3 A). The compartmentalization of the nucleus into discrete domains contributes to the complexity of processes involved in gene expression and its regulation. Various detection methods have revealed an increasing number of distinct subnuclear structures, and the characterization of the proteins contained within these domains opens up the possibility to investigate their function. The best characterized compartment to date, the nucleolus, is the site of rRNA synthesis and pre-ribosomal assembly, whereas the functions of most other subnuclear structures are much less clear (reviewed in Matera 1999; Spector 1993; Nickerson et al. 1995). Several classes of subnuclear domains have been observed. Some, such as the nucleolus and coiled bodies, fulfill particular roles in the maturation of processing RNAs, for example snRNPs or rRNA, and were therefore called nuclear factories (Matera 1999). Other Clofoctol nuclear factories, such as the Oct1/PTF/transcription (OPT) domain, constitute a compartment where a specific group of genes is brought together, thereby making transcriptional regulation more efficient (Pombo et al. 1998). Recently, it was shown that at least a subset of promyelocytic leukemia (PML) bodies and the perinuclear compartment (PNC) rapidly accumulate FITC-labeled nucleotides, suggesting that they may be sites of transcriptional activity (Huang et al. 1998; LaMorte et al. 1998). However, it cannot be excluded that the transcripts were initially synthesized elsewhere and then translocated into these compartments. Another domain class is formed by the human polycomb group complex (PcG), which localizes to specific heterochromatic regions, suggesting a role in the constitutive repression of transcription (Saurin et al. 1998). Some nuclear domains are storage compartments, where certain proteins are kept in an inactive or inaccessible form. Regulatory mechanisms, such as phosphorylation, control the release of these proteins into the nucleoplasm, where they assemble into functional units. Prominent members of this class are the speckles, which are considered to be storage compartments for splicing factors (Spector 1993; Misteli and Spector 1998). Moreover, some transcription factors have been shown to localize in discrete dots throughout the nucleus, and it is thought that these may also represent storage compartments since they do not coincide with regions of transcriptional activity (van Steensel et al. 1995). Nuclear factories and storage compartments are dynamically linked to RNA polymerase activity. Speckles change their morphology under the influence of transcriptional inhibitors (Spector et al. 1983; Carmo-Fonseca et al. 1992; Misteli et al. 1997; Nayler et al. 1998b) and coiled bodies change their composition upon transcriptional inhibition and eventually disperse (Carmo-Fonseca et al. 1992; Matera 1999). Recently, it was shown that transcription and replication sites, which are both active during S-phase, are found in distinct and separate subnuclear domains, and it was proposed that overlapping sites are temporally separated. This implicates that a given site is either transcriptionally active or replicates (Wei et al. 1998). Clofoctol Together with the characterization of novel subnuclear domains, these results provide Clofoctol further evidence for the existence of a dynamically regulated nuclear architecture supporting the compartmentalization of the nucleus (Nakayasu and Berezney 1989; Jackson et al. 1993; Ma et al. 1998). We previously identified a nuclear protein, YT521-B, as a 110-kD protein containing an amino-terminally located glutamic acidCrich domain (E-box) and a characteristic glutamic acid/arginineCrich domain (ER-domain) at the carboxy-terminal end (Hartmann et al. 1999). ER-domain proteins comprise a growing number of molecules and several family members are involved in RNA metabolism (Hartmann et al. 1999). Moreover, it was suggested that ER repeat proteins may contribute to the development of neurodegenerative diseases (Assier et al. 1999). Transiently expressed EGFP-YT521-B fusion proteins localized to the nucleus and displayed a characteristic pattern Rabbit Polyclonal to AIBP of nuclear bodies, which varied in number and size. Clofoctol Furthermore, transient expression of YT521-B modulated splicing of reporter minigenes in a dose dependent manner. Using immunoprecipitation and yeast two-hybrid experiments, we have shown that YT521-B interacts with scaffold attachment factor B (SAF-B) and the 68-kD Src substrate associated during mitosis (Sam68; Hartmann et al. Clofoctol 1999). SAF-B forms a ternary complex with RNA polymerase II and SR proteins at the so-called scaffold or matrix attachment regions (SAR/MAR; Nayler et al. 1998a). Sam68, an RNA binding.