACCC repeat (at nucleotides -149 to -145 and -139 to -135). An oligonucleotide containing the distal CACCC motif was gel shifted by THP-1 cell nuclear extract and was supershifted by antisera to Sp1 and Sp3. Cotransfection of an Sp1 expression plasmid into Drosophila SL2 cells with a -228 to -3 LTC(4)S reporter construct transactivated the reporter gene, whereas mutations at the CACCC repeat region reduced Sp1 transactivation by approximately 66%. Similarly, the Kruppel-like factor Zf9/CPBP (core promoter-binding protein) transactivated the -228 construct in COS cells but not its CACCC mutant construct. These findings indicate the involvement of Sp1 and an Inr in non-cell-specific regulation and a Kruppel-like transcription factor and Sp1 in the cell-specific regulation of the LTC(4)S gene. These are the first such analyses of a member of a newly recognized superfamily of membrane-associated proteins involved in eicosanoid and glutathione metabolism, which contains key proteins involved in the generation of both prostanoids and cysteinyl leukotrienes.

Base Sequence↗

PubMed2001

Transcriptional regulation of the p67phox gene: role of AP-1 in concert with myeloid-specific transcription factors.

We have investigated the myeloid-specific transcriptional regulation of p67(phox), an essential component of phagocyte respiratory burst NADPH oxidase. Analysis was carried out on the p67(phox) 5'-flanking region from -3669 to -4 (relative to ATG), including the first exon and intron and part of the second exon. The construct extending from -985 to -4 produced the highest luciferase activity in myeloid HL-60 cells but was not active in HeLa or Jurkat cells, indicating myeloid-specific expression. Four active elements were identified: Sp1/Sp3 at -694, PU.1 at -289, AP-1 at -210, and PU.1/HAF1 at -182, the latter three being in the first intron. These cis elements bound their cognate transacting factors both in vitro and in vivo. Mutation of the Sp1, PU.1, or PU.1/HAF1 site each decreased promoter activity by 35-50%. Mutations in all three sites reduced promoter activity by 90%. However, mutation of the AP-1 site alone nearly abolished promoter activity. The AP-1 site bound Jun and Fos proteins from HL-60 cell nuclear extract. Co-expression with Jun B in AP-1-deficient cells increased promoter activity by 3-fold. These data show that full p67(phox) promoter activity requires cooperation between myeloid-specific and nonmyeloid transcription factors, with AP-1 being the most critical for function.

Base Sequence↗

PubMed2000

Catalase expression in Drosophila melanogaster is responsive to ecdysone and exhibits both transcriptional and post-transcriptional regulation.

In the present study, we have examined catalase protein and mRNA levels and the factors that may regulate catalase expression in Drosophila melanogaster during development. Both mRNA and protein changes are in general accord with variations in ecdysteroid titer during development. Differences in mRNA and protein accumulation profiles, particularly in embryos and young adults, suggest that catalase may be regulated at both transcriptional and post-transcriptional levels. It was possible to induce catalase expression by administering exogenous 20-hydroxyecdysone (Ec) in culture at certain stages of development (usually at time points corresponding to previously observed hormone and catalase peaks). Experiments with exogenous administration of Ec, cycloheximide, and actinomycin D suggest a complex interplay of factors affecting catalase expression. In cultured third instar larvae, superinduction of catalase occurred in the presence of both Ec and cycloheximide. If ecdysteroid production was suppressed prior to antibiotic treatment by temperature upshift of the conditional mutant dre4(e55), superinduction occurred mostly at the protein level. In cultured adult abdomens, we observed induction by Ec and superinduction in the presence of hormone and translation or transcription inhibitors. Unlike what was observed in larvae, superinduction of catalase protein was dramatically more pronounced in control flies.

Animals↗

PubMed1990

The zebrafish homeobox gene hox-2.2: transcription unit, potential regulatory regions and in situ localization of transcripts.

The data presented in this report strongly suggest that the genome of the zebrafish, Brachydanio rerio, has a homeobox cluster which is equivalent to the murine Hox-2 locus. In support of this conclusion, we have found two closely linked zebrafish genes which are true homologues of the mouse Hox-2.1 and Hox-2.2 genes. Here we describe structural and functional properties of the zebrafish Hox-2.2 homologue hox-2.2. Furthermore, we have identified another zebrafish gene related to hox-2.2 which appears to correspond to the Hox-6.1 gene of the murine Hox-3 locus. In order to characterize the zebrafish hox-2.2 gene we have determined the genomic DNA sequence of a 3.4 kb SalI fragment. This revealed that the hox-2.2 transcription unit encodes a putative protein of 228 amino acids. The homeodomains of the murine Hox-2.2 and the zebrafish hox-2.2 proteins are almost identical and extensive sequence identity exists in other regions of the two proteins, which share 160 (70%) of the amino acid residues. Also in terms of expression, strong similarities were observed relative to the murine Hox-2.2 gene. Transcripts derived from zebrafish hox-2.2 start to accumulate when somite formation is initiated. Later in development these transcripts are detected mainly in the central nervous system. Reminiscent of Hox-2.2, the rostral boundary of zebrafish hox-2.2 expression is located in the posterior region of the hindbrain. Notably, untranslated regions of the hox-2.2 gene contain several short sequences closely related to a known homeodomain recognition sequence.

Amino Acid Sequence↗

PubMed2006

Platelet derived growth factor induced tenascin-C transcription is phosphoinositide 3-kinase/Akt-dependent and mediated by Ets family transcription factors.

Previous studies have identified several cytokines as inducers of tenascin-C (TN-C) expression in various tissue culture systems. However, the signaling pathways of the regulation of TN-C expression are almost unknown. In this study, we clarified the molecular mechanism(s) underlying the regulation of the TN-C gene by platelet derived growth factor (PDGF) in cultured human dermal fibroblasts. PDGF induced the expression of TN-C protein as well as mRNA in a dose-dependent manner. Actinomycin D, an RNA synthesis inhibitor, significantly blocked the PDGF-mediated upregulation of TN-C mRNA expression, whereas cycloheximide, a protein synthesis inhibitor, did not. The PDGF-mediated induction of TN-C expression was inhibited by the treatment of fibroblasts with a selective phosphoinositide 3-kinase (PI3K) inhibitor, wortmannin, or LY294002. These results suggest that PDGF induced the expression of TN-C at a transcriptional level via phosphoinositide3-kinase/Akt signaling pathways. We performed serial 5' deletions and a transient transfection analysis to define the region in the TN-C promoter mediating the responsiveness to PDGF. Overexpression of Sp1, Ets1, or Ets2 activated the TN-C promoter and superinduced TN-C promoter activity stimulated by PDGF, whereas overexpression of Fli1 inhibited the effects of PDGF on TN-C expression. Mutation of the Sp1/3 binding sites or Ets binding sites in the TN-C promoter region responsible to PDGF abrogated the PDGF-inducible promoter activity. Immunoprecipitation analysis revealed that Sp1, Ets1, and Ets2 form a transcriptionally active complex. On the other hand, the interaction of Fli1 with Sp1 decreased after PDGF treatment. These results suggest that the upregulation of TN-C expression by PDGF involves Ets family transcription factors, co-operating with Sp1.

Base Sequence↗

PubMed1996

Transcriptional and post-transcriptional regulation of GM-CSF-induced IL-1 beta gene expression in PMN.

Polymorphonuclear leukocytes (PMN) play an important role in inflammation, immune responses, and tissue repair by secreting interleukin- 1 beta (IL-1 beta). We investigated the regulation of IL-1 beta gene expression in human PMN treated with granulocyte-macrophage colony-stimulating factor (GM-CSF). GM-CSF induced IL-1 beta mRNA accumulation at 0.1 ng/ml and maximal induction was observed at 1 ng/ml. IL- 1 beta mRNA levels reached a maximum with 1-2 h after stimulation with GM-CSF and returned to baseline levels by 4-6 h. The time course of IL-1 beta mRNA induction by GM-CSF was more protracted than previously reported for PMN stimulated with tumor necrosis factor-alpha (TNF-alpha, 10 ng/ml). Nuclear run-on analysis indicated that GM-CSF, like TNF, increases IL-1 beta transcription. Kinetic studies with the RNA synthesis inhibitor, actinomycin D, showed that GM-CSF induces stable IL-1B mRNA. Cycloheximide enhanced the IL-1 beta mRNA accumulation by GM-CSF at the level of mRNA stabilization, but blocked IL-1 beta mRNA expression by TNF. Thus, GM-CSF increases IL-1 beta message accumulation in PMN at both the transcriptional and post-transcriptional levels by mechanisms that are different from TNF induction of IL-1 beta gene expression.

Cells, Cultured↗

PubMed2002

Hes-1, a known transcriptional repressor, acts as a transcriptional activator for the human acid alpha-glucosidase gene in human fibroblast cells.

Hes-1, the mammalian homologue 1 of Drosophila hairy and Enhancer of split proteins, belongs to a family of basic helix-loop-helix proteins that are essential to neurogenesis, myogenesis, hematopoiesis, and sex determination. Hes-1 is a transcriptional repressor for a number of known genes including the human acid alpha-glucosidase (GAA) gene as we have previously shown in Hep G2 cells. The human GAA gene encodes the enzyme for glycogen breakdown in lysosomes, deficiency of which results in Glycogen Storage Disease type II (Pompe syndrome). Using constructs containing the DNA element that demonstrates repressive activity in Hep G2 cells and conditions in which the same transcription factors, Hes-1 and YY1, bind, we have shown that this element functions as an enhancer in human fibroblasts. Site-directed mutagenesis and overexpression of Hes-1 showed that Hes-1 functions as a transcriptional activator. The dual function of Hes-1 we have found is likely to contribute to the subtle tissue-specific control of this housekeeping gene.

Adult↗

PubMed1996

A transcription map in the CATCH22 critical region: identification, mapping, and ordering of four novel transcripts expressed in heart.

The acronym CATCH22 is used to indicate collectively a group of related phenotypes, namely velocardiofacial syndrome (VCFS), DiGeorge anomaly (DGA), and conotruncal anomaly face, which are associated with deletions within 22q11.2 in the great majority of patients. A deletion map has allowed us to delimit a smallest region of deletion overlap, considerably smaller than the commonly deleted region. We have mapped within this region the chromosomal breakpoint of a balanced translocation patient presenting with a DGA/VCFS phenotype, making this region the strongest candidate for the location of the gene(s) responsible for the disease phenotype. We report a systematic gene search in this region and show the presence of at least six distinct transcripts, two of which have been previously described. The region searched was approximately 270 kb; therefore, an average of one transcript every 45 kb was found. We generated eight new ESTs and mapped two ESTs present in public databases. All six transcripts are expressed in heart, an organ involved in 70%-80% of CATCH22 patients. We show that the multimethod approach to search for expressed sequences is effective and indeed necessary for a comprehensive search and provides molecular tools for further characterization of the potential genes identified.

Abnormalities, Multiple↗

PubMed1996

The mouse DNA binding protein Rc for the kappa B motif of transcription and for the V(D)J recombination signal sequences contains composite DNA-protein interaction domains and belongs to a new family of large transcriptional proteins.

Rc is a DNA binding protein with dual specificities for the V(D)J recombination signal sequences and for the B motif of the immunoglobulin kappa chain gene enhancer. The largest Rc transcript present in lymphoid cells/tissues is approximately 9 kb. Molecular cloning and sequence determination for 8822 bp of mouse Rc cDNA revealed an open reading frame of 2282 amino acids and long 5'- and 3'-untranslated regions. The derived amino acid sequence contains multiple DNA and protein interaction domains. Composite ZAS structures with tandem zinc fingers, an acidic motif, and a Ser/Thr-rich segment are located near the N-terminal and the C-terminal regions. The middle region of Rc contains a lone zinc finger, an acidic motif, a Ser-rich region, a nucleus localization signal, and GTPase motifs. Cloning and characterization of a mouse Rc gene show that the Rc cDNA corresponds to seven exons located in a genomic region spanning 70 kb. Exon 2 is exceptionally large, with 5487 bp. cDNA cloning and Northern blot analyses revealed multiple Rc transcripts, probably generated by alternative splicings. Sequence comparisons show that Rc belongs to a ZAS protein family that is involved in gene transcription and/or DNA recombination. The major histocompatibility complex class I gene enhancer binding proteins MBP1 and MBP2 are other representatives of this ZAS protein family.

Amino Acid Sequence↗

PubMed2001

Differential melting of the transcription start site associated with changes in RNA polymerase-promoter contacts in initiating transcription complexes.

Formaldehyde cross-linking was used in a kinetic analysis of RNA polymerase-lacUV5 promoter interactions in open complexes (RP(o)). RP(o) quenched from 37 degrees C to 14 degrees C isomerised to a closed, competitor resistant, complex (RP(LT)). We observed that contacts of the beta' and sigma subunits with the positions -3, -5 of the non-template DNA strand disappeared very quickly during the first 30 seconds after the temperature downshift. The re-annealing of the DNA downstream of the transcription start site takes place in the same time scale. However re-annealing of the upstream part of the transcription bubble was slower and completed within five minutes. The results support a two-step model of promoter melting and suggest that conformational changes in the RNA polymerase occur concurrently with the melting around the transcription start site.

Cross-Linking Reagents↗

PubMed1992

CFTR illegitimate transcription in lymphoid cells: quantification and applications to the investigation of pathological transcripts.

Since the isolation of the cystic fibrosis transmembrane conductance regulator gene (CFTR) and the characterization of the main mutation (delta F508) in 1989, a large number of rare mutations has been found. Full screening of the CFTR gene is difficult because it is split into 27 exons covering 250 kb of genomic DNA. This gene is essentially expressed in the lung and intestinal tract, neither of which are easily accessible for routine investigations. The recent description of a faint transcription of highly tissue-specific genes in any cell, a phenomenon known as illegitimate transcription, would facilitate the research of mutations and the characterization of truncated m-RNA caused by splicing mutations. Using the polymerase chain reaction on cDNA (cDNA-PCR), we detected transcripts of the CFTR gene in lymphocytes and lymphoblast cells at a very low level (about 300 times less than in lung or intestine). This strategy allowed us to obtain a sufficient amount of cDNA-PCR product compatible with further molecular analyses. We have, therefore, analyzed a cDNA fragment overlapping exons 10 and 11 by polyacrylamide gel electrophoresis and direct sequencing, and detected the delta F508 mutation at this level. Our protocol can be generalized to the investigation of the total 4.5-kb CFTR coding sequence.

Animals↗

PubMed1996

Regulation of the apolipoprotein E by dietary lipids occurs by transcriptional and post-transcriptional mechanisms.

The aim of the present investigation was to study the regulation of apolipoprotein E by two dietary nutrients, saturated fat and cholesterol, known to raise plasma cholesterol levels. ApoE is a protein component of several classes of lipoproteins including VLDL and HDL, and dietary lipids may regulate VLDL and apoE-containing HDL particles through their effects on apoE gene. Male rats and mice were fed the following 4 diets: control diet (C); high cholesterol diet with 0.5% cholesterol (HC); high fat diet with 20% hydrogenated coconut oil (HF); and high fat plus high cholesterol diet with 0.5% cholesterol and 20% fat (HF/ C). Plasma cholesterol levels remained unchanged on HC diet, but in mice VLDL-cholesterol increased by 31%. HF diet increased VLDL and LDL by 15-17% in rats, and 21% in mice. A combination of fat and cholesterol diet showed pronounced effects on plasma lipoprotein concentrations, raising apoB-containing particles by 21% and 44% in mice and rats, respectively. Plasma apoE levels increased significantly on all diets. The mechanism of regulation of increased plasma apoB and apoE levels was examined. Quantification of hepatic apoB mRNA showed a lack of correlation between plasma apoB and hepatic apoB mRNA levels, suggesting that posttranscriptional regulation increased plasma apoB-containing lipoproteins in animals fed saturated fat diets. Hepatic apoE mRNA levels increased significantly in animals fed cholesterol-rich diets. However, despite increased plasma apoE levels on diet containing only saturated fat, hepatic apoE mRNA did not change. Synthesis of apoE on the liver polysomes increased selectively on cholesterol-rich diets. These results suggest that cholesterol-rich diets altered apoE, in part, by transcriptional mechanism, and saturated fat-rich diets increased plasma apoE levels by posttranscriptional mechanism, possibly decreased receptor-mediated uptake of apoE-containing particles. The regulation of LDL receptor was also studied since plasma apoB and E levels may be altered by LDL receptor-mediated uptake by the hepatocytes. As expected, high cholesterol diet decreased LDL receptor mRNA by 30-40%. However, the LDL receptor protein on liver membranes did not change on any of the test diets in both animal species. Hepatic cholesterol content increased several fold selectively on high cholesterol diets. These findings suggest that: 1) both transcriptional and posttranscriptional mechanisms are important in regulating plasma apoB and E containing lipoproteins; 2) dietary cholesterol regulates apoE gene by a transcriptional mechanism and dietary saturated fat by posttranscriptional mechanism; and 3) changes in the hepatic apoE and LDL receptor mRNA are associated with the changes in intracellular cholesterol concentrations.

Animals↗