Table of Contents

Volume 3, Issue 3

  May/June 2012

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LANDES HIGHLIGHTS

Open Access Article

Landes Highlights

Eva M. Riedmann
http://dx.doi.org/10.4161/nucl.20453
PDF




EXTRA VIEWS

Open Access Article

The discrepancy between chromatin factor location and effect

Tineke Lenstra and Frank C.P. Holstege
http://dx.doi.org/10.4161/nucl.19513
Abstract | Full Text | PDF

The influence of chromatin on many cellular processes is well appreciated. Much has been learnt by studying the role of chromatin remodeling and modifying complexes on individual genes. The seemingly straightforward models that inevitably arise from such studies are challenged by genome-wide analyses. Two recent studies in Saccharomyces cerevisiae provide unprecedented coverage of both the genome-wide location and the effect on gene expression for the majority of chromatin factors. Comparison of the overlap between location and expression effects reveals a large disconnect, with on average only 2.5% of occupied genes showing changes in expression. It is also interesting that only 24% of all expression effects are associated with chromatin factor occupancy. The large difference between location and effect likely reflects general properties inherent to regulation of gene expression through chromatin in yeast. Explanations for the discrepancy include gene-specific properties that exert a requirement for certain factors only on specific genes, as well as functional redundancy, whereby loss of a particular factor is compensated by others that function in a distinct but nevertheless compensatory manner. Since the majority of chromatin factor perturbations do show significant effects on specific subsets of genes, this implies the presence of different types of gene-specific properties that determine which chromatin factors a particular gene requires for proper expression. Understanding these gene-specific properties should be the focus of future studies aimed at understanding regulation of gene expression through chromatin.

Divergence of mitotic strategies in fission yeasts

Ying Gu, Candice Yam and Snezhana Oliferenko
http://dx.doi.org/10.4161/nucl.19514
Abstract | PDF

The aim of mitosis is to produce two daughter nuclei, each containing a chromosome complement identical to that of the mother nucleus. This can be accomplished through a variety of strategies, with “open” and “closed” modes of mitosis positioned at the opposite ends of the spectrum and a range of intermediate patterns in between. In the “closed” mitosis, the nuclear envelope remains intact throughout the nuclear division. In the “open” division type, the envelope of the original nucleus breaks down early in mitosis and reassembles around the segregated daughter genomes. In any case, the nuclear membrane has to remodel to accommodate the mitotic spindle assembly, chromosome segregation and formation of the daughter nuclei. We have recently shown that within the fission yeast clade, the mitotic control of the nuclear surface area may determine the choice between the nuclear envelope breakdown and a fully “closed” division. Here we discuss our data and argue that comparative cell biology studies using two fission yeast species, Schizosaccharomyces pombe and Schizosaccharomyces japonicus, could provide unprecedented insights into physiology and evolution of mitosis.

Open Access Article

Nuclear envelope insertion of spindle pole bodies and nuclear pore complexes

Sue L. Jaspersen and Suman Ghosh
http://dx.doi.org/10.4161/nucl.20148
Abstract | Full Text | PDF

The defining feature of eukaryotic cells is the double lipid bilayer of the nuclear envelope (NE) that serves as a physical barrier separating the genome from the cytosol. Nuclear pore complexes (NPCs) are embedded in the NE to facilitate transport of proteins and other macromolecules into and out of the nucleus. In fungi and early embryos where the NE does not completely breakdown during mitosis, microtubule-organizing centers such as the spindle pole body (SPB) must also be inserted into the NE to facilitate organization of the mitotic spindle. Several recent papers have shed light on the mechanism by which SPB complexes are inserted into the NE. An unexpected link between the SPB and NPCs suggests that assembly of these NE complexes is tightly coordinated. We review the findings of these reports in light of our current knowledge of SPB, NPC and NE structure, assembly and function.

A lamin in lower eukaryotes?

Petros Batsios, Tatjana Peter, Otto Baumann, Reimer Stick, Irene Meyer and Ralph Gräf
http://dx.doi.org/10.4161/nucl.20149
Abstract | PDF

Lamins are the major components of the nuclear lamina and serve not only as a mechanical support, but are also involved in chromatin organization, epigenetic regulation, transcription, and mitotic events. Despite these universal tasks, lamins have so far been found only in metazoans. Yet, recently we have identified Dictyostelium NE81 as the first lamin-like protein in a lower eukaryote. Based on the current knowledge, we draw a model for nuclear envelope organization in Dictyostelium in this Extra View and we review the experimental data that justified this classification. Furthermore we provide unpublished data underscoring the requirement of posttranslational CaaX-box processing for proper protein localization at the nuclear envelope. Sequence comparison of NE81 sequences from four Dictyostelia with bona fide lamins illustrates the evolutional relationship between these proteins. Under certain conditions these usually unicellular social amoebae congregate to form a multicellular body. We propose that the evolution of the lamin-like NE81 went along with the invention of multicellularity.

Organization and roles of nucleosomes at mouse meiotic recombination hotspots

Irina Getun, Zhen Wu and Philippe R.J. Bois
http://dx.doi.org/10.4161/nucl.20325
Abstract | Full Text | PDF

Meiotic double strand breaks (DSBs) occur at discrete regions in the genome coined hotspots. Precisely what directs site selection of these DSBs is hotly debated and in particular it is unclear which chromatin features and regulatory factors are necessary for a genomic region to initiate and resolve DSBs as a crossover (CO) event. In human and mouse, one layer of hotspot selection control is a recognition sequence element present at these sites that is bound by the Prdm9 zinc-finger protein. Furthermore, an overall open chromatin structure is thought to be required to allow access of the recombination machinery, and this is often dictated by the packaging of DNA around nucleosomes. We recently defined the nucleosome occupancy maps of four mouse recombination hotspots throughout meiosis. These analyses revealed no obvious dynamic changes in nucleosome occupancy, suggesting an intrinsic nature of recombinogenic sites, yet they also revealed that nucleosomes define zones of exclusion for CO resolution. Here, we discuss new evidence implicating nucleosome occupancy in recombinogenic repair and its potential roles in controlling chromatin structure at mouse meiotic hotspots.

Palmitoylation in the nucleus: A little fat around the edges

Catherine A. Fox and Marc Gartenberg
http://dx.doi.org/10.4161/nucl.20391
Abstract | Full Text | PDF

Many proteins are post-translationally modified by lipid moieties such as palmitoyl or prenyl (e.g., farnesyl) groups, creating functional proteolipids. Lipid modifications share the property of increasing a protein’s hydrophobicity and thus the propensity of that protein to associate with a membrane. These modifications are used to control the localization and activity of membrane-associated proteins. A well-recognized paradigm is farnesylation of the Ras GTPase that helps target this critical signaling protein to the plasma membrane.¹

 

Protein palmitoylation, the addition of a 16-carbon fatty acid side chain to cysteines via a reversible thioester linkage, has received attention because of the relatively recent discovery of a conserved family of palmitoyltransfereases (PATs for protein acyl transferase) that catalyze this modification.^2,3 PATs are integral membrane proteins that contain a catalytic domain with the signature asp-his-his-cys (DHHC) sequence that faces the cytosol. Mammalian cells are estimated to encode over 20 PATs, the sheer number suggesting expansive regulatory roles for palmitoylation in biology. The budding yeast Saccharomyces cerevisiae, where PATs were originally discovered, encodes seven such enzymes. Each shows a distinct cellular localization pattern, with individual PATs enriched in either the endoplasmic reticulum (ER), golgi, vacuole or plasma membranes.⁴ In general, each PAT modifies a distinct set of proteins in vivo.⁵

Palmitoylation helps anchor substrate proteins or protein domains to membranes and in this capacity often influences vesicular trafficking between membranes. For instance, while farnesylation of some forms of Ras is sufficient to anchor Ras to the ER membrane, subsequent palmitoylation is required to target these Ras proteins to the plasma membrane.³ Similarly, trafficking of the integral membrane protein yeast chitin synthase from the ER to the plasma membrane requires palmitoylation.⁶ As these two examples show, palmitoylation regulates the specificity of protein-membrane interactions in the cytosol.

Proteolipids also act in the nucleus but the list of examples is considerably less extensive. Foremost are the metazoan lamins, a set of intermediate filaments that form a fibrous network at the inner nuclear membrane (INM) called the nuclear lamina.⁷ The mature B-type lamins are farnesylated constitutively whereas lamin A is farnesylated only during maturation. Lamins stabilize the shape of the nucleus while binding specific chromatin domains at the nuclear edge, and thus influence a variety of chromosomal processes. Mutations of the lamin-encoding genes, such as those responsible for Hutchinson-Gilford Progeria Syndrome, yield misshapen nuclei, changes in chromatin distribution and modification, as well as loss of perinuclear heterochromatin.⁷

While the model eukaryote yeast Saccharomyces cerevisiae lacks lamins, significant aspects of chromosome organization at the INM are preserved. In particular, heterochromatin regions are anchored to the INM where assembly of these compact and transcriptionally silent chromatin structures is favored.^8,9 Telomeres, which contain adjacent domains of heterochromatin, are also anchored to the INM, thereby enhancing the genomic stability of chromosome ends.¹⁰ By virtue of the precedent set by proteolipids in the cytosol and lamins in the metazoan nucleus, one might expect that proteolipids also influence the localization, assembly or regulation of these chromosomal landmarks. However, while extensive studies in budding yeast have identified a set of proteins that anchor telomeres and other heterochromatic domains, to date none of these proteins have been shown to function as proteolipids.⁸

Here we summarize our recent study, published in the August 2011 issue of PNAS, in which we show that the conserved nuclear protein Rif1 in the budding yeast Saccharomyces cerevisiae is palmitoylated, and that this modification governs the localization of Rif1 to the INM.¹¹ Rif1 is a telomere binding protein that regulates both telomere length and heterochromatin-mediated gene silencing near telomeres and other internal chromosomal sites. We found that some but not all of these roles for Rif1 are influenced by palmitoylation. We conclude that protein palmitoylation can play a significant role in targeting chromosomal regulatory proteins to the INM.

The function of lamins in the context of tissue building and maintenance

Youngjo Kim, Katie McDole and Yixian Zheng
http://dx.doi.org/10.4161/nucl.20392
Abstract | Full Text | PDF

Lamins are the major structural components of the nuclear lamina found in metazoan organisms. Extensive studies using tissue culture cells have shown that lamins are involved in a wide range of basic cell functions. This has led to the prevailing idea that a given animal cell needs at least one lamin protein for its basic proliferation and survival. However, recent studies have shown that lamins are dispensable for the proliferation and survival of mouse embryonic stem cells (ESC). In contrast to a lack of essential functions in ESCs, certain differentiated cells lacking B-type lamins exhibit increased cell cycle exit rates and enhanced senescence. In this Extraview, we discuss how studies using animal models and cell cultures have begun to reveal cell-type specific functions of lamins in tissue building and homeostasis.




REVIEW

Open Access Article

PML body meets telomere: The beginning of an ALTernate ending?

Inn Chung, Sarah Osterwald, Katharina I. Deeg and Karsten Rippe
http://dx.doi.org/10.4161/nucl.20326
Abstract | Full Text | PDF

The unlimited proliferation potential of cancer cells requires the maintenance of their telomeres. This is frequently accomplished by reactivation of telomerase. However, in a significant fraction of tumors an alternative lengthening of telomeres (ALT) mechanism is active. The molecular mechanism of the ALT pathway remains elusive. In particular, the role of characteristic complexes of promyelocytic leukemia nuclear bodies (PML-NBs) with telomeres, the ALT-associated PML-NBs (APBs), is currently under investigation. Here, we review recent findings on the assembly, structure and functions of APBs. It is discussed how genomic aberrations in ALT-positive cancer cells could result in the formation of APBs and in ALT activity. We conclude that they are important functional intermediates in what is considered the canonical ALT pathway and discuss deregulations of cellular pathways that contribute to the emergence of the ALT phenotype.




RESEARCH PAPERS

Dualistic function of Daxx at centromeric and pericentromeric heterochromatin in normal and stress conditions

Viacheslav M. Morozov, Ekaterina V. Gavrilova, Vasily V. Ogryzko and Alexander M. Ishov
http://dx.doi.org/10.4161/nucl.20180
Abstract | Full Text | PDF

Nuclear structures ND10/PML NBs are linked to multiple processes, including the maintenance of intranuclear homeostasis by sequestering proteins into “nuclear depot.” This function presumes release of proteins from PML NBs and their redistribution to the alternative, supposedly “active” locations, in response to the external stress application. To further investigate this nuclear depot function, we focused on the intranuclear distribution of protein Daxx that in normal conditions is mainly accumulated at PML NBs, and has a minor association with centromeres and pericentromeres (CEN/periCEN). Here we report that application of physiological Heat Shock (HS) changes this balance forcing very robust and reversible accumulation of Daxx on CEN/periCEN heterochromatin.

 

Heterochromatin architecture is essential for the proper orchestration of nuclear processes, while transcription from this part of genome is required for its maintenance. To understand functional consequences of Daxx deposition at CEN/periCEN, we tested for Daxx-dependency of heterochromatin transcription. Depletion of Daxx reduces accumulation of CEN RNA in normal conditions and periCEN RNA after HS application. Searching for the mechanism of Daxx-dependent regulation of heterochromatin transcription, we found that depletion of Daxx decreases incorporation of transcription-associated histone H3 variant, H3.3, into both CEN and periCEN. Surprisingly, HS-induced deposition of Daxx does not further elevate incorporation of H3.3 into CEN/periCEN that remained steady during stress and recovery. Instead, depletion of Daxx leads to HS-induced changes in the balance of epigenetic modifications at heterochromatin, most dramatically elevating levels of active H3K4Me2 modification at periCEN. We propose dualistic function of Daxx-containing complexes at CEN/periCEN: (1) regulation of H3.3 loading in normal conditions, and (2) protection of epigenetic status upon stress-induced accumulation, thus collectively guarding epigenetic identity of CEN/periCEN heterochromatin.

Did the modulation of expression noise shape the evolution of three dimensional genome organizations in eukaryotes?

Kuljeet Singh Sandhu
http://dx.doi.org/10.4161/nucl.20263
Abstract | Full Text | PDF

The pervasive role of distant chromatin interactions in transcriptional regulation is increasingly becoming evident. There is a possibility that the greater diversity in chromatin interactions of a genomic locus could contribute to stochastic variation in its gene expression. However, this issue has not been addressed. Here, I present a few lines of evidence, which suggest that the variation in trans chromatin interactions might occur at the cost of expression noise. Genomic regions with nucleosome depletion, abundant and rapid transcription and with essential gene clusters exhibit relatively fewer trans chromatin interactions in the nucleus. Moreover, loci with greater number of interactions tend to show higher expression noise. Based on these observations, I hypothesize that the three dimensional organization of eukaryotic genomes might have evolved under a selective pressure to minimize the expression noise of essential gene clusters in the nucleus.

Open Access Article

Profilin is associated with transcriptionally active genes

Emilia Söderberg, Viktoria Hessle, Anne von Euler and Neus Visa
http://dx.doi.org/10.4161/nucl.20327
Abstract | Full Text | PDF

We have raised antibodies against the profilin of Chironomus tentans to study the location of profilin relative to chromatin and to active genes in salivary gland polytene chromosomes. We show that a fraction of profilin is located in the nucleus, where profilin is highly concentrated in the nucleoplasm and at the nuclear periphery. Moreover, profilin is associated with multiple bands in the polytene chromosomes. By staining salivary glands with propidium iodide, we show that profilin does not co-localize with dense chromatin. Profilin associates instead with protein-coding genes that are transcriptionally active, as revealed by co-localization with hnRNP and snRNP proteins. We have performed experiments of transcription inhibition with actinomycin D and we show that the association of profilin with the chromosomes requires ongoing transcription. However, the interaction of profilin with the gene loci does not depend on RNA. Our results are compatible with profilin regulating actin polymerization in the cell nucleus. However, the association of actin with the polytene chromosomes of C. tentans is sensitive to RNase, whereas the association of profilin is not, and we propose therefore that the chromosomal location of profilin is independent of actin.

Nuclear actin depolymerisation in transcriptionally active avian and amphibian oocytes leads to collapse of intranuclear structures

Antonina Maslova and Alla Krasikova
http://dx.doi.org/10.4161/nucl.20393
Abstract | PDF

Actin, which is normally depleted in the nuclei of somatic cells, accumulates in high amounts in giant nuclei of amphibian oocytes. The supramolecular organization and functions of this nuclear pool of actin in growing vertebrate oocyte are controversial. Here, we investigated the role of nuclear actin in the maintenance of the spatial architecture of intranuclear structures in avian and amphibian growing oocytes. A meshwork of filamentous actin was not detected in freshly isolated or fixed oocyte nuclei of Xenopus, chicken, or quail. We found that the actin meshwork inside the oocyte nucleus could be induced by phalloidin treatment. Actin polymerisation is demonstrated to be required to stabilize the specific spatial organization of nuclear structures in avian and amphibian growing oocytes. In experiments with the actin depolymerising drugs cytochalasin D and latrunculin A, we showed that disassembly of nuclear actin polymers led to chromosome condensation and their transportation to a limited space within the oocyte nucleus. Experimentally induced ‘collapsing’ of chromosomes and nuclear bodies, together with global inhibition of transcription, strongly resembled the process of karyosphere formation during oocyte growth.