Cells respond to biotin deficiency by increasing the expression of the biotin transporters SMVT and MCT1, which serve as """"""""checkpoints"""""""" for biotin entry into cells. It is unknown, however, how cells sense biotin status and which mechanisms mediate regulation of biotin transporters. Evidence has been provided that biotin regulates gene expression directly at the chromatin level. Previous studies suggested that holocarboxylase synthetase (HCS) mediates the binding of biotin to histones (DNA-binding proteins) H2A, H3, and H4, and that biotinylation of histones causes gene repression. Our studies are consistent with the hypotheses that biotin-dependent nuclear translocation of HCS serves as a biotin sensor, and that binding of biotin to histones by HCS is associated with chromatin remodeling events that regulate the transcription of biotin transporter genes. Long-term objective: Our long-term objective is to elucidate mechanisms of biotin homeostasis in humans. We seek to identify both sensors of cellular biotin and mechanisms that regulate the """"""""checkpoints"""""""" for biotin entry into cells, SMVT and MCT1. We also seek to elucidate mechanisms of HCS regulation in humans by identifying and characterizing HCS-binding proteins.
Specific aim : To identify mechanisms of biotin-dependent nuclear translocation of HCS, and to characterize HCS-dependent chromatin remodeling events that affect gene transcription at biotin transporter loci.
This aim will test the following hypotheses. (1) HCS serves as a biotin sensor in human cytoplasm. Increased cellular concentrations of biotin are associated with HCS-mediated biotinylation of HCS-binding proteins, triggering nuclear translocation of HCS. (2) Nuclear HCS-binding proteins recruit HCS to specific regions in chromatin, including SMVT and MCT1 loci. (3) HCS catalyzes biotinylation of histones at target loci;the increased biotinylation of histones at SMVT and MCT1 loci in response to biotin supplementation decreases the transcription of biotin transporter genes SMVT and MCT1. (4) Collectively, intracellular biotin directly controls the expression of biotin transporters, mediated by HCS-dependent chromatin remodeling. Methods: HCS-binding proteins in cytoplasm and nucleus will be identified by using techniques such as yeast-two-hybrid assays, in silico domain searches, co-immunoprecipitations, and transgenic cell lines. The relative enrichment of HCS and biotinylated histones at biotin transporter loci will be quantified by chromatin immunoprecipitation assays and real-time PCR in both human biotin supplementation studies and human cell lines. Transcription of SMVT and MCT1 will be quantified by using real-time PCR and reporter-gene constructs in both human biotin supplementation studies and transgenic cell lines.

Public Health Relevance

Relevance to public health: Biotinylation of histones is a unique epigenetic mark because it depends on the dietary intake of the essential vitamin biotin. Biotin deficiency is prevalent among Americans, and moderate biotin deficiency has been observed in up to 50% of pregnant women. Previous studies suggest that biotinylation of histones plays a critical role in gene regulation and genomic stability, thereby decreasing the risk for chromosomal abnormalities and cancer.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK077816-03
Application #
8019073
Study Section
Integrative Nutrition and Metabolic Processes Study Section (INMP)
Program Officer
Maruvada, Padma
Project Start
2009-02-01
Project End
2013-01-31
Budget Start
2011-02-01
Budget End
2013-01-31
Support Year
3
Fiscal Year
2011
Total Cost
$247,108
Indirect Cost
Name
University of Nebraska Lincoln
Department
Miscellaneous
Type
Schools of Education
DUNS #
555456995
City
Lincoln
State
NE
Country
United States
Zip Code
68588
Cordonier, Elizabeth L; Adjam, Riem; Teixeira, Daniel Camara et al. (2015) Resveratrol compounds inhibit human holocarboxylase synthetase and cause a lean phenotype in Drosophila melanogaster. J Nutr Biochem 26:1379-84
Liu, Dandan; Zempleni, Janos (2014) Transcriptional regulation of the albumin gene depends on the removal of histone methylation marks by the FAD-dependent monoamine oxidase lysine-specific demethylase 1 in HepG2 human hepatocarcinoma cells. J Nutr 144:997-1001
Baier, Scott R; Nguyen, Christopher; Xie, Fang et al. (2014) MicroRNAs are absorbed in biologically meaningful amounts from nutritionally relevant doses of cow milk and affect gene expression in peripheral blood mononuclear cells, HEK-293 kidney cell cultures, and mouse livers. J Nutr 144:1495-500
Liu, Dandan; Zempleni, Janos (2014) Low activity of LSD1 elicits a pro-inflammatory gene expression profile in riboflavin-deficient human T Lymphoma Jurkat cells. Genes Nutr 9:422
Zempleni, Janos; Liu, Dandan; Camara, Daniel Teixeira et al. (2014) Novel roles of holocarboxylase synthetase in gene regulation and intermediary metabolism. Nutr Rev 72:369-76
Li, Yong; Malkaram, Sridhar A; Zhou, Jie et al. (2014) Lysine biotinylation and methionine oxidation in the heat shock protein HSP60 synergize in the elimination of reactive oxygen species in human cell cultures. J Nutr Biochem 25:475-82
Liu, Dandan; Zempleni, Janos (2014) Holocarboxylase synthetase interacts physically with nuclear receptor co-repressor, histone deacetylase 1 and a novel splicing variant of histone deacetylase 1 to repress repeats. Biochem J 461:477-86
Baier, Scott R; Zbasnik, Richard; Schlegel, Vicki et al. (2014) Off-target effects of sulforaphane include the derepression of long terminal repeats through histone acetylation events. J Nutr Biochem 25:665-8
Xue, Jing; Wijeratne, Subhashinee S K; Zempleni, Janos (2013) Holocarboxylase synthetase synergizes with methyl CpG binding protein 2 and DNA methyltransferase 1 in the transcriptional repression of long-terminal repeats. Epigenetics 8:504-11
Singh, Mahendra P; Wijeratne, Subhashinee S K; Zempleni, Janos (2013) Biotinylation of lysine 16 in histone H4 contributes toward nucleosome condensation. Arch Biochem Biophys 529:105-11

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