THE BEHAVIOR OF THE COILED BODY IN CELLS INFECTED WITH ADENOVIRUS IN-VITRO

The coiled body is a phylogenetically conserved nuclear organelle whos e function is not known. Probes for detection of p80-coilin, an 80 kDa protein enriched in the coiled body, have made possible studies deter mining the behavior of the coiled body during the cell cycle, in proli ferating cells, as well as reports suggesting some relationship of the coiled body to mRNA splicing and to the nucleolus. The objective of t his study is to examine the distribution of p80-coilin and nucleolar p roteins in cells infected with adenovirus in vitro. HeLa cells grown a s monolayers were infected with successive dilutions of type 5 human a denovirus culture and fixed in methanol/acetone at different time poin ts. Single and double indirect immunofluorescence was performed with h uman autoantibodies to p80-coilin, fibrillarin, NOR-90/hUBF, RNA polym erase I, PM-Sd, and To, as well as rabbit polyclonal serum to p80-coil in (R288) and mouse monoclonal antibody to adenovirus 72-kDa DNA-bindi ng protein. Indirect immunofluorescence (IIF) with anti- p80-coilin an tibodies showed that the usual bright dot-like coiled body staining pa ttern was replaced in infected cells by 1-5 clusters of tiny dots at t he periphery of the nucleus. This phenomenon was first detected within 12 h of infection and affected more severely cells with increased len gth and load of infection. Cells subjected to heat shock presented no such alteration. Double IIF showed that cells with abnormal coiled bod y appearance expressed the viral 72-kDa DNA-binding protein. Nucleolar proteins RNA polymerase I and NOR-90/hUBF became associated with the p80-coilin-enriched clusters and were no longer detected in the nucleo lus. Other nucleolar proteins, like PM-Sd and To, remained associated to the nucleolus and were not detected in the newly formed clusters. F ibrillarin had a heterogeneous behavior, being restricted to the nucle olus in some infected cells while in some others it was associated wit h the p80-coilin-enriched clusters. Thus our results showed that in vi tro adenovirus infection induced radical redistribution of nucleolar a nd coiled body constituents into newly formed structures characterized by clusters of tiny dots in the periphery of the nucleus. The fact th at three major proteins involved in rRNA synthesis and processing colo calized with p80-coilin in these clusters may bring additional support to the idea that the coiled body and p80-coilin may be implicated in functions related to the nucleolus.