This dissertation contains four divisions represented in three chapters. The first chapter introduces transcription factor Rev-Erb, a transcriptional repressor belonging to the superfamily of nuclear receptors. Chapter 1 then continues with the physiological roles and functional mechanisms of enhancers. These are DNA regulatory elements key to orchestrate temporal and spatial expression of genes. Our work on Rev-Erb in macrophages, described in the second chapter, provides evidences of a novel transcription regulatory mechanism at enhancer elements. We identified novel target genes of Rev-Erb, such as matrix metalloproteases 9 (mmp9) and fractalkine receptor (Cx3cr1), which are negatively regulated by Rev-Erb from distal enhancer elements. In fact, genome-wide binding analysis shows that most Rev-Erb localizes to putative macrophage enhancers. A recent discovery demonstrated that RNAs are actively transcribed at enhancers on a genomic- scale. In line with this finding, Rev-Erb represses RNA transcription at enhancers. The mechanism is similar to its action at promoters by inhibition of RNA polymerase II recruitment and deacetylation of histones. Given that both enhancer-RNAs (eRNAs) and mRNAs are negatively regulated by Rev-Erb, we further test whether eRNA is a functional intermediate for target gene regulation. Using Mmp9 as a model, targeting its eRNA with RNA interference decreases mRNA expression. Furthermore, in Rev-Erb deficient macrophages, both Mmp9 eRNA and mRNA are derepressed; but targeting eRNA is sufficient to reverse Mmp9 mRNA derepression. Our finding suggests a functional role for enhancer-RNA. When acting through enhancers, Rev-Erb regulates target genes by repressing transcription at enhancers as a mechanistic intermediate. The last chapter explores ways to utilize the enormousness of genome-scale data from human populations to study functions of transcription regulatory elements relevant to diseases through forward genetics. Understanding mechanisms of transcriptional regulation, including discoveries of functional and targetable intermediates such as enhancer- RNAs, suggests new methods of disease detection and treatment

Transcription factor programs mediating the immune response to coronavirus disease 2019 (COVID-19) are not fully understood. Capturing active transcription initiation from cis-regulatory elements such as enhancers and promoters by capped small RNA sequencing (csRNA-seq), in contrast to capturing steady-state transcripts by conventional RNA-seq, allows unbiased identification of the underlying transcription factor activity and regulatory pathways. Here, we profile transcription initiation in critically ill COVID-19 patients, identifying transcription factor motifs that correlate with clinical lung injury and disease severity. Unbiased clustering reveals distinct subsets of cis-regulatory elements that delineate the cell type, pathway-specific, and combinatorial transcription factor activity. We find evidence of critical roles of regulatory networks, showing that STAT/BCL6 and E2F/MYB regulatory programs from myeloid cell populations are activated in patients with poor disease outcomes and associated with COVID-19 susceptibility genetic variants. More broadly, we demonstrate how capturing acute, disease-mediated changes in transcription initiation can provide insight into the underlying molecular mechanisms and stratify patient disease severity.