The nucleoli accumulate rRNA genes and are the sites of rRNA synthesis and rRNA assembly into ribosomes. During mitosis, nucleoli dissociate, but nucleolar remnants remain on the rRNA gene loci, forming distinct nucleolar organizer regions (NORs). Little is known about the composition and structure of NORs, but upstream binding factor (UBF) has been established as its master organizer. In this study, we sought to establish new proteins in NORs. Using UBF-Sepharose to isolate UBF-binding proteins, we identified histone H1.2 as a candidate partner but were puzzled by this observation, given that UBF is known to be located predominantly in nucleoli, whereas H1.2 distributed broadly among the chromatins in interphase nuclei. We then examined cells undergoing mitosis and saw that both H1.2 and UBF were recruited into NORs in this state, reconciling the results of our UBF pulldowns. Inhibiting rRNA synthesis in interphase nuclei also induced NOR-like structures containing both UBF and H1.2. When chromosomes were isolated and spread on coverslips, NORs appeared separated from the chromosomes containing both UBF and H1.2. After chromosomes were fragmented by homogenization, intact NORs remained visible. Results collectively suggest that NORs are independent structures and that the linker histone H1.2 is a novel component of this structure.
Keywords: UBF; chromatin; histone; linker histone H1.2; mitosis; nucleolus; nucleolus organizer region (NOR); ribosomal ribonucleic acid (rRNA) (ribosomal RNA).
© 2018 by The American Society for Biochemistry and Molecular Biology, Inc.