Towards a comprehensive regulatory map of Mammalian Genomes.

Genome mapping studies have generated a nearly complete collection of genes for the human genome, but we still lack an equivalently vetted inventory of human regulatory sequences. Cis-regulatory modules (CRMs) play important roles in controlling when, where, and how much a gene is expressed. We developed a training data-free CRM-prediction algorithm, the Mammalian Regulatory MOdule Detector (MrMOD) for accurate CRM prediction in mammalian genomes. MrMOD provides genome position-fixed CRM models similar to the fixed gene models for the mouse and human genomes using only genomic sequences as the inputs with one adjustable parameter - the significance p-value. Importantly, MrMOD predicts a comprehensive set of high-resolution CRMs in the mouse and human genomes including all types of regulatory modules not limited to any tissue, cell type, developmental stage, or condition. We computationally validated MrMOD predictions used a compendium of 21 orthogonal experimental data sets including thousands of experimentally defined CRMs and millions of putative regulatory elements derived from hundreds of different tissues, cell types, and stimulus conditions obtained from multiple databases. In ovo transgenic reporter assay demonstrates the power of our prediction in guiding experimental design. We analyzed CRMs located in the chromosome 17 using unsupervised machine learning and identified groups of CRMs with multiple lines of evidence supporting their functionality, linking CRMs with upstream binding transcription factors and downstream target genes. Our work provides a comprehensive base pair resolution annotation of the functional regulatory elements and non-functional regions in the mammalian genomes.

Genome-wide chromatin accessibility analyses provide a map for enhancing optic nerve regeneration.

Retinal Ganglion Cells (RGCs) lose their ability to grow axons during development. Adult RGCs thus fail to regenerate their axons after injury, leading to vision loss. To uncover mechanisms that promote regeneration of RGC axons, we identified transcription factors (TF) and open chromatin regions that are enriched in rat embryonic RGCs (high axon growth capacity) compared to postnatal RGCs (low axon growth capacity). We found that developmental stage-specific gene expression changes correlated with changes in promoter chromatin accessibility. Binding motifs for TFs such as CREB, CTCF, JUN and YY1 were enriched in the regions of the chromatin that were more accessible in embryonic RGCs. Proteomic analysis of purified rat RGC nuclei confirmed the expression of TFs with potential role in axon growth such as CREB, CTCF, YY1, and JUND. The CREB/ATF binding motif was widespread at the open chromatin region of known pro-regenerative TFs, supporting a role of CREB in regulating axon regeneration. Consistently, overexpression of CREB fused to the VP64 transactivation domain in mouse RGCs promoted axon regeneration after optic nerve injury. Our study provides a map of the chromatin accessibility during RGC development and highlights that TF associated with developmental axon growth can stimulate axon regeneration in mature RGC.

Ascending dorsal column sensory neurons respond to spinal cord injury and downregulate genes related to lipid metabolism.

Regeneration failure after spinal cord injury (SCI) results in part from the lack of a pro-regenerative response in injured neurons, but the response to SCI has not been examined specifically in injured sensory neurons. Using RNA sequencing of dorsal root ganglion, we determined that thoracic SCI elicits a transcriptional response distinct from sciatic nerve injury (SNI). Both SNI and SCI induced upregulation of ATF3 and Jun, yet this response failed to promote growth in sensory neurons after SCI. RNA sequencing of purified sensory neurons one and three days after injury revealed that unlike SNI, the SCI response is not sustained. Both SCI and SNI elicited the expression of ATF3 target genes, with very little overlap between conditions. Pathway analysis of differentially expressed ATF3 target genes revealed that fatty acid biosynthesis and terpenoid backbone synthesis were downregulated after SCI but not SNI. Pharmacologic inhibition of fatty acid synthase, the enzyme generating palmitic acid, decreased axon growth and regeneration in vitro. These results support the notion that decreased expression of lipid metabolism-related genes after SCI, including fatty acid synthase, may restrict axon regenerative capacity after SCI.

Gene Co-expression Analysis Indicates Potential Pathways and Regulators of Beef Tenderness in Nellore Cattle.

Beef tenderness, a complex trait affected by many factors, is economically important to beef quality, industry, and consumer's palatability. In this study, RNA-Seq was used in network analysis to better understand the biological processes that lead to differences in beef tenderness. Skeletal muscle transcriptional profiles from 24 Nellore steers, selected by extreme estimated breeding values (EBVs) for shear force after 14 days of aging, were analyzed and 22 differentially expressed transcripts were identified. Among these were genes encoding ribosomal proteins, glutathione transporter ATP-binding cassette, sub-family C (CFTR/MRP), member 4 (ABCC4), and synaptotagmin IV (SYT4). Complementary co-expression analyses using Partial Correlation with Information Theory (PCIT), Phenotypic Impact Factor (PIF) and the Regulatory Impact Factor (RIF) methods identified candidate regulators and related pathways. The PCIT analysis identified ubiquitin specific peptidase 2 (USP2), growth factor receptor-bound protein 10 (GBR10), anoctamin 1 (ANO1), and transmembrane BAX inhibitor motif containing 4 (TMBIM4) as the most differentially hubbed (DH) transcripts. The transcripts that had a significant correlation with USP2, GBR10, ANO1, and TMBIM4 enriched for proteasome KEGG pathway. RIF analysis identified microRNAs as candidate regulators of variation in tenderness, including bta-mir-133a-2 and bta-mir-22. Both microRNAs have target genes present in the calcium signaling pathway and apoptosis. PIF analysis identified myoglobin (MB), enolase 3 (ENO3), and carbonic anhydrase 3 (CA3) as potentially having fundamental roles in tenderness. Pathways identified in our study impacted in beef tenderness included: calcium signaling, apoptosis, and proteolysis. These findings underscore some of the complex molecular mechanisms that control beef tenderness in Nellore cattle.