An enzymatic mechanism has been proposed by which biotinidase may catalyze biotinylation of histones. Here, human cells were found to covalently bind biotin to histones H1, H2A, H2B, H3, and H4. Cells respond to proliferation with increased biotinylation of histones; biotinylation increases early in the cell cycle and remains increased during the cycle. Notwithstanding the catalytic role of biotinidase in biotinylation of histones, mRNA encoding biotinidase and biotinidase activity did not parallel the increased biotinylation of histones in proliferating cells. Biotinylation of histones might be regulated by enzymes other than biotinidase or by the rate of histone debiotinylation.

Biotin supply may affect transcription of genes and biotinylation of proteins in cells. In this study, Jurkat cells were used to model effects of biotin supply on biotin homeostasis and interleukin-2 metabolism in immune cells. Cells were cultured in media containing deficient (25 pmol/L), physiologic (250 pmol/L), or pharmacologic concentrations (10,000 pmol/L) of biotin for 4 wk. Activities of the biotin-dependent enzyme propionyl-CoA carboxylase paralleled the biotin concentrations in media [pmol bicarbonate fixed/(min x 10(6) cells)]: 1.9 +/- 0.7 (25 pmol/L biotin) vs. 19 +/- 1.2 (250 pmol/L biotin) vs. 40 +/- 2.0 (10,000 pmol/L biotin). Cells responded to biotin deficiency with increased expression of biotin transporter genes. Biotin-deficient cells maintained normal biotinylation of histones but contained reduced levels of biotinylated carboxylases, suggesting compartmentalization of intracellular biotin distribution. Rates of cell proliferation and activities of the apoptotic enzyme caspase-3 were similar among treatment groups, suggesting that net proliferation was not affected by biotin status. Net secretion of interleukin-2 by Jurkat cells was inversely associated with the biotin concentration in media [kU/(L x 24 h x 10(6) cells)]: 21 +/- 1.8 (25 pmol/L biotin) vs. 15 +/- 5.4 (250 pmol/L biotin) vs. 6.1 +/- 1.8 (10,000 pmol/L biotin), suggesting increased secretion or decreased internalization of interleukin-2 by biotin-deficient cells. This study provides evidence that biotin supply affects biotinylation of proteins, gene expression and metabolism of interleukin-2 in Jurkat cells. The physiological significance of effects of biotin status on metabolism of interleukin-2 remains to be elaborated.

Histones are modified post‐translationally, e.g. by methylation of lysine and arginine residues, and by phosphorylation of serine residues. These modifications regulate processes such as gene expression, DNA repair, and mitosis and meiosis. Recently, evidence has been provided that histones are also modified by covalent binding of the vitamin biotin. The aims of this study were to identify biotinylation sites in histone H3, and to investigate the crosstalk among histone biotinylation, methylation and phosphorylation. Synthetic peptides based on the sequence of human histone H3 were used as substrates for enzymatic biotinylation by biotinidase; biotin in peptides was probed using streptavidin peroxidase. These studies provided evidence that K4, K9 and K18 in histone H3 are good targets for biotinylation; K14 and K23 are relatively poor targets. Antibodies were generated to histone H3, biotinylated either at K4, K9 or K18. These antibodies localized to nuclei in human placental cells in immunocytochemistry and immunoblotting experiments, suggesting that lysines in histone H3 are biotinylated in vivo. Dimethylation of R2, R8 and R17 increased biotinylation of K4, K9 and K18, respectively, by biotinidase; phosphorylation of S10 abolished biotinylation of K9. These observations are consistent with crosstalk between biotinylation of histones and other known modifications of histones. We speculate that this crosstalk provides a link to known roles for biotin in gene expression and cell proliferation.

Covalent modifications of histones play crucial roles in chromatin structure and genomic stability. Recently, we reported a novel modification of histones: biotinylation of lysine residues. Here we provide evidence that K12-biotinylated histone H4 (K12Bio H4) maps specifically to both heterochromatin (alpha satellite repeats in pericentromeric regions) and transcriptionally repressed chromatin (g-G globin and interleukin-2) in human lymphoblastoma cells. The abundance of K12Bio H4 in these regions was similar to that of K9-dimethylated histone H3, a known marker for heterochromatin. Likewise, K8-biotinylated histone H4 (K8Bio H4) mapped to heterochromatin, but the relative enrichment was smaller compared with K12Bio H4. Stimulation of interleukin-2 transcriptional activity with phorbol-12-myristate-13-acetate and phytohemagglutinin caused a rapid depletion of K12Bio H4 in the gene promoter. These data are consistent with a novel role for biotin in chromatin structure and transcriptional activity of genes. D

Biotin in breakdown products of biotinylated carboxylases serves as substrate for biotinylation of histones by biotinidase. Here we determined whether biotinylation of histones might play a role in repair of damaged DNA and in apoptosis. Jurkat cells were exposed to UV light to induce DNA damage. Abundance of thymine dimers increased about three times in response to UV exposure, consistent with DNA damage. Biotin-containing carboxylases were degraded in response to UV exposure, as judged by Western blot analysis and carboxylase activities. Mitochondrial integrity decreased in response to UV exposure (as judged by confocal microscopy), facilitating the release of breakdown products of carboxylases from mitochondria. Biotinylation of histones increased in response to UV exposure; biotinylation of histones did not occur specifically at sites of newly repaired DNA. UV exposure triggered apoptosis, as judged by caspase-3 activity and analysis by confocal microscopy. In summary, this study provided evidence that increased biotinylation of histones in DNA-damaged cells might either be a side product of carboxylase degradation or a step during apoptosis.