Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
Filters applied. Clear all
. 2016 Jul;44(7):1027-37.
doi: 10.1124/dmd.116.069344. Epub 2016 Feb 26.

Role of Chromatin Structural Changes in Regulating Human CYP3A Ontogeny

Affiliations
Free PMC article

Role of Chromatin Structural Changes in Regulating Human CYP3A Ontogeny

Nicholas L Giebel et al. Drug Metab Dispos. .
Free PMC article

Abstract

Variability in drug-metabolizing enzyme developmental trajectories contributes to interindividual differences in susceptibility to chemical toxicity and adverse drug reactions, particularly in the first years of life. Factors linked to these interindividual differences are largely unknown, but molecular mechanisms regulating ontogeny are likely involved. To evaluate chromatin structure dynamics as a likely contributing mechanism, age-dependent changes in modified and variant histone occupancy were evaluated within known CYP3A4 and 3A7 regulatory domains. Chromatin immunoprecipitation using fetal or postnatal human hepatocyte chromatin pools followed by quantitative polymerase chain reaction DNA amplification was used to determine relative chromatin occupancy by modified and variant histones. Chromatin structure representing a poised transcriptional state (bivalent chromatin), indicated by the occupancy by modified histones associated with both active and repressed transcription, was observed for CYP3A4 and most 3A7 regulatory regions in both postnatal and fetal livers. However, the CYP3A4 regulatory regions had significantly greater occupancy by modified histones associated with repressed transcription in the fetal liver. Conversely, some modified histones associated with active transcription exhibited greater occupancy in the postnatal liver. CYP3A7 regulatory regions also had significantly greater occupancy by modified histones associated with repressed transcription in the fetus. The observed occupancy by modified histones is consistent with chromatin structural dynamics contributing to CYP3A4 ontogeny, although the data are less conclusive regarding CYP3A7. Interpretation of the latter data may be confounded by cell-type heterogeneity in the fetal liver.

Figures

Fig. 1.
Fig. 1.
CYP3A4 (A) and CYP3A7 (B) promoters and known regulatory regions. The major regulatory regions, the CLEM4, XREM, C/EBPβ, and proximal promoter are expanded, with transcription factor binding sites indicated. AP1, activator protein 1; HNF4, hepatocyte nuclear factor 4; KLF9, Kruppel-like factor 9; NF1/HNF3, nuclear factor 1/hepatocyte nuclear factor 3; PAR, proline acid rich factor; PXR, pregnane X receptor; Sp/XKLF, sephacryl phosphocellulose/Kruppel-like factor; AP1, activator protein 1; USF1, upstream factor 1.
Fig. 2.
Fig. 2.
Chromatin occupancy by H3K27me3 and C/EBPβ within the CYP3A4 C/EBPβ binding region in fetal (○) and postnatal (●) hepatic chromatin pools. The C/EBPβ binding region with targeted amplicons and transcription factor binding sites is indicated. Median ΔCq values are shown plus the data range (* = P ≤ 0.05; *** = P ≤ 0.001, Mann-Whitney U test).
Fig. 3.
Fig. 3.
CYP3A4 promoter occupancy by modified and variant histones or transcription factors. (A) Shown are the four known CYP3A4 regulatory domains probed in the current study, all known transcription factors binding within each domain, and the qPCR amplicons used to evaluate ChIP enrichment. ChIP was performed with antibodies for modified and variant histones, as well as a single transcription factor known to bind within each domain, followed by qPCR to quantify occupancy. The data for pooled fetal (N = 14) (B) and postnatal (N = 10) (C) hepatic chromatin are shown as blue-tinted heat maps representing a range of median ΔCq values from a minimum of three experiments for each amplicon within the indicated CYP3A4 region. N/A, not assayed. Modified and variant histones are grouped into those associated with active (green font), poised (blue font), or inactive transcription (red font). The relevant occupancy by select transcription factors (black font) is also shown. AP1, activator protein 1; HNF4, hepatocyte nuclear factor 4; KLF9, Kruppel-like factor 9; NF1/HNF3, nuclear factor 1/hepatocyte nuclear factor 3; PAR, proline acid rich factor; PXR, pregnane X receptor; USF1, upstream factor 1; Sp/XKLF, sephacryl phosphocellulose/Kruppel-like factor.
Fig. 4.
Fig. 4.
Differences in CYP3A4 promoter occupancy by modified and variant histones between pooled fetal (N = 14) and pooled postnatal (N = 10) hepatic chromatin were evaluated using a Mann-Whitney U test. The data are shown as red- or green-tinted heat maps (see legend) of the P value ranges. Red and green tints indicate increasingly greater degrees of significant differences between chromatin occupancy between postnatal and fetal chromatin, respectively. Yellow indicates no significant difference (NS). N/A, not assayed. Modified and variant histones are grouped into those associated with active (green font), poised (blue font), or inactive transcription states (red font). The relevant occupancy by transcription factors (black font) is also shown.
Fig. 5.
Fig. 5.
CYP3A7 promoter occupancy by modified and variant histones or transcription factors. (A) Shown are the three known CYP3A7 regulatory domains probed in the current study, all known transcription factors binding within each domain, and the qPCR amplicons used to evaluate ChIP enrichment. ChIP was performed with antibodies for modified and variant histones, as well as a single transcription factor known to bind within each domain, followed by qPCR to quantify occupancy. The data for pooled fetal (N = 14) (B) and postnatal (N = 10) (C) hepatic chromatin are shown as blue-tinted heat maps representing a range of median ΔCq values from a minimum of three experiments for each amplicon within the indicated CYP3A7 region. N/A, not assayed. Modified and variant histones are grouped into those associated with active (green font), poised (blue font), or inactive transcription (red font). The relevant occupancy by select transcription factors (black font) is also shown. HNF4, hepatocyte nuclear factor 4; KLF9, Kruppel-like factor 9; NF1/HNF3, nuclear factor 1/hepatocyte nuclear factor 3; PXR, pregnane X receptor.
Fig. 6.
Fig. 6.
Differences in CYP3A7 promoter occupancy by modified and variant histones or transcription factors between pooled fetal (N = 14) and pooled postnatal (N = 10) hepatic chromatin were evaluated using a Mann-Whitney U test. The data are shown as red- or green-tinted heat maps (see legend) of the P value ranges. Red and green tints indicate greater occupancy in the postnatal and fetal chromatin, respectively. Yellow indicates no significant difference (NS). N/A, not assayed. Modified and variant histones are grouped into those associated with active (green font), poised (blue font), or inactive transcription (red font). The relevant occupancy by transcription factors (black font) is also shown.

Similar articles

See all similar articles

MeSH terms

LinkOut - more resources

Feedback