IN-VIVO CHROMATIN STRUCTURE OF THE MURINE INTERLEUKIN-5 GENE REGION -A NEW INTACT CELL SYSTEM

The study of chromatin involvement in the regulation of gene expressio n has traditionally required the isolation of nuclei. However, cell fr actionation techniques are subject to redistribution of proteins durin g the isolation procedure, which prevents rigorous physiologically rel evant analysis. To eliminate the need to isolate nuclei and to analyze chromatin structures in vivo in response to agents regulating murine interleukin-5 (IL-5) gene activation, we have established a novel lyso lecithin permeabilized intact cell system for suspension cell types, i n this case T cells. Nuclear integrity of permeabilized cells is demon strated by nuclear transport assays using confocal laser scanning micr oscopy. Results are identical in unstimulated and stimulated T cells, indicating that the chromatin structure after activation is not the re sult of gross alterations in nuclear protein transport properties. Pot ential new IL-5 gene regulatory regions are identified by DNase I hype rsensitivity mapping. Our lysolecithin permeabilized intact cell syste m is amenable to physiologically relevant analysis of responses to sig naling pathways at the level of chromatin, nuclear protein translocati on and possibly other cellular functions in a variety of suspension an d adherent cell types.