The linear DNA in a human cell is compacted over a million fold, so much that, when compacted, 60,000 feet of DNA can fit on the tip of a needle. The first stage of compaction, nucleosome formation, is already known to impact gene regulation. The further compaction of nucleosomes into a chromatin fiber has also been shown to limit accessibility of DNA to DNA binding proteins. Despite the nucleosome crystal structure and decades of biochemical assays, the structure of this fiber is still a matter of debate. There are two competing structures proposed for the chromatin fiber: a two start and a one start model. A major difference between these models is the path of the linker DNA. Computational studies of chromatin structure suggest that details such as the length of linker DNA between nucleosomes can cause changes in the type of structure adopted by the fiber. Experiments using different linker lengths and different solution conditions support this claim and provide evidence in support of each model of the chromatin fiber. No study, however, has unambiguously accounted for the in vivo variability of linker lengths found in natural chromatin. We are developing a protocol that will allow us to assay the structure of chromatin in vivo using yeast DNA to account for the true variability of in vivo nucleosome spacing. We have shown our assay is sensitive to the structure of chromatin and are assaying how that structure differs at different genomic locations. We anticipate that the overall structure of the fiber at each genomic location is important for regulation. Our future aim is to explicitly test this prediction by first using our models to correlate changes in the fiber structure to differences in transcriptional regulation.
Reduced estrogen concentrations following menopause are associated with increases in the incidence of cardiovascular disease. Calmodulin (CaM) is required for the activities of numerous cardiac proteins yet is not sufficiently expressed for its binding targets. We have begun to examine the effects of 17b‐estradiol (E2) on the cardiac network of CaM‐binding proteins. Female rats received sham surgery or ovariectomy, followed by treatment with vehicle or E2 for 2 weeks. E2 treatment in sham animals increases the interaction between endogenous CaM and the cardiac α adrenergic receptor type 1A. Surprisingly, ovariectomy substantially increases this interaction, while E2 replacement now reduces it. To examine the effects of these treatments on the population of cardiac CaM‐binding sites that are unsaturated by endogenous CaM, lysate from left ventricle was subjected to saturating Ca2+ concentration and processed through a CaM sepharose column. Flow through contained CaM‐binding sites saturated by endogenous CaM, while sepharose‐bound fraction represented endogenously unsaturated CaM‐binding sites. The fractions eluted from the CaM sepharose were subsequently used in competitive binding assays using purified CaM and a CaM biosensor. E2 treatment in sham animals increases the number of unsaturated CaM‐binding sites. Ovariectomy further increases this number, while E2 treatment now reduces it. The data indicate that E2 treatment exerts opposing effects in the presence and absence of ovaries on specific endogenous CaM‐target associations and the number of endogenously unsaturable Ca2+‐dependent CaM‐binding sites in the heart. These results suggest that estrogen replacement may have unpredictable functional outcomes.Support or Funding InformationSupported by NIH Grant HL112184 to Quang‐Kim Tran and Grants from the Iowa Osteopathic and Educational Research Funds to Sarah Clayton and Quang‐Kim Tran.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.