title: High-resolution, high-throughput microscopy analyses of nuclear receptor and coregulator function.
abstract: Steroid nuclear receptors are ligand-dependent transcription factors that have been studied since the early 1960s by principally biochemical and reporter assay approaches. From these studies an elegant and complex model of nuclear receptor transcription regulation has been developed. Inherent to both biochemical and reporter assay approaches is the generation of averaged responses and it is not generally considered that individual cells could exhibit quite varied responses. In some cases, recent microscopic single-cell analyses provide markedly different responses relative to traditional approaches based on population averaging and underscore the need to continue refinement of the current model of nuclear receptor-regulated transcription. While single-cell analyses of nuclear receptor action have been hindered by the predominantly qualitative nature of the approach, high-throughput microscopy is now available to resolve this issue. This chapter demonstrates the utility of high-throughput microscopic analyses of nuclear receptor and nuclear receptor coregulator function. The ability of high-throughput microscopy to generate physiologically appropriate test populations by filtering based on morphological and protein of interest expression criteria is demonstrated. High-resolution, high-throughput microscopy is illustrated that provides quantitative subcellular information for both androgen and estrogen receptors. Efforts are ongoing to develop model systems that provide additional multiplex data and with refined image analyses to achieve true high-content imaging screens.
authors.affiliation: Integrated Microscopy Core, Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, USA.
authors.forename: Valeria
authors.initials: V
authors.lastname: Berno
authors.forename: Cruz A
authors.initials: CA
authors.lastname: Hinojos
authors.forename: Larbi
authors.initials: L
authors.lastname: Amazit
authors.forename: Adam T
authors.initials: AT
authors.lastname: Szafran
authors.forename: Michael A
authors.initials: MA
authors.lastname: Mancini
mesh_terms: D000465:Algorithms; D000818:Animals; D001331:Automation; D004247:DNA / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D005557:Formaldehyde / Q000737:chemistry; D006367:HeLa Cells; D006801:Humans; D008853:Microscopy / Q000379:methods*; D011107:Polylysine / Q000737:chemistry; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry*; D011960:Receptors, Estrogen / Q000378:metabolism; D011987:Receptors, Steroid / Q000737:chemistry*; D015398:Signal Transduction; D014162:Transfection
chemical_list: D018160:Receptors, Cytoplasmic and Nuclear; D011960:Receptors, Estrogen; D011987:Receptors, Steroid; D005557:Formaldehyde; D011107:Polylysine; D004247:DNA