The Nucleolus and the Four Ribonucleoproteins of Translation

Abstract

The classical view of the nucleolus as solely committed to ribosome biosynthesis has been modified by recent studies pointing to additional roles for this nuclear domain. These newly recognized features include the nucleolar presence of several nonribosomal RNAs transcribed by RNA polymerase III, as well as nucleolar roles in gene silencing, cell cycle progression, and cellular senescence. The signal recognition particle (SRP) 1 RNA, and several protein components of the SRP also recently have been detected in the nucleolus. Thus, the large and small ribosomal subunits, the 5S rRNA-ribonucleoprotein complex, and now the SRP, are known to be assembled in or pass through the nucleolus. These findings, together with the recent observations that some transfer RNA precursor molecules and the pretransfer RNA processing enzyme, RNase P, are also found in the nucleolus, raise the possibility that these translational components are congressed in the nucleolus in order to probatively interact with one another, perhaps as a test of proper conformational fit. We hypothesize that such interactions may be an important checkpoint during nucleolar assembly of the translational machinery at steps ranging from the regulation of nascent transcript processing to a possible transient preassembly of the entire translational apparatus.

Human ribonuclease P: Subunits, function, and intranuclear localization

“…The ribonucleoprotein core of human nuclear RNase P has at least 10 distinct proteins associated with H1 RNA+ The next major challenge will be the determination of the minimal subunits sufficient for reconstitution of RNase P activity in vitro and that will set the stage for future work on the structural biology of this riboHuman ribonuclease P 5 nucleoprotein+ This can have general implications in understanding the function of other small nuclear and nucleolar ribonucleoproteins involved in RNA processing+ RNase P and its multiple subunits are spread in the nuclear space of mammalian cells+ Coordination between distinct nuclear compartments and the cytoplasm should take place to ensure RNase P production and accurate processing of tRNA+ The function of RNase P should not be disconnected from that of RNase MRP and thereby from rRNA processing+ The findings that protein synthesis can be coupled to transcription within the nucleus (Iborro et al+, 2001) and that RNase P biosynthesis is linked to that of ribonucleoprotein complexes of translation (Pederson & Politz, 2000) may reveal new roles of this holoenzyme and its subunits in gene transcription, RNA processing, and translation+ …”

“…The biosynthesis of human RNase P should involve the coordination of expression of the genes coding for its RNA and protein subunits+ In addition, the biogenesis of RNase P should be tightly coupled to the translation machinery (Pederson & Politz, 2000)+ Nonetheless, control of expression of a single Rpp subunit may regulate the entire holoenzyme and its activity in tRNA processing+ Examples of regulation mechanisms that modulate RNase P activity or the expression of its proteins in human and yeast cells are described below+…”

“…In situ RNA hybridization analysis and microinjection of labeled H1 RNA into human cell lines demonstrate that this RNA transiently enters the nucleolus before it diffuses in the nucleoplasm (Jacobson et al+, 1997; Table 2)+ The endogenous H1 RNA was also detected in the cytoplasm, the nucleoli, and the perinucleolar compartment (see Wolin & Matera, 1999)+ Because H1 RNA is essential for enzyme activity and exists in the nucleoplasm, in contrast to the S. cerevisiae RNase P RNA that is predominantly found in the nucleolus (Bertrand et al+, 1998;Kendall et al+, 2000;Lewis & Tollervey, 2000), human RNase P may function in this compartment+ The nucleolus of human cells seems to serve as an assembly site for RNase P (Pederson & Politz, 2000)+ In contrast to H1 RNA, several protein subunits, including Rpp14, Rpp29, Rpp38, hPop1, and hPop5, are mainly localized in nucleoli of human cells (Lygerou et al+, 1996b;Jarrous et al+, 1999b;Savino et al+, 1999;van Eenennaam et al+, 2001)+ Rpp29 and Rpp38 are also found in Cajal bodies (Table 2; Jarrous et al+, 1999b), nuclear structures in which transcription and processing machineries preassemble (Lamond & Earnshaw, 1998;Gall, 2000Gall, , 2001)+ Unexpectedly, endogenous Rpp21 is mainly found in the nucleoplasm and a small fraction of it exists in nucleoli ( Table 2; )+ In addition, Rpp21 is not concentrated in Cajal bodies, as are Rpp29 and Rpp38+ These observations indicate that the nuclear localization patterns of Rpp21, as well as H1 RNA, are different from those of other Rpp subunits (Table 2)+ Moreover, protein subunits of RNase P may move rapidly from one compartment to another in the nucleus of human cells+ Thus, fluorescence recovery after photobleaching (FRAP) and fluorescence loss in photobleaching (FLIP) techniques reveal that Rpp29 exchanges quickly between the nucleolus and nucleoplasm of HeLa cells (Chen & Huang, 2001)+ The high mobility of Rpp29 suggests that the intranuclear pool of this protein is not in association with a stationary complex+ A high mobility in the nucleus has also been demonstrated for RNA and nucleolar proteins required for mRNA processing (Politz et al+, 1998;Phair & Misteli, 2000;Chen & Huang, 2001;Pederson, 2001)+ Moreover, nuclear organelles, such as Cajal bodies, are in continuous movement and contact each other in the nucleus (Platani et al+, 2000)+ Therefore, the localization pattern of Rpp29 in the nucleus does not ...…”

“…There is a controversy over the location of active nuclear RNase P and tRNA processing in human cells (see Wolin & Matera, 1999;)+ Some recent observations suggest that human RNase P is a nucleolar enzyme (Jacobson et al+, 1997;Jarrous et al+, 1999b;Lewis & Tollervey, 2000;Pederson & Politz, 2000)+ However, the dynamic and flexible organization of nuclear structures, including the nucleolus, and their building components (Wei et al+, 1998;Wolffe & Hansen, 2001) may imply that nuclear RNase P is not a fixed entity+ The elucidation of the composition, subunit interactions, and intracellular distribution patterns of human RNase P, as summarized above, sheds new light on …”

“…In the nucleus, two issues should be stressed to understand the mechanism of the assembly of active RNase P+ First, H1 RNA and its protein subunits are differentially distributed in several compartments, including nucleoli, Cajal bodies, the perinucleolar compartment and nucleoplasm (Table 2)+ These compartments are transcriptionally specialized sites and their structural integrity is dependent on continuous RNA synthesis (Lamond & Earnshaw, 1998;Pombo et al+, 1999;Cook, 1999;Gall, 2001)+ Accordingly, RNase P is close to sites of active gene transcription+ Second, the rapid mobility of protein subunits of RNase P, at least as shown for Rpp29 (Chen & Huang, 2001), supports the idea that these subunits associate with and dissociate from RNase P in a continuous manner (Lewis & Tollervey, 2000)+ Furthermore, the synthesis of new substrate RNAs, for example, tRNA transcripts, and their interaction with subunits of RNase P, such as Rpp14, Rpp21, Rpp29 , and the La antigen , may play a role in the final formation and reorganization of the catalytic form of RNase P+ However, there is an additional factor that may play a role in the assembly of RNase P+ At least six protein subunits of human RNase P, namely Rpp20, Rpp29, Rpp30, Rpp38, hPop1, and hPop5, have been shown to be associated with RNase MRP (Lygerou et al+, 1996b;Pluk et al+, 1999;van Eenennaam et al+, 1999van Eenennaam et al+, , 2001)+ This overlap in subunit composition raises the possibility that the assembly processes of these two ribonucleoproteins are related+ The conservation of the P3 domains in H1 RNA and MRP RNA (Schmitt, 1999;Frank et al+, 2000) and their common interacting proteins (Pluk et al+, 1999;) support a functional relationship+ This domain is required for the entry of the H1 and MRP RNAs to the nucleolus (Jacobson et al+, 1995, 1997), a major assembly site for ribonucleoproteins (Pederson & Politz, 2000;Grosshans et al+, 2001)+ Moreover, new ideas have been presented in which the assembly of RNase P in the nucleolus is coordinated with ribonucleoproteins of translation, for example, ribosome, SRP, and 5S RNP (Pederson & Politz, 2000)+ Taken together, recent findings support the conclusion that human RNase P ribonucleoproteins are dynamically assembled and recruited to discrete sites of active gene transcription and RNA processing+ Accordingly, RNase P is not restricted to a specific nuclear compartment+…”