Wagner and Valentin were the first to describe the nucleolus in two independent publications in the 1830s. The nucleolus is a nuclear sub-organelle that varies in size and number depending on cell type. The main function of the nucleolus is to synthesize and assemble ribosomes for later transport to the cytoplasm, where translation takes place. The nucleolus has also been found to be involved in cell cycle regulation and cellular stress responses. Example images of proteins localized to the nucleoli can be seen in Figure 1. Of all human proteins, 1314 (7%) have been experimentally shown to localize to nucleoli (Figure 2). A Gene Ontology (GO)-based functional enrichment analysis of the nucleolar proteins shows enriched terms for biological processes related to rRNA processing. Approximately 87% (n=1140) of the nucleolar proteins localize to other cellular compartments in addition to nucleoli, and 33% (n=430) only localize to other nuclear structures. The most common additional localizations except for the nucleoplasm are the cytosol and mitochondria.

UTP6 - A-431

RPF1 - SK-MEL-30


Figure 1. Examples of proteins localized to the nucleoli. UTP6 is suggested to be involved in processing of pre rRNA (detected in A-431 cells). RPF1 is a protein believed to be required for ribosome biogenesis (detected in SK-MEL-30 cells). NIFK is known to localize to the nucleoli, but its function is still unclear (detected in U-2 OS cells).

  • 7% (1314 proteins) of all human proteins have been experimentally detected in the nucleoli by the Human Protein Atlas.
  • 400 proteins in the nucleoli are supported by experimental evidence and out of these 108 proteins are enhanced by the Human Protein Atlas.
  • 1140 proteins in the nucleoli have multiple locations.
  • 302 proteins in the nucleoli show a cell to cell variation. Of these 259 show a variation in intensity and 53 a spatial variation.

  • Nucleolar proteins are mainly involved in rRNA processing.

Figure 2. 7% of all human protein-coding genes encode proteins localized to the nucleoli. Each bar is clickable and gives a search result of proteins that belong to the selected category.

The structure of nucleoli


  • Nucleoli: 1027
  • Nucleoli fibrillar center: 287

The nucleoli are non-membrane enclosed, highly conserved, sub organelles within the nucleus. They are formed around nucleolus organizer regions (NORs) consisting of ribosomal DNA (rDNA) and are structurally organized into three different sub regions; fibrillar center (FC), dense fibrillar component (DFC) and granular component (GC) (Boisvert FM et al, 2007; Scheer U et al, 1999). A selection of proteins localized to the nucleoli, which are suitable as nucleoli markers, can be found in Table 1.

Table 1. Selection of proteins suitable as markers for the nucleoli or its substructures.

Gene Description Substructure
DDX47 DEAD-box helicase 47 Nucleoli
RPF1 Ribosome production factor 1 homolog Nucleoli
UTP6 UTP6, small subunit processome component Nucleoli
NOL10 Nucleolar protein 10 Nucleoli
FTSJ3 FtsJ RNA methyltransferase homolog 3 Nucleoli
UBTF Upstream binding transcription factor, RNA polymerase I Nucleoli fibrillar center

A majority of the nucleolar proteins show staining throughout the whole nucleolar area, while roughly 20% display a more refined staining pattern. The staining of fibrillar centers and/or dense fibrillar component appears as clusters of spots for most cell lines while for others, for example MCF-7 and U-251, only one larger spot is seen. Some proteins localize to the rim of the nucleolus, which is visible as a thin circle around the nucleolus and could be associated to either the GC or the perinucleolar heterochromatin surrounding the nucleolus (Németh A et al, 2011). A recent study suggests that the protein MKI67, localized to nucleoli rim, functions like a surfactant to create non-membranous barriers in the cell. Therefore, proteins with similar staining patterns could have similar function (Cuylen S et al, 2016). MKI67 and other immunofluorescent images of different nucleolar substructures can be seen in Figure 3. During mitosis when transcription shuts down, the nucleoli are disassembled as no ribosome assembly is required. The size of the nucleolus has also been suggested to correlate with the proliferative ability of cells (Derenzini M et al, 2000).

KRI1 - HEK 293


KRI1 - U-2 OS

NOLC1 - HEK 293


MKI67 - U-251 MG

Figure 3. Examples of the morphology of the nucleoli in different cell lines as well as the nucleolar substructures and staining patterns. Immunofluorescent staining of KRI1 in HEK 239, MCF-7 and U-2 OS cells. NOLC1 might play a role in maintaining the structure of the fibrillar center and the dense fibrillar component in the nucleoli. NOLC1 is localized to the fibrillar center (detected in HEK293 cells). UBTF is involved in the activation of RNA polymerase I and is localized to the fibrillar center (detected in U-2 OS cells). MKI67 has been found to maintain mitotic chromosome integrity and is a well-known cellular proliferation marker. MKI67 is localized to the nucleoli rim (detected in U-251 cells). Note that nucleoli rim is currently not a location annotated in the Cell Atlas.

Figure 4. 3D-view of nucleoli in U-2 OS, visualized by immunofluorescent staining of NOP56. The morphology of nucleoli in human induced stem cells can be seen in the Allen Cell Explorer.

The function of nucleoli

The nucleolus is responsible for the synthesis, processing and assembly of ribosomes, a complex process controlled in the nucleolar sub regions; fibrillar center, dense fibrillar component and the granular component (Boisvert FM et al, 2007; Scheer U et al, 1999; Németh A et al, 2011). The border between the FC and the DFC contains proteins from the RNA polymerase I complex and is the region where pre-ribosomal RNA (pre-rRNA) is transcribed from rDNA. The pre-rRNA is later modified by proteins in the DFC followed by assembly of the ribosome subunits in the GC (Scheer U et al, 1999). As is the case for the majority of organelles, the proteome of the nucleolus is dynamic and has been shown to consist of multiple overlapping sets of proteins that are interchanging dependent on the cell state. The need for translational capacity varies with different cell cycle phases, and transcriptional capacity is heavily dependent on the amount of ribosomes available. In addition to being responsible for ribosome assembly, the nucleolus has also been found to comprise proteins involved in cell cycle regulation and cellular stress responses (Boisvert FM et al, 2007; Visintin R et al, 2000).

Several genetic disorders such as Werner, fragile X and Treacher Collins syndrome have been linked to nucleolar proteins (Marciniak RA et al, 1998; Tamanini F et al, 2000; Willemsen R et al, 1996; Isaac C et al, 2000). Moreover, the nucleolar size increases with the cells proliferative ability, suggesting that the nucleoli play an important role in development of cancer and could therefore be a potential target for cancer therapy (Drygin D et al, 2010).

Gene Ontology (GO) analysis of the proteins mainly localized to the nucleoli shows functions that are well in-line with already known functions for the structure. The enriched terms for the GO domain Biological Process are related to the rRNA processing and ribosome assembly (Figure 5a), while enrichment analysis of the GO domain Molecular Function gave enriched results for RNA binding activities (Figure 5b). A list of highly expressed nucleolar proteins are summarized in Table 2.

Figure 5a. Gene Ontology-based enrichment analysis for the nucleolar proteome showing the significantly enriched terms for the GO domain Biological Process. Each bar is clickable and gives a search result of proteins that belong to the selected category.

Figure 5b. Gene Ontology-based enrichment analysis for the nucleolar proteome showing the significantly enriched terms for the GO domain Molecular Function. Each bar is clickable and gives a search result of proteins that belong to the selected category.

Table 2. Highly expressed single localized nucleolar proteins across different cell lines.

Gene Description Average NX
EBNA1BP2 EBNA1 binding protein 2 35
FBL Fibrillarin 34
NOLC1 Nucleolar and coiled-body phosphoprotein 1 30
RSL1D1 Ribosomal L1 domain containing 1 28
NOP56 NOP56 ribonucleoprotein 28
NIFK Nucleolar protein interacting with the FHA domain of MKI67 27
GNL2 G protein nucleolar 2 26
WDR82 WD repeat domain 82 25
NOC2L NOC2 like nucleolar associated transcriptional repressor 25
LLPH LLP homolog, long-term synaptic facilitation 24

Nucleolar proteins with multiple locations

Of the nucleolar proteins identified in the Cell Atlas, approximately 87% (n=1140) also localize to other cell compartments (Figure 6). Of these, 33% (n=430) only localize to other nuclear structures. The network plot shows that the most common locations shared with nucleoli are nucleoplasm, cytosol and mitochondria. Given that the nucleoli are responsible for synthesis and assembly of ribosomes that later are exported to the cytoplasm, many of the proteins localized to both the nucleoli and the cytoplasmic structures are most likely involved in translation. The number of proteins localized to the nucleoli and the nucleoplasm as well as the nucleoli and mitochondria are seen more often than expected with the current distribution of multilocalizing proteins, while nucleolar proteins that additionally localize to vesicles, the Golgi apparatus, the cytosol or the centrosomes are significantly underrepresented. Examples of multilocalizing proteins within the nucleolar proteome can be seen in Figure 7.

Figure 6. Interactive network plot of nucleolar proteins with multiple localizations. The numbers in the connecting nodes show the proteins that are localized to the nucleoli and to one or more additional locations. Only connecting nodes containing more than one protein and at least 0.5% of proteins in the nucleolar proteome are shown. The circle sizes are related to the number of proteins. The cyan colored nodes show combinations that are significantly overrepresented, while magenta colored nodes show combinations that are significantly underrepresented as compared to the probability of observing that combination based on the frequency of each annotation and a hypergeometric test (p≤0.05). Note that this calculation is only done for proteins with dual localizations. Each node is clickable and results in a list of all proteins that are found in the connected organelles.

EXOSC10 - U-2 OS



Figure 7. Examples of multilocalizing proteins in the nucleolar proteome. The examples show common or overrepresented combinations for multilocalizing proteins in the nucleolar proteome. EXOSC10 is known to be involved in multiple RNA processing pathways in the nucleolus, nucleus and the cytoplasm (detected in U-2 OS cells). APTX is known to be involved in DNA repair and is localized to the nucleoplasm and the nucleoli (detected in PC-3 cells). SPATS2L is known to localize to the nucleoli and into cytoplasmic stress granules during oxidative stress but the function is unknown (detected in U-2 OS cells).

Expression levels of nucleoli proteins in tissue

Transcriptome analysis and classification of genes into tissue distribution categories (Figure 8) shows that genes encoding proteins that localize to nucleoli are more likely to either be detected in a single tissue or detected in all tissues, compared to all genes presented in the Cell Atlas. Significantly lower portions of nucleoli-associated genes are detected in some or many of the tissues. Thus, these genes tend to either be ubiqutously expressed, or show a strict tissue-specific expression. ,

Figure 8. Bar plot showing the percentage of genes in different tissue distribution categories for nucleoli-associated protein-coding genes compared to all genes in the Cell Atlas. Asterisk marks a statistically significant deviation (p≤0.05) in the number of genes in a category based on a binomial statistical test. Each bar is clickable and gives a search result of proteins that belong to the selected category.

Relevant links and publications

Thul PJ et al, 2017. A subcellular map of the human proteome. Science.
PubMed: 28495876 DOI: 10.1126/science.aal3321

Andersen JS et al, 2005. Nucleolar proteome dynamics. Nature.
PubMed: 15635413 DOI: 10.1038/nature03207

Boisvert FM et al, 2007. The multifunctional nucleolus. Nat Rev Mol Cell Biol.
PubMed: 17519961 DOI: 10.1038/nrm2184

Cuylen S et al, 2016. Ki-67 acts as a biological surfactant to disperse mitotic chromosomes. Nature.
PubMed: 27362226 DOI: 10.1038/nature18610

Derenzini M et al, 2000. Nucleolar size indicates the rapidity of cell proliferation in cancer tissues. J Pathol.
PubMed: 10861579 DOI: 10.1002/(SICI)1096-9896(200006)191:2<181::AID-PATH607>3.0.CO;2-V

Drygin D et al, 2010. The RNA polymerase I transcription machinery: an emerging target for the treatment of cancer. Annu Rev Pharmacol Toxicol.
PubMed: 20055700 DOI: 10.1146/annurev.pharmtox.010909.105844

Isaac C et al, 2000. Characterization of the nucleolar gene product, treacle, in Treacher Collins syndrome. Mol Biol Cell.
PubMed: 10982400 

Marciniak RA et al, 1998. Nucleolar localization of the Werner syndrome protein in human cells. Proc Natl Acad Sci U S A.
PubMed: 9618508 

Németh A et al, 2011. Genome organization in and around the nucleolus. Trends Genet.
PubMed: 21295884 DOI: 10.1016/j.tig.2011.01.002

Scheer U et al, 1999. Structure and function of the nucleolus. Curr Opin Cell Biol.
PubMed: 10395554 DOI: 10.1016/S0955-0674(99)80054-4

Tamanini F et al, 2000. The fragile X-related proteins FXR1P and FXR2P contain a functional nucleolar-targeting signal equivalent to the HIV-1 regulatory proteins. Hum Mol Genet.
PubMed: 10888599 

Visintin R et al, 2000. The nucleolus: the magician's hat for cell cycle tricks. Curr Opin Cell Biol.
PubMed: 10801456 

Willemsen R et al, 1996. Association of FMRP with ribosomal precursor particles in the nucleolus. Biochem Biophys Res Commun.
PubMed: 8769090 DOI: 10.1006/bbrc.1996.1126