Effect of 5-Aza-2´-Deoxycytidine and Trichostatin A on Endogenous Versus Ectopic Expression of Placental Members of the Human Growth Hormone Gene Family
| dc.contributor.author | Ganguly, Esha | |
| dc.contributor.examiningcommittee | Nachtigal, Mark (Biochemistry and Medical Genetics) Dodd, Janis (Physiology and Pathophysiology) | en_US |
| dc.contributor.supervisor | Cattini, Peter (Physiology and Pathophysiology) | en_US |
| dc.date.accessioned | 2016-03-07T16:46:24Z | |
| dc.date.available | 2016-03-07T16:46:24Z | |
| dc.date.issued | 2016 | |
| dc.degree.discipline | Physiology and Pathophysiology | en_US |
| dc.degree.level | Master of Science (M.Sc.) | en_US |
| dc.description.abstract | Background: The genes coding for human (h) chorionic somatomammotropin (CS), hCS-A and hCS-B, and placental growth hormone (GH-V), hGH-V are located at a single locus on chromosome 17q22-24. Local regulatory (5´ P and 3´ enhancer) sequences and a remote locus control region (LCR) containing a placenta-specific hypersensitive site (HS) IV, have been implicated in the efficient expression of the placental hCS/GH-V genes, in part through gene transfer studies in placental and non-placental tumor cell lines. However, low levels of endogenous expression are reported in placental tumor cells compared to normal term placenta. Thus it was hypothesized that the hCS/GH-V chromatin structure in human choriocarcinoma cells is less accessible to regulatory regions essential for efficient expression due to DNA and/or histone modifications, specifically methylation and acetylation, respectively. Approach: To assess individual hCS-A, hCS-B and hGH-V gene expression in placental and non-placental tumor cells, and assess the effect of increasing “chromatin accessibility” on hCS/GH-V RNA levels by inhibiting DNA methylation and histone deacetylation using 5-aza-2´-deoxycytidine (azadC) and trichostatin A (TSA). Principal Findings: Low levels of hCS-A, hCS-B and hGH-V RNA were detected in placental and non-placental tumor cells compared to term placenta. A significant >5-fold increase in promoter activity was seen in placental but not non-placental cells transfected with hybrid hCS promoter luciferase genes containing 3´-enhancer sequences. Placental JEG-3 cells pretreated with azadC and TSA resulted in a significant >10-fold increase in hCS-A, hCS-B and hGH-V RNA levels compared to TSA treatment alone, however, a modest ~3-fold effect was seen in non-placental MCF-7 cells. By contrast to the effect of pretreatment with azadC, post-treatment with azadC mutes the stimulatory effects of TSA on hCS/GH-V transcripts. The specificity of the response suggests that azadC treatment, and presumably hypomethylation of DNA, results in an increase in response to TSA and histone hyperacetylation at the hGH/CS locus. An assessment of histone H3/H4 hyperacetylation in JEG-3 cells treated with azadC and TSA versus TSA alone revealed significant increases consistent with a more open chromatin structure including the hCS 3´-enhancer sequences and LCR. Conclusions: These observations suggest that accessibility of remote and local regulatory regions required for efficient placental hGH/CS expression can be restricted by DNA methylation and histone acetylation status. This includes restricting access of the hCS 3´-enhancer sequences to available placental enhancer transcription factors. | en_US |
| dc.description.note | May 2016 | en_US |
| dc.identifier.uri | http://hdl.handle.net/1993/31147 | |
| dc.language.iso | eng | en_US |
| dc.rights | open access | en_US |
| dc.subject | 5-Aza-2´-Deoxycytidine , Trichostatin A, Acetylation, DNA Methylation, Placental, GH-CS | en_US |
| dc.title | Effect of 5-Aza-2´-Deoxycytidine and Trichostatin A on Endogenous Versus Ectopic Expression of Placental Members of the Human Growth Hormone Gene Family | en_US |
| dc.type | master thesis | en_US |