The notion of a “plurifunctional” nucleolus is now well established. However, molecular mechanisms underlying the biological processes occurring within this nuclear domain remain only partially understood. As a first step in elucidating these mechanisms we have carried out a proteomic analysis to draw up a list of proteins present within nucleoli of HeLa cells. This analysis allowed the identification of 213 different nucleolar proteins. This catalog complements that of the 271 proteins obtained recently by others, giving a total of ∼350 different nucleolar proteins. Functional classification of these proteins allowed outlining several biological processes taking place within nucleoli. Bioinformatic analyses permitted the assignment of hypothetical functions for 43 proteins for which no functional information is available. Notably, a role in ribosome biogenesis was proposed for 31 proteins. More generally, this functional classification reinforces the plurifunctional nature of nucleoli and provides convincing evidence that nucleoli may play a central role in the control of gene expression. Finally, this analysis supports the recent demonstration of a coupling of transcription and translation in higher eukaryotes.

Productive infection by herpes simplex virus type 1 (HSV-1), which occurs in the host cell nucleus, is accompanied by dramatic modifications of the nuclear architecture, including profound alterations of nucleolar morphology. Here, we show that the three most abundant nucleolar proteins-nucleolin, B23, and fibrillarin-are redistributed out of the nucleoli as a consequence of HSV-1 infection. We show that the amount of nucleolin increases progressively during the course of infection. We demonstrate for the first time that a nucleolar protein, i.e., nucleolin, colocalizes with ICP8 in the viral replication compartments, at the time when viral replication is effective, suggesting an involvement of nucleolin in the HSV-1 DNA replication process. At later times of infection, a granular form of nucleolin localizes to the cytoplasm, in structures that display the characteristic features of aggresomes, indicating that this form of nucleolin is very probably destined for degradation. The delocalization of nucleolin from the nucleoli requires the viral ICP4 protein or a factor(s) whose expression involves ICP4. Using small interfering RNA technology, we show that viral replication requires a high level of nucleolin expression, demonstrating for the first time a direct role for a nucleolar protein in herpes simplex virus biology.Herpes simplex virus type 1 (HSV-1) is a human herpesvirus consisting of an outer envelope, a tegument, a capsid, and a linear double-stranded DNA. Productive infection consists of a highly ordered program of viral gene expression, DNA replication, and virion assembly that leads to the formation of infectious viral progeny and cell death (48). The viral DNA contains at least 80 genes whose expression is sequentially and temporally regulated by complex regulatory mechanisms. Viral genes can be divided into immediate-early, early, and late genes, according to their kinetics of expression. Proteins encoded by immediate-early genes are involved in the regulation of the synthesis of early and late proteins. Proteins encoded by early genes participate in viral DNA replication, and proteins encoded by late genes are mainly the structural components of the viral particles.Viral transcription, DNA replication, assembly of new capsids, and packaging of HSV-1 DNA occur in the host cell nucleus. As a consequence, HSV-1-infected cells undergo a variety of changes including dramatic modifications of the nuclear architecture. The formation of viral replication compartments (VRC), which are the sites of replication, transcription, and encapsidation of HSV-1 genomes, is accompanied by the marginalization of chromatin and the disruption of the nuclear lamina and of PML bodies, as well as by a profound modification of nucleolar morphology (2,15,33,39). Soon after infection, nucleoli increase in size, localize close to the nuclear membrane, and finally become fragmented in small pieces (39,49). In addition, several viral proteins, including ICP0, ICP4, ICP27, US11, and gamma 34.5, localize at least transiently to nucl...