As an approach to understanding the composition and structure of transcriptionally regulated genes, we have devised methods for purification of yeast episomal chromatin. One of these, using conventional biochemical methods, had been described previously. We have improved this method by inserting the E. coli lac operator into the plasmid DNA and then using repressor-operator affinity and antibodies to allow partial purification (greater than 50%) in a a single day. We have now inserted several transcriptionally regulated genes into the plasmid and will purify chromatin and determine its structure when the genes are either active or repressed. One of these genes, HSP26, appears to be regulated in identical fashion in the multicopy plasmid environment and as a single copy in the yeast genome. The role of chromatin structure in transcriptional regulation has been studied in model systems. An investigation of transcriptional regualtion has been studied in model systems. An investigation of transcription of the 5S rRNA gene in Xenopus oocyte extracts suggests that initiation is blocked by histones but elongation is not. Studies of thermal untwisting of DNA show that DNA is constrained on the surface of nucleosomes containing chicken histones and relatively unconstrained when associated with yeast histones. Messenger RNA for a collagen gene is expressed only in cells of the mesenchyme lineage during sea urchin embryogenesis. The protein coded by the gene was localized immunologically to the primary mesenchyme cells which elaborate the larval endoskeleton. Other studies of urchin embryogenesis have documented the presence of insulin-like molecules which may function as factors involved in general or specific aspects of development and have led to cloning of the gene for a putative cell adhesion molecule which is expressed in a temporally and spatially specific fashion.
