Haider Neena B, Mollema Nissa, Gaule Meghan, Yuan Yang, Sachs Andrew J, Nystuen Arne M, Naggert Jürgen K, Nishina Patsy M
Department of Genetics, Cell Biology, and Anatomy, University of Nebraska Medical Center, 985805 Nebraska Medical Center, Omaha, NE 68198-5805, USA.
Exp Eye Res. 2009 Sep;89(3):365-72. doi: 10.1016/j.exer.2009.04.006. Epub 2009 Apr 18.
The retinal transcription factor Nr2e3 plays a key role in photoreceptor development and function. In this study we examine gene expression in the retina of Nr2e3(rd7/rd7) mutants with respect to wild-type control mice, to identify genes that are misregulated and hence potentially function in the Nr2e3 transcriptional network. Quantitative candidate gene real time PCR and subtractive hybridization approaches were used to identify transcripts that were misregulated in Nr2e3(rd7/rd7) mice. Chromatin immunoprecipitation assays were then used to determine which of the misregulated transcripts were direct targets of NR2E3. We identified 24 potential targets of NR2E3. In the developing retina, NR2E3 targets transcription factors such as Ror1, Rorg, and the nuclear hormone receptors Nr1d1 and Nr2c1. In the mature retina NR2E3 targets several genes including the rod specific gene Gnb1 and cone specific genes blue opsin, and two of the cone transducin subunits, Gnat2 and Gnb3. In addition, we identified 5 novel transcripts that are targeted by NR2E3. While mislocalization of proteins between rods and cones was not observed, we did observe diminished concentration of GNB1 protein in adult Nr2e3(rd7/rd7) retinas. These studies identified novel transcriptional pathways that are potentially targeted by Nr2e3 in the retina and specifically demonstrate a novel role for NR2E3 in regulating genes involved in phototransduction.
视网膜转录因子Nr2e3在光感受器的发育和功能中起关键作用。在本研究中，我们检测了Nr2e3(rd7/rd7)突变体视网膜相对于野生型对照小鼠的基因表达情况，以鉴定调控异常从而可能在Nr2e3转录网络中发挥作用的基因。采用定量候选基因实时PCR和消减杂交方法来鉴定在Nr2e3(rd7/rd7)小鼠中调控异常的转录本。随后利用染色质免疫沉淀试验来确定哪些调控异常的转录本是NR2E3的直接靶点。我们鉴定出了24个NR2E3的潜在靶点。在发育中的视网膜中，NR2E3的靶点包括转录因子，如Ror1、Rorg以及核激素受体Nr1d1和Nr2c1。在成熟视网膜中，NR2E3的靶点包括多个基因，如视杆细胞特异性基因Gnb1、视锥细胞特异性基因蓝视蛋白以及视锥转导素的两个亚基Gnat2和Gnb3。此外，我们还鉴定出了5个受NR2E3靶向的新转录本。虽然未观察到视杆细胞和视锥细胞之间蛋白质的错误定位，但我们确实观察到成年Nr2e3(rd7/rd7)视网膜中GNB1蛋白浓度降低。这些研究确定了视网膜中Nr2e3可能靶向的新转录途径，并特别证明了NR2E3在调节参与光转导的基因方面的新作用。
