The nucleolus directly regulates p53 export and degradation

The correlation between stress-induced nucleolar disruption and abrogation of p53 degradation is evident after a wide variety of cellular stresses. This link may be caused by steps in p53 regulation occurring in nucleoli, as suggested by some biochemical evidence. Alternatively, nucleolar disruption also causes redistribution of nucleolar proteins, potentially altering their interactions with p53 and/or MDM2. This raises the fundamental question of whether the nucleolus controls p53 directly, i.e., as a site where p53 regulatory processes occur, or indirectly, i.e., by determining the cellular localization of p53/MDM2-interacting factors. In this work, transport experiments based on heterokaryons, photobleaching, and micronucleation demonstrate that p53 regulatory events are directly regulated by nucleoli and are dependent on intact nucleolar structure and function. Subcellular fractionation and nucleolar isolation revealed a distribution of ubiquitylated p53 that supports these findings. In addition, our results indicate that p53 is exported by two pathways: one stress sensitive and one stress insensitive, the latter being regulated by activities present in the nucleolus.     

Nucleolus: the fascinating nuclear body

Nucleoli are the prominent contrasted structures of the cell nucleus. In the nucleolus, ribosomal RNAs are synthesized, processed and assembled with ribosomal proteins. RNA polymerase I synthesizes the ribosomal RNAs and this activity is cell cycle regulated. The nucleolus reveals the functional organization of the nucleus in which the compartmentation of the different steps of ribosome biogenesis is observed whereas the nucleolar machineries are in permanent exchange with the nucleoplasm and other nuclear bodies. After mitosis, nucleolar assembly is a time and space regulated process controlled by the cell cycle. In addition, by generating a large volume in the nucleus with apparently no RNA polymerase II activity, the nucleolus creates a domain of retention/sequestration of molecules normally active outside the nucleolus. Viruses interact with the nucleolus and recruit nucleolar proteins to facilitate virus replication. The nucleolus is also a sensor of stress due to the redistribution of the ribosomal proteins in the nucleoplasm by nucleolus disruption. The nucleolus plays several crucial functions in the nucleus: in addition to its function as ribosome factory of the cells it is a multifunctional nuclear domain, and nucleolar activity is linked with several pathologies. Perspectives on the evolution of this research area are proposed.     

Treacle and TOPBP1 control replication stress response in the nucleolus

Replication stress is one of the main sources of genome instability. Although the replication stress response in eukaryotic cells has been extensively studied, almost nothing is known about the replication stress response in nucleoli. Here, we demonstrate that initial replication stress–response factors, such as RPA, TOPBP1, and ATR, are recruited inside the nucleolus in response to drug-induced replication stress. The role of TOPBP1 goes beyond the typical replication stress response; it interacts with the low-complexity nucleolar protein Treacle (also referred to as TCOF1) and forms large Treacle–TOPBP1 foci inside the nucleolus. In response to replication stress, Treacle and TOPBP1 facilitate ATR signaling at stalled replication forks, reinforce ATR-mediated checkpoint activation inside the nucleolus, and promote the recruitment of downstream replication stress response proteins inside the nucleolus without forming nucleolar caps. Characterization of the Treacle–TOPBP1 interaction mode leads us to propose that these factors can form a molecular platform for efficient stress response in the nucleolus.     

Charge versus sequence for nuclear/nucleolar localization of plant ribosomal proteins

Ribosomal subunit assembly in the nucleolus is dependent on efficient targeting of ribosomal proteins (RPs) from the cytoplasm into the nucleus and nucleolus. Nuclear/nucleolar localization of a protein is generally mediated by one or more specific stretches of basic amino acids—nuclear/nucleolar localization signals (NLSs/NoLSs). Arabidopsis thaliana RPL23aA has eight putative NLSs/NoLSs (pNLSs/NoLSs). Here we mutated all eight NLS/NoLSs individually and in groups and showed, via transient expression in tobacco cells that nucleolar localization of RPL23aA was disrupted by mutation of various combinations of five or more pNLSs/NoLSs. Mutation of all eight pNLSs/NoLSs, a 50 % reduction in total basic charge of RPL23aA, resulted in a complete disruption of nucleolar localization, however, the protein can still localize to the nucleus. As no individual or specific combination of NoLSs was absolutely required for nucleolar localization, we suggest that nucleolar localization/retention of RPL23aA is dependent on the overall basic charge. In addition to the optimal basic charge conferred by these NoLSs, nucleolar localization/retention of RPL23aA also required a C-terminal putative 26S rRNA binding site. In contrast, in the RPs RPS8A and RPL15A, mutation of just two and three N-terminal pNLSs, respectively, disrupted both nuclear and nucleolar localization of these two RPs, indicating differential signal requirements for nuclear and nucleolar localization of the three Arabidopsis RPs RPL23aA, RPL15A and RPS8A.     

Nucleolar topography of nuclei in histologic sections. Application of a nonparametric approach to the study of breast cancer and non-Hodgkin's lymphoma

The position of the nucleolus within a near-convex nuclear profile was defined, without taking stereology into account, in terms of two distances, P and G, where P = pi p2/SN, G = pi 2/SN, p and g being, respectively, the smallest and greatest distances between the center of the nucleolus (considered as a disk) and the superficial nuclear membrane, and SN being the surface area of the nucleus. Nuclear elongation (ND) was defined by the ratio ND = SN/A, where A is the area of the largest inscribed disk. The nucleolus-nucleus ratio RS = Sn/SN (i.e., the surface of the nucleolus over that of the nucleus) describes the size of the nucleolus. A four-class classification of nucleolar topography was then developed from the model of an ellipsoid nuclear profile (1 less than ND less than 2.5), for which the probability that a randomly located nucleolus (with RS less than 0.23) will be classified as central (G less than 2.5 and 0.40 less than or equal to P less than 1) is less than 0.14, as paracentral (G less than 2.5 and 0.23 less than or equal to P less than 0.40) is less than 0.13, as "transversely eccentric" (G less than 2.5 and P less than 0.23) is less than 0.10 and as "longitudinally eccentric" (G greater than or equal to 2.5) is greater than 0.66. This theoretical distribution into four classes was compared to that of nuclear profiles in four cases of breast cancer and in typical cases of immunoblastic, centroblastic and Burkitt's non-Hodgkin's lymphomas. The findings illustrate the ability of this nonparametric method to indicate characteristic nucleolar locations in relation to the number of nucleoli, their size and their nuclear profile elongation.