Compartmentalization has long been known to have a key role in regulation of cellular processes. By keeping enzymes and regulatory complexes in compartments where the delivery of substrate or exit of product is controlled, competing reactions can occur simultaneously in different parts of the cell. Moreover, spatial confinement facilitates the working of molecules participating in reaction chains and is crucial for coupling unfavourable with energetically favourable chemical reactions. Although in many cases intracellular compartmentalization relies on boundaries imposed by membranes, several non-membrane-bounded compartments exist in eukaryotic cells. One of these, the nucleolus, has recently attracted much attention. The emerging view is that molecular confinement in the nucleolus actively contributes to the control of cellular survival and proliferation.

To be or not to be in the nucleolus

African trypanosomes are unicellular, eukaryotic parasites that live extracellularly in a wide range of mammals, including humans. They have a surface coat, composed of variant surface glycoprotein (VSG), which probably is essential and acts as a defence against general innate immunity and against acquired immunity directed at invariant surface antigens. In effect, the VSG is the only antigen that the host can target, and each trypanosome expresses only one VSG. To counter specific antibodies againts the VSG, trypanosomes periodically undergo antigenic variation, the change to expression of another VSG. Antigenic variation belongs to the general survival strategy of enhanced phenotypic variation, where a subset of ‘contigency’ genes of viruses, bacteria and parasites hypermutate, allowing adaptation to hostile or changing environments. A fundamental feature of antigenic variation is its link with the population dynamics of trypanosomes within the single host. Antigenic variants appear hierachically within the mammalian host, with a mixture of order and randomness. The underlying mechanisms of this are not understood, although differential VSG gene activation may play a prominent part. Trypanosome antigenic variation has evolved a second arm in which the infective metalcyclic population in the tsetse fly expresses a defined mixture of VSGs, although again each trypanosome expresses a single VSG. Differential VSG expression enhances transmission to new hosts, in the case of blood-stream trypanosomes by prolonging infection, and in the metacyclic population by generating diversity that may counter existing partial immunity in reservoir hosts. Antigenic variation employs a huge repertoire of VSG genes. Only one is expressed at a time bloodstream trypanosomes, as a result of transcription being restricted to a set of about 20 bloodstream expression sites ( BES s), which are at chromosome telomeres. Only one BES is active at a time, probably through transcriptional elongation being inhibited in the silent BES s. Although transcriptional switching between BES s can effect a VSG switch, the most prolific switch route involves homologous recombination of deoxyribonucleic acid, usually by the copying of a silent gene into a BES . Hierarchical expression of VSGs may be dictated in part by the different types of locus occupied by VSG genes. The VSG genes expressed int the metacyclic population also occupy telomeric sites, which appear to be derived from BES s but have a simpler structure. Their differential expression is achieved by random transcriptional activation; the detailed story requires direct study of the metacylic stage itself. Available evidence suggests that the VSG originated as a surface receptor, and it can be proposed that a number of selective events have contributed to the evolution of the complex, multisystem phenomenon that antigenic variation has become.

Antigenic variation in trypanosomes: enhanced phenotypic variation in a eukaryotic parasite.

Ribosome biogenesis centres both physically and functionally on the activity of the ribosomal RNA (rRNA) genes. Ribosome assembly occurs co-transcriptionally on these genes, requires the coordinated expression and assembly of many hundreds of proteins and is finely tuned to cell and organism growth. This review presents contemporary understanding of the mode and the means of rRNA gene transcription and how growth factors, oncogenes and tumour suppressors regulate this transcription. It is argued that transcription elongation is a key mechanism regulating rRNA gene transcription. This unorthodox view provides a logical framework to explain the co-transcriptional phase of ribosome biogenesis.

A housekeeper with power of attorney: the rRNA genes in ribosome biogenesis

▪ Abstract Eukaryotic genomes are distributed on linear chromosomes that are grouped together in the nucleus, an organelle separated from the cytoplasm by a characteristic double membrane studded with large proteinaceous pores. The chromatin within chromosomes has an as yet poorly characterized higher-order structure, but in addition to this, chromosomes and specific subchromosomal domains are nonrandomly positioned in nuclei. This review examines functional implications of the long-range organization of the genome in interphase nuclei. A rigorous test of the physiological importance of nuclear architecture is achieved by introducing mutations that compromise both structure and function. Focussing on such genetic approaches, we address general concepts of interphase nuclear order, the role of the nuclear envelope (NE) and lamins, and finally the importance of spatial organization for DNA replication and heritable gene expression.

The Function of Nuclear Architecture: A Genetic Approach

Maturation and Intranuclear Transport of Pre-Ribosomes Requires Noc Proteins

Using Saccharomyces cerevisiae strains with genetically modified nucleoli, we show here that changing parameters as critical as the tandem organization of the ribosomal genes and the polymerase transcribing rDNA, although profoundly modifying the position and the shape of the nucleolus, only partially alter its functional subcompartmentation. High-resolution morphology achieved by cryofixation, together with ultrastructural localization of nucleolar proteins and rRNA, reveals that the nucleolar structure, arising upon transcription of rDNA from plasmids by RNA polymerase I, is still divided in functional subcompartments like the wild-type nucleolus. rRNA maturation is restricted to a fibrillar component, reminiscent of the dense fibrillar component in wild-type cells; a granular component is also present, whereas no fibrillar center can be distinguished, which directly links this latter substructure to rDNA chromosomal organization. Although morphologically different, the mininucleoli observed in cells transcribing rDNA with RNA polymerase II also contain a fibrillar subregion of analogous function, in addition to a dense core of unknown nature. Upon repression of rDNA transcription in this strain or in an RNA polymerase I thermosensitive mutant, the nucleolar structure falls apart (in a reversible manner), and nucleolar constituents partially relocate to the nucleoplasm, indicating that rRNA is a primary determinant for the assembly of the nucleolus.

Frédéric Teulières

Papers

Assembly and functional organization of the nucleolus: ultrastructural analysis of Saccharomyces cerevisiae mutants.