Session: SUN 649-677-Adipocyte Biology
Poster Board SUN-673
Methods: We used the 3T3-L1 adipocyte cell system to perform all studies. ChIP-qPCR was performed to determine enrichment of histone H3 modifications K4-dimethyl (H3K4m2), K4-trimethyl (H3K4m3), K9-dimethyl (H3K9m2). PCR primer sets were designed to tile across the oblocus, such that amplicons were ~100bp in size and separated by ~500bp since well defined PPAR response elements (PPRE) are unknown. In the case of gyk locus, we designed 1 primer set near the described PPREs and 1 primer set ~4kb proximal to that site as a control region where we did not expect to find enrichment of PPAR-γ or of histone marks associated with transcriptional activation.
Results: We found strong enrichment of H3K4m2 and –m3, near the reported gyk PPRE (gyk -1893) but not in a region proximal (gyk -6085). On the ob locus, we found enrichment of H3K4m2 ~ but not m3 ~ 16kb proximal to the reported transcriptional start site (TSS). Interestingly, there was only background enrichment of H3K4m2 near the ob TSS. After treatment with rosiglitazone (1uM) for 6 hours, we found that at gyk -1893 H3K4m2 decreased but H3K4m3 was increased. In contrast, at ob -16kb H3K4m2 decreased and H3K4m3 remained low.
Conclusion: Based on the above data we hypothesize that there may be differential recruitment of histone modifying enzymes in response to rosiglitazone at gyk vs. ob response elements. Further work is focused on identifying which histone demethylases or histone methyltransferases are involved in this regulation.
Nothing to Disclose: AS, VPL, FEW
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