Histone Lysine Demethylase 1A Is a Master Regulator of Genes Necessary for Trophoblast Cell Proliferation

Abstract

Histone lysine demethylase 1A is a master regulator of genes necessary for trophoblast cell proliferation.A proper functioning placenta is critical for pregnancy, fetal growth and development and postnatal health. Trophoblast cell proliferation and differentiation is critical for placental development and function. Recently we demonstrated that the histone lysine demethylase KDM1A binds to androgen receptor (AR) in human and sheep trophoblast cells, and targets the same promoter region of vascular endothelial growth factor A (VEGFA), suggesting a role for KDM1A and AR in early placental angiogenesis. The goal of this study was to determine the function of KDM1A during early placental development. We hypothesized that KDM1A regulates genes that are necessary for trophoblast cell proliferation, and early placental development. To this end, both in vitro and in vivo approaches were used in this study. ACH-3P cells (human first trimester trophoblast cells (CT and EVT) fused with the choriocarcinoma cell line AC1-1) were used, and a KDM1A knock out (KO) cell line was generated using CRISPR-Cas 9 based genome editing. KDM1A KO in ACH-3P cells led to significant (P<0.05) reduction in AR and VEGFA. Furthermore, factors important for cell proliferation and trophoblast cell development high mobility group AT-hook 1 (HMGA1), LIN28, and MYC protooncogene (cMYC) were significantly (P<0.05) lower in KDM1A KO ACH-3P cells. Cell proliferation assays revealed a significant (P<0.05) reduction in KDM1A KO ACH-3P cells compared to scramble controls. An in vivo experiment was conducted to demonstrate a role for KDM1A in placental development, using the sheep as a model. Day 9 hatched blastocysts were flushed and infected with a Lenti-CRISPRv2 KDM1A target construct (n=4) to knockout KDM1A specifically in the trophectoderm, or with SC (n=5). Infected embryos were transferred to recipient ewes and embryos were collected at gestational day 16. Data suggests that KDM1A KO in trophoblast cells is necessary for conceptus elongation. Current experiments are ongoing to determine the effects of KDM1A and AR knockdown using shRNA lentiviral target vectors on conceptus elongation and pregnancy. Collectively these results indicate that KDM1A plays a central role in regulating genes necessary for trophoblast cell proliferation.This project was supported by Agriculture and Food Research Initiative Competitive Grant no. 2019-67015-29000 from the USDA National Institute of Food and Agriculture.