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Editorial overview: Cell nucleus: The nucleus:

a dynamic organelle

Michael P Rout and Gary H Karpen

Current Opinion in Structural Biology 2014, 28:iv–vii

For a complete overview see the Issue

Available online 4th June 2014

http://dx.doi.org/10.1016/j.ceb.2014.05.005

0955-0674/Published by Elsevier Ltd.

The nucleus is perhaps the defining feature of ‘eukaryotes’ (Greek ‘eu-’

Michael P Rout

(with) ‘-karyon’ (kernel, nucleus)), and the emperor of all eukaryotic

Laboratory of Cellular and Structural Biology,

The Rockefeller University, New York, organelles in terms of scale and complexity of organization. It is, of course,

NY 10065, United States the repository for almost all genomic information, encoded in DNA

e-mail: [email protected]

sequences wrapped into chromatin as discrete polymer packages termed

chromosomes, surrounded in turn by a double-membrane nuclear envelope

Michael P Rout received his Ph.D. in 1990

(NE). Our investment in genomics in humans and other eukaryotes has been

from the Medical Research Council

rightly huge; however, the return has been slow, as we discovered that

Laboratory of Molecular Biology at the

University of Cambridge, working with John sequence assemblies alone reveal little about how genetic information is

Kilmartin. He joined Rockefeller in 1990 as a utilized. Instead, genome functions are greatly influenced by processes that

postdoctoral fellow in Gu¨ nter Blobel’s control nuclear and chromosome architecture and dynamics — chromatin

laboratory and was appointed assistant

factors and modifications, epigenetic mechanisms, and pathways for con-

professor and head of laboratory in 1997 and

trolling transport of molecules into and out of the nucleus.

professor in 2008.

Gary H Karpen Moreover, the genome is not just a computer that reads software — rather, in

Life Sciences Division, Lawrence Berkeley

the nucleus we cannot separate the control system from the machinery. For

National Lab, and Department of Molecular

example, DNA not only provides the genetic code but also forms the

and Cell Biology, University of California at

dynamic scaffold for nuclear structures, and the way chromatin packaging

Berkeley, Berkeley, CA 94720, United States

e-mail: [email protected] proteins are modified can also be part of the heritable code. A huge portion of

the eukaryotic cellular machinery is involved in nuclear maintenance and

Gary H Karpen is the Director of Berkeley Lab function, such that likely over one-third of all proteins, and almost all RNA

Life Sciences Division. He received his PhD

and DNA, make the nucleus an integral part of their lifestyle in the cell.

from the University of Washington,

Therefore, if we ever want a full return on our investment, we must now

Department of Genetics, in 1987. He

devote resources to fully understand how the nucleus functions as an

performed postdoctoral work with Allan

organelle, actively manages the hierarchical organization, expression, and

Spradling at the Carnegie Institute, and was

on the faculty at The Salk Institute (La Jolla) communication across the genome, in both healthy and diseased cells. The

from 1991 to 2003, before moving to LBNL Opinions here reflect this changing view of the nucleus as an organelle; not a

and UC Berkeley. passive repository for genetic information, but an active, dynamic super-

structure whose processes dictate how that information is organized,

accessed and used.

The Opinions presented here consider recent findings on the spatial and

temporal organization of chromatin, the structure and dynamic organization

of the nuclear periphery, and the mechanisms and regulation of nucleocy-

toplasmic exchange. As reflected in these various articles, our understanding

of the architecture of the nucleus, specifically the 3D localization of genomic

regions, whole chromosome and sub-chromosome domains, and nuclear

bodies or ‘factories’ associated with transcription, replication and DNA

repair, has advanced considerably in recent years due to important techno-

logical advances. They also discuss how many parts of the nucleus

have multiple and dynamically changing functionalities, involving the rapid

Current Opinion in Cell Biology 2014, 28:iv–vii www.sciencedirect.com

Editorial overview: Cell nucleus: The nucleus: a dynamic organelle Rout and Karpen v

re-purposing and re-localization of assemblies and pro- may be ‘moonlighting’ at the spindle during mitosis,

cesses once considered to have only one set of defined when their normal roles are suspended. However, some

roles. degree of nucleocytoplasmic partitioning is maintained in

the spindle even in the absence of traditional nuclear

We start at the beginning of the nucleus’ story, where structures including the NE, indicating that these com-

Devos et al. discuss how valuable insights into the ponents may still retain their segregationist roles even in

nucleus’ evolutionary origins can be gained by compara- this alternate environment.

tive studies. By contrasting nuclear architectures in

familiar fungal and vertebrate ‘model’ systems with those Mitosis, though, is not the only cellular process that can

in evolutionary distant eukaryotic groups, hints are emer- significantly alter and regulate the overall architecture

ging with respect to the prokaryotic ancestors of some key of the nucleus. Nuclear morphology can differ strik-

nuclear components. The nucleus appears to have formed ingly among the eukaryotes, and even between differ-

as an extension of an evolving internal membrane system ent tissues in a single organism. As discussed in Jevtic´

along with the Golgi and ER, first as an open reticulum et al., structural components of the nuclear periphery,

surrounding the chromatin and later developing a traf- such as the lamina and associated assemblies, play a

ficking system that sealed the nucleus and differentiated major role in determining nuclear shape, as do forces

its contents from the cytoplasm. The ‘protocoatomer acting on the nucleus from the cytoplasm. However,

hypothesis’ suggests that this process involved the repur- much of how the nucleus maintains its morphology and

posing of early membrane coating complexes into the volume is still not well understood, though clearly it is

structural and trafficking mechanisms that defined these an exquisitely controlled process, and numerous disease

different organelles. Evidence for this idea can be found states seem intimately linked to alterations in nuclear

buried in the heart of the nuclear pore complex (NPC), morphology.

the sole mediator of trafficking across the NE, and thus

between the nucleus and cytoplasm. Trafficking is One of the more surprising findings in recent years has

mediated by a host of protein transport factors that been the demonstration that forces generated by cyto-

recognize cognate cargoes and chaperone them through plasmic skeletal components, such as microtubules and

the NPC. The NPC and transport factors are surprisingly actin, can be transmitted to inside the nucleus and act to

well conserved across all eukaryotes and contain the regulate nuclear positioning and meiotic chromosome

signature structures of coating complexes, supporting movements/pairing. Luxton and Starr review recent find-

the protocoatomer idea and indicating that once its basic ings about LINC complexes, composed of SUN/KASH

architectures and mechanisms were defined, they have proteins, that together span the nuclear envelope and

remained stable through the last billion years of eukar- thereby directly link cytoplasmic force generation to

yotic evolution with only some lineage-specific areas of nuclear dynamics. In particular, new research has shown

divergence. By contrast, the nuclear lamina — the scaf- that the KASH family of proteins, which are located in the

fold that stabilizes the NE and anchors key regions of outer nuclear envelope, interact with a wide spectrum of

chromatin to the nuclear periphery — varies considerably cytoplasmic elements, including microtubule motors,

among the major eukaryotic lineages, though the reasons actin and myosin, intermediate filaments, and RanGAP.

for this evolutionary complexity and architectural flexi- These new insights hold promise for a deeper molecular

bility remain unclear. understanding of LINC complexes and force generation/

mechanics in the nucleus, information that will contribute

The spindle is another example of a nuclear-associated to diagnosis and treatment of human diseases associated

structure with complex evolutionary origins that may with LINC complex mutations, such as autism, hearing

have involved centriole-like systems that organized both loss and cancer.

the flagellum and spindle. These later duplicated into two

separate structures, specialized for organizing either the The NPCs stand at the border of the nucleus and are seen

flagellum/cilia or spindle, the latter retaining a core set of as its ‘gatekeepers’, being perhaps the most prominent

proteins involved in building and stabilizing microtubules NE components. The major roles NPCs play in both the

as well as motors to manipulate chromosome separation flow of information into and out of the nucleus, and in the

during cell division. Indeed, the precise composition of regulation of gene expression, are the focus of three

the spindle, and its structural and functional relationship Opinion articles. Mor et al. discuss the different factors

with other nuclear-associated structures, is a matter of that mediate each of the multiple transport pathways that

both ongoing debate and active research. As Schweizer import and export macromolecules across the NE. A new

et al. describe, a number of proteins more traditionally understanding is emerging into how defects at various

associated with other roles in the nucleus may also aug- levels of the transport machinery — from the signals in

ment the microtubule scaffold of the spindle with a cargos being recognized by transport factors, through the

dynamic ‘spindle matrix’. This includes components transport factors themselves, to the NPC itself — are at

normally associated with the NPC and lamina, which the root of many of the most important viral diseases and

www.sciencedirect.com Current Opinion in Cell Biology 2014, 28:iv–vii

vi Cell nucleus

cancers. Excitingly, new drugs that target these defects nuclear morphology and genome organization, and con-

are being developed and show tremendous promise. tributes to human disease states.

However, a still poorly understood aspect of nuclear The mysterious genomic domain known as heterochro-

trafficking is how membrane proteins, synthesized in matin shows strong preferences for associations with the

the cytoplasmic endoplasmic reticulum, are moved to nuclear lamina, as well as the nucleolus. Padeken and

the inner membrane of the NE. Laba et al. describe Heun review recent literature that provides new insights

recent progress in this area that has finally begun to bear into how these associations and dynamics are regulated. It

fruit. Although signal mediated trafficking is involved in is now clear that multiple factors and pathways are

membrane protein transport to the inner NE, it is a involved in tethering heterochromatin to the lamina

complex process that involves the integration of multiple and the nucleolus, including transcription factors, histone

sorting signals, of which the canonical nuclear localization modifications, DNA sequences and non-coding RNAs.

signals form only one part. Moreover, the NPC is emer- Interestingly, localization to the lamina and nucleolar

ging as a key platform for the organization of multiple sub-compartments is usually but not always correlated

functionalities, including RNA processing and proofread- with transcriptional silencing, and the authors propose

ing, mitotic control, DNA repair, and gene expression. that expression fate may be accomplished by a combi-

Indeed, though long a subject of speculation, recent nation of localization and pre-existing chromatin states.

progress discussed by Ptak et al. has started to reveal The challenge now is to dissect the relative roles these

the crucial and direct role the NPC plays in organizing different factors and pathways play in regulating hetero-

chromatin architecture and gene regulation. Individual chromatin associations, and how they impact heterochro-

NPC components, spread throughout the structure of the matin and other nuclear functions.

NPC and not just (as expected) localized to its nuclear

periphery, interact directly with assemblies on chromatin Various versions of Chromosome Conformation Capture

and the transcriptional machinery, regulating chromatin technologies (i.e. HiC), combined with imaging/FISH,

structure, gene transcription and subsequent transcript have contributed to a deeper understanding of how

processing. The NPC is thus emerging as an archetypal nearby and distant genomic regions interact in the 3D

example of how there can be multiple functionalities for nucleus, how they change dynamically over time and in

each nuclear structure, and it is expected that the study of different cell types, and the roles of such associations in

the underlying mechanics of these different functional- nuclear functions. It is now clear that interphase chromo-

ities will yield important insights into the underlying somes are folded in a hierarchical manner — the highest

causes of the numerous NPC-associated diseases. frequencies of association are local, which fold into sur-

prisingly conserved 1 Mb Topologically-Associated-

Moving to the inside of the nuclear envelope, the nuclear Domains (TADs), which in turn associate to form whole

lamina is a fascinating structure composed of a meshwork chromosome domains. Nicodemi and Pombo describe

of lamins and associated proteins, some of which are how quantitative modeling approaches pioneered in phy-

embedded in the inner envelope and cross the lamina sics research can be applied to improve our understanding

(e.g. the Lamin B receptor, LBR). The lamina has been of chromosome topology, and propose one model (Strings

well known for its role in providing physical support for & Binders Switch) that can recapitulate the observed

the nuclear envelope, and for its role in human diseases hierarchies and dynamics from fundamental biophysical

(laminopathies) and aging. Amendola and van Steensel properties. Whether or not this particular model stands

discuss recent studies revealing that the lamina plays the test of time, it is clear that integration of biophysical

important roles in regulating genome functions and approaches and models will ultimately reveal the basic

organization, and displays unexpected dynamics. principles responsible for initiating and accomplishing

Methods aimed at identifying specific genome-lamina interphase chromosome architecture, and how these pro-

interactions (ChIP-seq and DAM-ID) show that associ- cesses impact nuclear functions. For example, Phillips-

ations between genomic regions and the lamina (known Cremins discusses how genome folding, revealed through

as Lamina-Associated-Domains, or LADs) are regulated CCC analyses, has impacted our understanding of cellular

by a combination of DNA sequence and post-translational differentiation in mammals, focusing on how genome

histone modifications, and correlations with RNA-seq architecture changes in the transitions from embryonic

data confirm a general trend for transcriptional silencing stem cells, to pluripotent cells, to terminally-differen-

within LADs. Most surprisingly, these studies reveal that tiated cells.

LADs are dynamic, and change during differentiation and

development. As reviewed in Wong et al., high-through- One of the most intriguing aspects of nuclear architecture is

put proteomic approaches have identified many nuclear the compartmentalization of different functions, such as

lamina and envelope proteins (NETs) that are differen- transcription, replication and DNA repair, into nuclear

tially expressed in different cell types. Recent results are bodies or ‘factories’ that lack restraining membranes. Slee-

revealing how differential regulation of NETs influences man and Trinkle-Mulcahy discuss recent and exciting

Current Opinion in Cell Biology 2014, 28:iv–vii www.sciencedirect.com

Editorial overview: Cell nucleus: The nucleus: a dynamic organelle Rout and Karpen vii

developments in our understanding of how nuclear sub- two years for studying actin polymerization in vivo hold

compartments, including nucleoli, Cajal bodies, and spli- great promise for future insights into the roles nuclear actin

cing speckles are formed and dynamically regulated, and and associated motors/factors play in nuclear dynamics.

how their dysfunction impacts human health. Although

there is ample evidence for self-assembly of some nuclear In summary, the recent researches summarized in these

bodies (such as nucleoli), we need to focus efforts on reviews demonstrate that we are in the middle of a

understanding how most nuclear bodies are formed and ‘revolution’ in our thinking about nuclear structure and

regulated. function. Gone are the days when we can conceptualize

nuclear events as simply requiring recognition of linear

Although it is clear that genomic regions move within the ‘two dimensional’ DNA/chromatin targets by macromol-

interphase nucleus, for example during the formation of ecular complexes such as RNA and DNA polymerases.

DNA repair centers or transcription ‘factories’, our un- Instead, we now recognize that nuclear functions are

derstanding of the molecules and mechanisms respon- accomplished in four dimensions, requiring spatial and

sible for mobility, and whether movements are active (i.e. temporal coordination and clustering of distant chromo-

directed) or passive (i.e. random), are sparse. The dis- some regions and macromolecular complex activities.

covery of actin fibers, actin-related proteins (ARPs) and Many nuclear structures previously viewed as ‘unifunc-

myosin motors in the interphase nucleus raised the tional’, such as NPCs, are now recognized to regulate

possibility that these genomic regions could move multiple, disparate functions, such as transport and gene

through the nucleus as motor-associated cargoes speeding expression, and can even change location and function

along actin ‘highways’. However, although the existence during the cell cycle. Unlocking the secrets of how

of actin and related complexes in the nucleus is no longer nuclear functions are regulated and impact human disease

questioned, many issues remain concerning their mol- requires that both investigators and funding agencies

ecular nature and dynamics. Hendzel reviews recent embrace the need for more multiscale, comprehensive,

studies that reveal how actin is transported into and and systems-level approaches to dissecting the mysteries

out of the nucleus, that actin can polymerize into fibers of this vital, ever-changing, highly interconnected and

inside the nucleus, and that actin polymerization is complex organelle. The time for a focused dissection of

critical for the retention of a serum response coactivator. this dynamic organelle, and the establishment of a nuclear

The new approaches and systems developed over the last systems biology, is at hand.

www.sciencedirect.com Current Opinion in Cell Biology 2014, 28:iv–vii