Use this URL to cite or link to this record in EThOS:
Title: Transcriptional regulation of PGC-1a in adipocyte cell lines
Author: Karamitri, Angeliki
ISNI:       0000 0001 3594 9134
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2009
Availability of Full Text:
Full text unavailable from EThOS.
Please contact the current institution’s library for further details.
PGC-la, a cold inducible coactivator of PPARy, has been reported to play an important role in the control ofpre-adipocyte differentiation to brown adipose tissue (BAT) and the expression of the BATspecific gene, uncoupling protein 1 (UCP I). The overall aim of this study was to compare the transcriptional control of PGC-la in brown (HIB-IB) and white (3T3-Ll) adipose tissue precursor cells. To achieve this, a 264bp reporter construct of the PGC-la proximal promoter (PGC-I a-pGL3), that was shown to support luciferase expression to a greater extent in HIB-IB compared with 3T3-LI cells, was used under basal and cAMP stimulated conditions. This fragment of the promoter contains a cAMP response element (CRE) at which a broad array of the bZIP family of transcription factors can potentially bind. Electrophoresis mobility shift assays using a PGC-Ia-CRE oligonucleotide indicated that CREB, ATF-2 and CIEBPfj but not CIEBPa or ClEBP5 bound to the CRE on the PGC-Ia promoter region in HffilB and 3T3-Ll cells. Co-transfection ofHIB-IB and 3T3-LI cells with PGCIa- pGL3 and expression plasmids for CIEBPa, CIEBPfj and ClEBP5 showed that C/EBP(3 but to a lesser extent C/EBP5 and CIEBPa, conferred a marked increase on the promoter activity under forskolin stimulated conditions. The stimulatory effect of CIEBP(3 was abolished when the CRE site was mutated demonstrating that CIEBP(3 stimulates transcription of PGC-la by acting through the CRE site. CIEBP(3 induction of the PGC-la expression was confirmed by demonstrating that PGC-Ia mRNA levels were increased after CIEBP(3 overxpression and forskolin stimulation to. Expression ~f UCP-I mRNA was also increased in 3T3-Ll cells under CIEBPI1 overxpression and forskolm stimulation suggesting that CIEBPI1 was able to reprogramme 3T3-LI cells to the BAT lineage. To establish the mechanism for the effect of CIEBP(3, I further characterised the PGC-Ia-CRE by investigating the effects of overexpressing CRE-associated transcription factors, using PGC-l reporter assays. CREB was able to increase PGC-la transcription, whereas ATF-2 and CHOP-tO overexpression resulted in decreases in transcriptional activation conferred by CIEBP(3. Results fro~ chromatin immunoprecipitation assays showed that CIEBPI1 and CREB were the strongest protem complexes bound to the CRE site under CIEBP,6 overexpresion and forskolin stimulated conditions, while ATF-2 and CHOP-tO binding was diminished. The results demonstrated that different combinations of transcription factors binding to the proximal CRE ofPGC-Ict. may be responsible for the differential response ofPGC-Ia. expression to cAMP in brown versus white adipose tissue.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available