Project description:Conditional knockout of the transcription factor Ronin (Thap11) in retinal progenitor cells (RPCs) results in a profound failure cell proliferation resulting in a hypoplastic adult retina that also suffers from photoreceptor degeneration. The goal of this study was to determine which genes are deregulated in response to loss of Ronin transcription factor activity in the developing retina. We generated Ronin flox/flox (Control) and Chx10-Cre::GFP+/tg; Ronin flox/flox (CKO) mice, in which Ronin loss occurs specifically within RPCs, and performed RNA-Seq analysis of embryonic day E14.5 (E14.5) retinae. Three independent pools of control and Ronin CKO retinae were collected consisting of a minimum of 10 retinae per pool and total RNA was extracted followed by polyA selection, fractionation (200-500 nucleotide range) and generation of cDNA. The resulting DNA was then used for standard Illumina adaptor ligation and sequencing. This experiment revealed decreased expression of a large group of mitochondrial genes including components of the electron transport chain (ETC), which have been recently implicated as direct regulators of the cell cycle.
Project description:Ronin (THAP11), an idiosyncratic DNA-binding protein that evolved from a primordial DNA transposon by molecular domestication, recognizes a hyperconserved promoter sequence through which it controls a variety of developmentally and metabolically essential genes in pluripotent stem cells. However, it remains unclear whether Ronin or related THAP proteins perform similar functions elsewhere in development. Here, we present evidence indicating that Ronin performs a novel function within the nascent heart as it arises from the mesoderm and forms a four-chambered organ. We show that Ronin is vital for cardiogenesis during midgestation through its control of a core set of critical genes. The activity of Ronin coincided with the recruitment of its cofactor, Hcf-1, and the elevation of H3K4me3 levels at specific target genes, suggesting the involvement of an epigenetic mechanism. On the strength of these findings, we propose that Ronin activity during cardiogenesis may offer a template that could be used to understand how important gene programs are sustained across different cell types within a developing organ, such as the heart.
Project description:Conditional knockout of the transcription factor Ronin (Thap11) in retinal progenitor cells (RPCs) results in a profound failure cell proliferation resulting in a hypoplastic adult retina that also suffers from photoreceptor degeneration. The goal of this study was to determine the genes that are transcriptionally regulated by Ronin during retinogenesis. P0 wild type retinae (CD-1 background) were pooled (>10 each) in ice-cold 1X PBS and immediately processed for chromatin extraction, fragmentation and immunoprecipitation using custom antibodies against Ronin G4275 (Dejosez et al., 2010), G4275 preimmune serum or normal rabbit IgG (Santa Cruz, sc-2027). The immunoprecipitated DNA fragments were then sequenced using the Ion Torrent PGM system.
Project description:To analyze cell lineage specific global transcriptional changes during retinogenesis, we sequenced mRNA from developing retina fractioned to Cd73+ photoreceptors and Cd73- other cells, in addition to the whole retina at embryonic period.