CHROMATIN DYNAMICS IN INTERPHASE NUCLEI AND ITS IMPLICATIONS FOR NUCLEAR-STRUCTURE

Translational dynamics of chromatin in interphase nuclei of living Swi ss 3T3 and HeLa cells was studied using fluorescence microscopy and fl uorescence recovery after photobleaching. Chromatin was fluorescently labeled using dihydroethidium, a membrane-permeant derivative of ethid ium bromide. After labeling, a laser was used to bleach small (similar to 0.4 mu m radius) spots in the heterochromatin and euchromatin of c ells of both types. These spots were observed to persist for >1 h, imp lying that interphase chromatin is immobile over distance scales great er than or equal to 0.4 mu m. Over very short times (<1 s), a partial fluorescence recovery within the spots was observed. This partial reco very is attributed to independent dye motion, based on comparison with results obtained using ethidium homodimer-1, which binds essentially irreversibly to nucleic acids. The immobility observed here is consist ent with chromosome confinement to domains in interphase nuclei. This immobility may reflect motion-impeding steric interactions that arise in the highly concentrated nuclear milieu or outright attachment of th e chromatin to underlying nuclear substructures, such as nucleoli, the nuclear lamina, or the nuclear matrix.