Trigeminal ganglion neurons comprise three main cell body-size types. This cell size heterogeneity provides an excellent neuronal model to study the cell size-dependent organization and dynamics of the nucleoli, Cajal (coiled) bodies (CBs), and nuclear speckles of pre-mRNA splicing factors, nuclear structures that play a key role in the normal neuronal physiology. We have analyzed the number of nucleoli and CBs and the structural and molecular organization of CBs and nuclear speckles in the three neuronal types by using immunofluorescence with antibodies that recognize nucleoli (fibrillarin), CBs (coilin), and nuclear speckles (snRNPs), confocal microscopy, and electron microscopy. Whereas the mean number of nucleoli per neuron decreases as a function of cell size, the number of CBs per cell significantly increases in large neurons in comparison with the small ones. In addition, large neurons have a higher proportion of CBs associated with the nucleolus. In all neuronal types, CBs concentrate coilin, fibrillarin, snRNPs, and the survival motor neuron protein (SMN). Immunostaining for snRNPs shows small speckle domains and extensive areas of diffuse nucleoplasmic signal in large neurons, in contrast with the large nuclear speckles found in small neurons. Furthermore, flow cytometric analysis shows that all neurons are in the range of diploid cells. These findings indicate that the fusion behavior of nucleoli, the formation of CBs and their relationships with the nucleolus, as well as the compartmentalization of the pre-mRNA splicing machinery, is related to cell body size in the trigeminal ganglion neurons. Because transcriptional activity is a basic determinant mechanism of cell size in diploid cells, we suggest that our findings reflect a distinct transcription-dependent organization of the nucleolus and splicing machinery in the three cell types of trigeminal ganglion neurons.

Nuclear bodies represent a heterogeneous class of nuclear structures. Herein, we describe that a subset of nuclear bodies is highly enriched in components of the ubiquitin-proteasome pathway of proteolysis. We coined the term clastosome (from the Greek klastos, broken and soma, body) to refer to this type of nuclear body. Clastosomes contain a high concentration of 1) ubiquitin conjugates, 2) the proteolytically active 20S core and the 19S regulatory complexes of the 26S proteasome, and 3) protein substrates of the proteasome. Although detected in a variety of cell types, clastosomes are scarce under normal conditions; however, they become more abundant when proteasomal activity is stimulated. In contrast, clastosomes disappear when cells are treated with proteasome inhibitors. Protein substrates of the proteasome that are found concentrated in clastosomes include the short-lived transcription factors c-Fos and c-Jun, adenovirus E1A proteins, and the PML protein. We propose that clastosomes are sites where proteolysis of a variety of protein substrates is taking place.

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