Quantitative analysis of promoter methylation in exfoliated epithelial cells isolated from breast milk of healthy women
EPIGENETICS
Authors: Wong, Chung M.; Anderton, Douglas L.; Smith-Schneider, Sallie; Wing, Megan A.; Greven, Melissa C.; Arcaro, Kathleen F.
Abstract
Promoter methylation analysis of genes frequently silenced in breast cancer is a promising indicator of breast cancer risk, as these methylation events are thought to occur long before presentation of disease. The numerous exfoliated epithelial cells present in breast milk may provide the breast epithelial DNA needed for detailed methylation analysis and assessment of breast cancer risk. Fresh breast milk samples and health, lifestyle and reproductive history questionnaires were collected from 111 women. Pyrosequencing analysis was conducted on DNA isolated from the exfoliated epithelial cells immunomagnetically separated from the total cell population in the breast milk of 102 women. A total of 65 CpG sites were examined in six tumor suppressor genes: PYCARD (also known as ASC or TMS1), CDH1, GSTP1, RBP1 (also known as CRBP1), SFRP1 and RASSF1. A sufficient quantity of DNA was obtained for meaningful analysis of promoter methylation; women donated an average of 86 ml of milk with a mean yield of 32,700 epithelial cells per ml. Methylation scores were in general low as expected of benign tissue, but analysis of outlier methylation scores revealed a significant relationship between breast cancer risk, as indicated by previous biopsy and methylation score, for several CpG sites in CDH1, GSTP1, SFRP1 and RBP1. Methylation of RASSF1 was positively correlated with women's age irrespective of her reproductive history. Promoter methylation patterns in DNA from breast milk epithelial cells can likely be used to assess breast cancer risk. Additional studies of women at high breast cancer risk are warranted.
The SR proteins SF2 and RBP1 regulate triglyceride storage in the fat body of Drosophila
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Authors: Bennick, Ryan A.; Nagengast, Alexis A.; DiAngelo, Justin R.
Abstract
In Western societies where food is abundant, these excess nutrients are stored as fats mainly in adipose tissue. Fats are stored in structures known as lipid droplets, and a genome-wide screen performed in Drosophila cells has identified several genes that are important for the formation of these droplets. One group of genes found during this screen included those that regulate mRNA splicing. Previous work from our lab has identified some splicing factors that play a role in regulating fat storage; however, the full complement of splicing proteins that regulate lipid metabolism is still unknown. In this study, the levels of a number of serine-arginine (SR) domain containing splicing factors (RSF1, RBP1, RBP1-like, SF2 and Srp-54) were decreased using RNAi in the adult fat body to assess their role in the control of Drosophila metabolism. Decreasing SF2 and RBP1 showed increased triglycerides, while inducing RNAi towards RSF1, RBP1-Like and Srp-54 had no effect on triglycerides. Interestingly, the increased triglyceride phenotype in the SF2-RNAi flies was due to an increase in the amount of fat stored per cell while the RBP1-RNAi flies have more fat cells. In addition, the splicing of the beta-oxidation enzyme, CPT1, was altered in the SF2-RNAi flies potentially promoting the increased triglycerides in these animals. Together, this study identifies novel splicing factors responsible for the regulation of lipid storage in the Drosophila fat body and contributes to our understanding of the mechanisms, which influence the regulation of fat storage in adipose-like cells. (C) 2019 Elsevier Inc. All rights reserved.