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Effect of dietary protein restriction on liver transcription factors.

ABSTRACT: The transcription of several genes that are preferentially expressed in the liver, including the serum albumin, transthyretin and carbamyl phosphate synthetase-I genes, is specifically decreased in animals consuming inadequate amounts of dietary protein. The high level of transcription of these genes in the liver is directed in part by a number of liver-enriched transcription factors, including hepatocyte nuclear factors (HNF)-1, -3, and -4, and proteins of the CCAAT/enhancer-binding protein (C/EBP) family. In the present study, we investigated the possibility that the co-ordinate decrease in transcription of the nutritionally sensitive genes in protein-deprived rats results from altered activity of one or more of the liver-enriched transcription factors. For HNF-4, Western blots indicated no change in the level of nuclear HNF-4 protein in liver of protein-deprived animals, whereas we observed a 40% reduction in the DNA binding activity of HNF-4 as measured by electrophoretic mobility shift assay (EMSA). Furthermore, the binding affinity of HNF-4 for DNA was unaltered by dietary protein deprivation, while the number of HNF-4 molecules able to bind to DNA (Bmax) was reduced, as determined by Scatchard analysis. This indicates that in the protein-restricted rats a portion of the pool of HNF-4 protein is inactivated or otherwise prevented from binding to DNA. The overall DNA binding activity of C/EBP alpha and beta was increased in protein-restricted animals. This change occurred in the absence of a change in the amount of the full-length forms of these two proteins, quantified by Western blotting. Interestingly, dietary protein restriction specifically increased the level of a truncated form of C/EBP beta (liver-enriched transcriptional inhibitory protein, LIP), which is a protein dominant negative inhibitor of C/EBP function. Analysis of HNF-3 DNA-binding activity by EMSA revealed that HNF-3 alpha and beta DNA binding was increased and that HNF-3 gamma DNA-binding activity was unchanged in protein-restricted animals. We also detected two apparently novel shift complexes with the HNF-3 probe by EMSA, both of which were decreased in protein-restricted animals. HNF-1 DNA-binding activity was increased by dietary protein restriction. We also examined the effect of protein restriction on the DNA-binding activity of two ubiquitous transcription factors, NF1 and Sp1. The DNA binding activity of the major NF1 isoforms was unchanged whereas the binding activity of Sp1 was increased in the protein-restricted animals. In summary, restriction of dietary protein resulted in a number of specific changes in the DNA-binding activity of various transcription factors. Because transcriptional activation typically involves the synergistic action of more than one transcription factor, small changes in the amount/activity of several factors, could have a strong net effect on the transcription of many genes.

SUBMITTER: Marten NW

PROVIDER: S-EPMC1217496 | biostudies-other | 1996 Jul

REPOSITORIES: biostudies-other

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Project description:Dietary restriction (DR), the process of decreasing overall food consumption over an extended period of time, has been shown to increase longevity across evolutionarily diverse species and delay the onset of age-associated diseases in humans. In Caenorhabditis elegans, the Myc-family transcription factors (TFs) MXL-2 (Mlx) and MML-1 (MondoA/ChREBP), which function as obligate heterodimers, and PHA-4 (orthologous to FOXA) are both necessary for the full physiological benefits of DR. However, the adaptive transcriptional response to DR and the role of MML-1::MXL-2 and PHA-4 remains elusive. We identified the transcriptional signature of C. elegans DR, using the eat-2 genetic model, and demonstrate broad changes in metabolic gene expression in eat-2 DR animals, which requires both mxl-2 and pha-4. While the requirement for these factors in DR gene expression overlaps, we found many of the DR genes exhibit an opposing change in relative gene expression in eat-2;mxl-2 animals compared to wild-type, which was not observed in eat-2 animals with pha-4 loss. Surprisingly, we discovered more than 2000 genes synthetically dysregulated in eat-2;mxl-2, out of which the promoters of down-regulated genes were substantially enriched for PQM-1 and ELT-1/3 GATA TF binding motifs. We further show functional deficiencies of the mxl-2 loss in DR outside of lifespan, as eat-2;mxl-2 animals exhibit substantially smaller brood sizes and lay a proportion of dead eggs, indicating that MML-1::MXL-2 has a role in maintaining the balance between resource allocation to the soma and to reproduction under conditions of chronic food scarcity. While eat-2 animals do not show a significantly different metabolic rate compared to wild-type, we also find that loss of mxl-2 in DR does not affect the rate of oxygen consumption in young animals. The gene expression signature of eat-2 mutant animals is consistent with optimization of energy utilization and resource allocation, rather than induction of canonical gene expression changes associated with acute metabolic stress, such as induction of autophagy after TORC1 inhibition. Consistently, eat-2 animals are not substantially resistant to stress, providing further support to the idea that chronic DR may benefit healthspan and lifespan through efficient use of limited resources rather than broad upregulation of stress responses, and also indicates that MML-1::MXL-2 and PHA-4 may have distinct roles in promotion of benefits in response to different pro-longevity stimuli.

Project description:Dietary restriction (DR), the process of decreasing overall food consumption over an extended period of time, has been shown to increase longevity across evolutionarily diverse species and delay the onset of age-associated diseases in humans. In Caenorhabditis elegans, the Myc-family transcription factors (TFs) MXL-2 (Mlx) and MML-1 (MondoA/ChREBP), which function as obligate heterodimers, and the forkhead box transcription factor A ortholog (PHA-4) are both necessary for the full physiological benefits of DR. However, the adaptive transcriptional response to DR and the role of MML-1::MXL-2 and PHA-4 remains elusive. We identified the transcriptional signature of C. elegans DR, using the eat-2 genetic model, and demonstrate broad changes in metabolic gene expression in eat-2 DR animals, which requires both mxl-2 and pha-4. While the requirement for these factors in DR gene expression overlaps, we found many of the DR genes exhibit an opposing change in relative gene expression in eat-2;mxl-2 animals compared to wild-type, which was not observed in eat-2 animals with pha-4 loss. We further show functional deficiencies of the mxl-2 loss in DR outside of lifespan, as eat-2;mxl-2 animals exhibit substantially smaller brood sizes and lay a proportion of dead eggs, indicating that MML-1::MXL-2 has a role in maintaining the balance between resource allocation to the soma and to reproduction under conditions of chronic food scarcity. While eat-2 animals do not show a significantly different metabolic rate compared to wild-type, we also find that loss of mxl-2 in DR does not affect the rate of oxygen consumption in young animals. The gene expression signature of eat-2 mutant animals is consistent with optimization of energy utilization and resource allocation, rather than induction of canonical gene expression changes associated with acute metabolic stress -such as induction of autophagy after TORC1 inhibition. Consistently, eat-2 animals are not substantially resistant to stress, providing further support to the idea that chronic DR may benefit healthspan and lifespan through efficient use of limited resources rather than broad upregulation of stress responses, and also indicates that MML-1::MXL-2 and PHA-4 may have different roles in promotion of benefits in response to different pro-longevity stimuli.

Project description:Background and objectivesMetabolomics is a relatively new field of "-omics" research, focusing on high-throughput identification of small molecular weight metabolites. Diet has both acute and chronic effects on metabolic profiles; however, alterations in response to dietary sodium restriction (DSR) are completely unknown. The goal of this study was to explore changes in urine metabolites in response to DSR, as well as their association with previously reported improvements in vascular function with DSR.Design, setting, participants, & measurementsUsing stored urine samples from a 10-week randomized placebo-controlled crossover study of DSR in 17 middle-aged/older adults (six men and 11 women; mean age 62±8 years) who had moderately elevated systolic BP (130-159 mmHg) and were otherwise healthy, a liquid chromatography/mass spectrometry-based analysis of 289 metabolites was performed. This study identified metabolites that were significantly altered between the typical (153±29 mmol/d) and low (70±29 mmol/d) sodium conditions, as well as their baseline (typical sodium) association with responsiveness to previously reported improvements in vascular endothelial function (brachial artery flow-mediated dilation) and large elastic artery stiffness (aortic pulse wave velocity).ResultsOf the 289 metabolites surveyed, 10 were significantly altered (nine were upregulated and one was downregulated) during the low sodium condition, and eight of these exceeded our prespecified clinically significant threshold of a >40% change. These metabolites were involved in biologic pathways broadly related to cardiovascular risk, nitric oxide production, oxidative stress, osmotic regulation, and metabolism. One metabolite, serine, was independently (positively) associated with previously reported improvements in the primary vascular outcome of brachial artery flow-mediated dilation.ConclusionsThis proof-of-concept study provides the first evidence that DSR is a stimulus that induces significant changes in urinary metabolomic profiles. Moreover, serine was independently associated with corresponding changes in vascular endothelial function after DSR. Larger follow-up studies will be required to confirm and further elucidate the metabolic pathways that are altered in response to DSR.

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Effect of dietary protein restriction on liver transcription factors.

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