Structure and Function in the Budding Yeast Nucleus
Abstract
Budding yeast, like other eukaryotes, carries its genetic information on chromosomes that are sequestered from other cellular constituents by a double membrane, which forms the nucleus. An elaborate molecular machinery forms large pores that span the double membrane and regulate the traffic of macromolecules into and out of the nucleus. In multicellular eukaryotes, an intermediate filament meshwork formed of lamin proteins bridges from pore to pore and helps the nucleus reform after mitosis. Yeast, however, lacks lamins, and the nuclear envelope is not disrupted during yeast mitosis. The mitotic spindle nucleates from the nucleoplasmic face of the spindle pole body, which is embedded in the nuclear envelope. Surprisingly, the kinetochores remain attached to short microtubules throughout interphase, influencing the position of centromeres in the interphase nucleus, and telomeres are found clustered in foci at the nuclear periphery. In addition to this chromosomal organization, the yeast nucleus is functionally compartmentalized to allow efficient gene expression, repression, RNA processing, genomic replication, and repair. The formation of functional subcompartments is achieved in the nucleus without intranuclear membranes and depends instead on sequence elements, protein–protein interactions, specific anchorage sites at the nuclear envelope or at pores, and long-range contacts between specific chromosomal loci, such as telomeres. Here we review the spatial organization of the budding yeast nucleus, the proteins involved in forming nuclear subcompartments, and evidence suggesting that the spatial organization of the nucleus is important for nuclear function.
THE cell nucleus not only harbors and expresses an organism’s essential genetic blueprint, but also ensures the proper expression, duplication, repair, and segregation of chromosomes while ensuring proper processing and export of messenger and ribosomal RNA (Spector 2003; Taddei et al. 2004b). The dense packing of highly charged molecules (DNA, RNA, histones, nonhistone proteins) in a limited nuclear space was once thought to constrain molecular dynamics, yet we now know that the nucleus is neither grid-locked nor a random jumble (Rouquette et al. 2010). Large rings of chromatin diffuse freely through the nuclear volume in a random diffusive walk (Gartenberg et al. 2004; Neumann et al. 2012), yet the nucleus can maintain functional subcompartments enriched for specific enzymes and chromatin states. Understanding this dichotomy is key to understanding how nuclear organization facilitates nuclear function (reviewed in Taddei et al. 2004b; Mekhail and Moazed 2010; Egecioglu and Brickner 2011; Rajapakse and Groudine 2011; Zimmer and Fabre 2011).
Chromosomes, and the nucleosomal fibers within them, can be thought of as basic structural elements of the nucleus. Long-range chromosome folding is constrained by the physics of polymer dynamics (Klenin et al. 1998; Dekker et al. 2002; Gehlen et al. 2006; Neumann et al. 2012), and the characteristics of the chromosome polymers themselves depend on the folding of the nucleosomal fiber (Rosa and Everaers 2008). Yet chromatin in interphase nuclei is not regularly compacted and is subject to reversible covalent modifications on both DNA and histones within the nucleosomal fiber. Therefore, crucial biophysical properties of long-range chromatin dynamics, such as persistence length, mass density, and diffusion rate, are variable and subject to changes induced by nucleosome remodelers and histone modifiers. Thus, the post-translational modification of histones contributes not only to local chromatin folding, but to the three-dimensional organization of the genome (van Steensel 2011).
A second major factor contributing to nuclear organization is the interaction between chromatin and stable structural elements of the nucleus. In budding yeast, the key structural elements are the nuclear envelope (NE), the nuclear pore complex (NPC), and the nucleolus. The NE encompasses different types of chromatin anchorage sites, including the spindle pole body (SPB), and unique protein components of the inner nuclear membrane that tether heterochromatin, the ribosomal DNA (rDNA), or different types of DNA damage (Akhtar and Gasser 2007; Mekhail and Moazed 2010). The NPC also plays a role in the transient anchoring of activated genes or of DNA damage that cannot be readily repaired by homologous recombination. Finally, long-range interaction of loci in trans, such as the clustering of telomeres or of transfer RNA (tRNA) genes, influences nuclear order. The combination of physical constraints on chromatin movement and protein–protein interactions helps generate nuclear subcompartments that are enriched for specific DNA sequences, factors, and enzymatic activities (Gasser et al. 2004; Rosa and Everaers 2008). How these subcompartments affect nuclear function remains a central topic of research.
The basic principles of nuclear organization can be observed in all eukaryotes from yeast to humans. This allows us to test the functional implications of nuclear organization in a single-celled organism, despite there being species- and tissue-specific nuclear features. With facile genetics, live microscopy, and genome-wide mapping approaches, budding yeast has proven to be extremely useful for testing the functional roles of nuclear structure, as reviewed below.
Acknowledgments
We thank S. Nagai for the repair figure and K. Bystricky for images in Figure 1. We thank V. Dion, C. Horigome, H. Ferreira, M. Oppikofer, and S. Kueng of the Gasser laboratory for a critical reading of the review. S.M.G. acknowledges support of the Novartis Research Foundation, the National Center for Competence in Research, Frontiers-in-Genetics, and the European Union Network Of Excellence Epigenome. A.T. is supported by the French Agence Nationale pour la Recherche and the European Research Council (ERC) under the European Community’s Seventh Framework Programme (FP7/2007-2013)/ERC grant agreement no. 210508.
Footnotes
Communicating editor: J. Boeke