A
High resolution 3D live-cell fluorescence image of a tunneling nanotube
(TNT) (white arrow) connecting two primary mesothelial cells. Scale
bar: 20 μm. B Depiction of a TNT (black arrow) between two cells with scanning electron microscopy. Scale bar: 10 μm. C Fluorescently labeled F-actin (white arrow) present in TNTs between individual HPMCs. Scale bar: 20 μm. D
Scanning electron microscope image of a potential TNT precursor (black
arrowhead). Insert shows a fluorescence microscopic image of
filopodia-like protrusions (white arrowhead) approaching a neighboring
cell. Scale bar: 2 μm.
A tunneling nanotube(TNT) or membrane nanotube is a term that has been applied to cytoskeletal protrusions that extend from the plasma membrane which enable different animal cells to connect over long distances, sometimes over 100 μm between certain types of cells. Tunneling nanotubes that are less than 0.7 micrometers in diameter, have an actin structure and carry portions of plasma membrane between cells in both directions. Larger TNTs (>0.7 μm) contain an actin structure with microtubules and/or intermediate filaments, and can carry components such as vesicles and organelles between cells, including whole mitochondria. The diameter of TNTs ranges from 0.05 μm to 1.5 μm and they can reach lengths of several cell diameters. There have been two types of observed TNTs: open ended and closed
ended. Open ended TNTs connect the cytoplasm of two cells. Closed ended
TNTs do not have continuous cytoplasm as there is a gap junction cap
that only allows small molecules and ions to flow between cells. These structures have shown involvement in cell-to-cell communication, transfer of nucleic acids such as mRNA and miRNA between cells in culture or in a tissue, and the spread of pathogens or toxins such as HIV and prions.
TNTs have observed lifetimes ranging from a few minutes up to several
hours, and several proteins have been implicated in their formation and
inhibition, including many that interact with Arp2/3.
History
3D
live-cell microscopy of immunofluorescent rat PC12 cells demonstrating
tunneling nanotubes. From one of the first published papers to describe
the phenomenon in 2004.
Membrane nanotubes were first described in a 1999 Cell article examining the development of Drosophila melanogaster wing imaginal discs. More recently, a Science article published in 2004 described structures that connected PC12 cells together, as well as other types of cell cultures. This study coined the term "tunneling nanotubes" and also showed that
nanotube formation between cells is correlated with both membrane and
organelle transfer.
Since these publications, more TNT-like structures have been recorded,
containing varying levels of F-actin, microtubules and other components,
but remaining relatively homogenous in terms of composition.
Formation
Several mechanisms may be involved in nanotube formation. These include molecular controls as well as cell-to-cell interactions.
Two primary mechanisms for TNT formation have been proposed. The
first involves cytoplasmic protrusions extending from one cell to
another, where they fuse with the membrane of the target cell.
The other mechanism occurs when two previously connected cells move
away from one another, and TNTs remain as bridges between the two cells.
Induction
Some dendritic cells and THP-1 monocytes
have been shown to connect via tunneling nanotubes and display evidence
of calcium flux when exposed to bacterial or mechanical stimuli.
TNT-mediated signaling has shown to produce spreading in target cells,
similar to the lamellipodia
produced when dendritic cells are exposed to bacterial products. The
TNTs demonstrated in this study propagated at initial speed of 35
micrometers/second and have shown to connect THP-1 monocytes with
nanotubes up to 100 micrometers long.
Phosphatidylserine exposure has demonstrated the ability to guide TNT formation from mesenchymal stem cells (MSCs) to a population of injured cells. The protein S100A4 and its receptor have been shown to guide the direction of TNT growth, as p53 activates caspase 3
to cleave S100A4 in the initiating cell, thereby generating a gradient
in which the target cell has higher amounts of the protein. These findings suggests that chemotactic gradients may be involved in TNT induction.
One study found that cell-to-cell contact was necessary for the formation of nanotube bridges between T cells.
p53 activation has also been implicated as a necessary mechanism for
the development of TNTs, as the downstream genes up-regulated by p53
(namely EGFR, Akt, PI3K, and mTOR) were found to be involved in nanotube formation following hydrogen peroxide treatment and serum starvation. Connexin-43 has shown to promote connection between bone marrow stromal cells (BMSCs) and alveolar epithelial cells, leading to the formation of nanotubes.
Cellular stress by rotenone or TNF-α was also shown to induce TNT formation between epithelial cells. Inflammation by lipopolysaccharides or interferon-γ has shown to increase the expression of proteins related to TNT formation.
Inhibition
While
TNTs have many components, their main inhibitors work by blocking or
limiting actin formation. TNT-like structures called streamers, which
are extremely thin protrusions, did not form when cultured with cytochalasin D, an F-actin depolymerizing compound. A separate study using cytochalasin B found that it impacted TNT formation without the destruction of existing TNTs. Latrunculin-B, another F-actin depolymerizing compound, was found to completely block TNT formation. Blocking CD38, which had been implicated in the release of mitochondria by astrocytes, also significantly decreased TNT formation.
TNFAIP2, also called M-Sec, is known to mediate TNT formation, and knockdown of this protein by shRNA reduced TNT development in epithelial cells by about two-thirds.
Inhibiting Arp2/3 directly resulted in different effects
depending on cell type. In human eye cells and macrophages, blocking
Arp2/3 led to a decrease in TNT formation. However, such inhibition in
neuronal cells resulted in an increase in the amount of cells connected
via TNTs, while lowering the total amount of TNTs connecting cells.
Role in intercellular transfer
Mitochondria
Tunneling nanotubes have been implicated as one mechanism by which whole mitochondria can be transferred from cell to cell. A recent study in Nature Nanotechnology has reported that cancer cells can hijack the mitochondria from immune cells via physical tunneling nanotubes. Mitochondrial DNA
damage appears to be the main trigger for the formation of TNTs in
order to traffic entire mitochondria, though the exact threshold of
damage necessary to induce TNT formation is yet unknown. The maximum speed of mitochondria traveling over TNTs was found to be about 80 nm/s, lower than the measured speed of 100-1400 nm/s of axonal transport of mitochondria; this could be due to the smaller diameter of TNTs inhibiting mitochondrial migration.
In one study, Ahmad et al. used four lines of mesenchymal stem cells, each expressing either a differing phenotype of the Rho-GTPase Miro1; a higher level of Miro1 was associated with more efficient mitochondrial transfer via TNTs.
Several studies have shown, through the selective blockage of TNT
formation, that TNTs are a primary mechanism for the trafficking of
whole mitochondria between heterogeneous cells.
Action Potential
Tunneling
nanotubes have been shown to propagate action potentials via their
extensions of endoplasmic reticulum that propagate Ca2+ influx through active diffusion.
Virus
Many
viruses can transfer their proteins to TNT-connected cells. Certain
types, such as influenza, have even been found to transfer their genome
via TNTs. Over two dozen types of viruses have been found to transfer through and/or modulate TNT. A 2022 study suggests that SARS-CoV-2 builds tunneling nanotubes from nose cells to gain access to the brain.
Nanomedicine
Tunneling
nanotubes have the potential to be involved in applications of
nanomedicine, as they have shown the ability to transfer such treatments
between cells. Future applications look to either inhibit TNTs to
prevent nanomedicine toxicity from reaching neighboring cells, or to
promote TNT formation to increase positive effects of the medicine.
TNT-like structures
While
TNT-like structures are all made of cytoskeletal cellular protrusions,
their fundamental difference with TNTs is in the connection between two
cells. TNT-like structures do not share intracellular contents such as
ions or small molecules between connected cells–a feature that is
present in both open ended and closed ended TNTs.
A TNT-like structure called a cytoneme enables exchanges between signaling centers. The formation of cytonemes towards a FGF homolog gradient has been observed, suggesting that chemotactic controls may also induce the formation of TNT-like structures. Cytonemes, however, do not always connect the membrane two cells and can act solely as environmental sensors.
Plasmodesmata have been identified as functional channels interconnecting plant cells, and stromules interconnect plastids.
Myopodia are actin-rich cytoplasmic extensions which have been observed in embryonic Drosophila. Similar structures have been observed in Xenopus and mouse models. Actin-containing cellular protrusions dubbed "streamers" have been observed in cultured B cells.
Vesicular transport in membrane nanotubes has been modeled utilizing a continuum approach. A variety of synthetic nanotubes, based on stacking of cyclic peptides and other cyclic molecules, have been investigated.
The cell nucleus (from Latinnucleus or nuculeus 'kernel, seed'; pl.: nuclei) is a membrane-bound organelle found in eukaryoticcells. Eukaryotic cells usually have a single nucleus, but a few cell types, such as mammalian red blood cells, have no nuclei, and a few others including osteoclasts have many. The main structures making up the nucleus are the nuclear envelope, a double membrane that encloses the entire organelle and isolates its contents from the cellular cytoplasm; and the nuclear matrix, a network within the nucleus that adds mechanical support.
The cell nucleus contains nearly all of the cell's genome. Nuclear DNA is often organized into multiple chromosomes – long strands of DNA dotted with various proteins, such as histones, that protect and organize the DNA. The genes within these chromosomes are structured
in such a way to promote cell function. The nucleus maintains the
integrity of genes and controls the activities of the cell by regulating
gene expression.
Because the nuclear envelope is impermeable to large molecules, nuclear pores are required to regulate nuclear transport of molecules across the envelope. The pores cross both nuclear membranes, providing a channel through which larger molecules must be actively transported by carrier proteins while allowing free movement of small molecules and ions. Movement of large molecules such as proteins and RNA
through the pores is required for both gene expression and the
maintenance of chromosomes. Although the interior of the nucleus does
not contain any membrane-bound subcompartments, a number of nuclear bodies exist, made up of unique proteins, RNA molecules, and particular parts of the chromosomes. The best-known of these is the nucleolus, involved in the assembly of ribosomes.
Diagram of the nucleus showing the ribosome-studded outer nuclear membrane, nuclear pores, DNA (complexed as chromatin), and the nucleolus.
The nucleus contains nearly all of the cell's DNA, surrounded by a network of fibrous intermediate filaments called the nuclear matrix, and is enveloped in a double membrane called the nuclear envelope. The nuclear envelope separates the fluid inside the nucleus, called the nucleoplasm, from the rest of the cell. The size of the nucleus is correlated to the size of the cell, and this ratio is reported across a range of cell types and species. In eukaryotes the nucleus in many cells typically occupies 10% of the cell volume. The nucleus is the largest organelle in animal cells. In human cells, the diameter of the nucleus is approximately six micrometres (μm).
A cross section of a nuclear pore on the surface of the nuclear envelope (1). Other diagram labels show (2) the outer ring, (3) spokes, (4) basket, and (5) filaments.
The nuclear envelope consists of two membranes, an inner and an outer nuclear membrane, perforated by nuclear pores.
Together, these membranes serve to separate the cell's genetic material
from the rest of the cell contents, and allow the nucleus to maintain
an environment distinct from the rest of the cell. Despite their close
apposition around much of the nucleus, the two membranes differ
substantially in shape and contents. The inner membrane surrounds the
nuclear content, providing its defining edge. Embedded within the inner membrane, various proteins bind the intermediate filaments that give the nucleus its structure. The outer membrane encloses the inner membrane, and is continuous with the adjacent endoplasmic reticulum membrane. As part of the endoplasmic reticulum membrane, the outer nuclear membrane is studded with ribosomes that are actively translating proteins across membrane. The space between the two membranes is called the perinuclear space, and is continuous with the endoplasmic reticulum lumen.
In a mammalian nuclear envelope there are between 3000 and 4000 nuclear pore complexes (NPCs) perforating the envelope. Each NPC contains an eightfold-symmetric ring-shaped structure at a position where the inner and outer membranes fuse. The number of NPCs can vary considerably across cell types; small glial cells only have about a few hundred, with large Purkinje cells having around 20,000. The NPC provides selective transport of molecules between the nucleoplasm and the cytosol. The nuclear pore complex is composed of approximately thirty different proteins known as nucleoporins. The pores are about 60–80 million daltons in molecular weight and consist of around 50 (in yeast) to several hundred proteins (in vertebrates).
The pores are 100 nm in total diameter; however, the gap through which
molecules freely diffuse is only about 9 nm wide, due to the presence of
regulatory systems within the center of the pore. This size selectively
allows the passage of small water-soluble molecules while preventing
larger molecules, such as nucleic acids
and larger proteins, from inappropriately entering or exiting the
nucleus. These large molecules must be actively transported into the
nucleus instead. Attached to the ring is a structure called the nuclear basket
that extends into the nucleoplasm, and a series of filamentous
extensions that reach into the cytoplasm. Both structures serve to
mediate binding to nuclear transport proteins.
Most proteins, ribosomal subunits, and some RNAs are transported
through the pore complexes in a process mediated by a family of
transport factors known as karyopherins.
Those karyopherins that mediate movement into the nucleus are also
called importins, whereas those that mediate movement out of the nucleus
are called exportins. Most karyopherins interact directly with their
cargo, although some use adaptor proteins. Steroid hormones such as cortisol and aldosterone, as well as other small lipid-soluble molecules involved in intercellular signaling, can diffuse through the cell membrane and into the cytoplasm, where they bind nuclear receptor proteins that are trafficked into the nucleus. There they serve as transcription factors when bound to their ligand; in the absence of a ligand, many such receptors function as histone deacetylases that repress gene expression.
In animal cells, two networks of intermediate filaments provide the nucleus with mechanical support: The nuclear lamina
forms an organized meshwork on the internal face of the envelope, while
less organized support is provided on the cytosolic face of the
envelope. Both systems provide structural support for the nuclear
envelope and anchoring sites for chromosomes and nuclear pores.
The nuclear lamina is composed mostly of lamin
proteins. Like all proteins, lamins are synthesized in the cytoplasm
and later transported to the nucleus interior, where they are assembled
before being incorporated into the existing network of nuclear lamina. Lamins found on the cytosolic face of the membrane, such as emerin and nesprin,
bind to the cytoskeleton to provide structural support. Lamins are also
found inside the nucleoplasm where they form another regular structure,
known as the nucleoplasmic veil, that is visible using fluorescence microscopy. The actual function of the veil is not clear, although it is excluded from the nucleolus and is present during interphase. Lamin structures that make up the veil, such as LEM3, bind chromatin and disrupting their structure inhibits transcription of protein-coding genes.
Like the components of other intermediate filaments, the lamin monomer contains an alpha-helical domain used by two monomers to coil around each other, forming a dimer structure called a coiled coil. Two of these dimer structures then join side by side, in an antiparallel arrangement, to form a tetramer called a protofilament. Eight of these protofilaments form a lateral arrangement that is twisted to form a ropelike filament.
These filaments can be assembled or disassembled in a dynamic manner,
meaning that changes in the length of the filament depend on the
competing rates of filament addition and removal.
Mutations in lamin genes leading to defects in filament assembly cause a group of rare genetic disorders known as laminopathies. The most notable laminopathy is the family of diseases known as progeria, which causes the appearance of premature aging in those with the condition. The exact mechanism by which the associated biochemical changes give rise to the aged phenotype is not well understood.
A mouse fibroblast nucleus in which DNA is stained blue. The distinct chromosome territories of chromosome 2 (red) and chromosome 9 (green) are stained with fluorescent in situ hybridization.
The cell nucleus contains the majority of the cell's genetic material
in the form of multiple linear DNA molecules organized into structures
called chromosomes. Each human cell contains roughly two meters of DNA. During most of the cell cycle these are organized in a DNA-protein complex known as chromatin, and during cell division the chromatin can be seen to form the well-defined chromosomes familiar from a karyotype. A small fraction of the cell's genes are located instead in the mitochondria.
There are two types of chromatin. Euchromatin is the less compact DNA form, and contains genes that are frequently expressed by the cell. The other type, heterochromatin, is the more compact form, and contains DNA that is infrequently transcribed. This structure is further categorized into facultative heterochromatin, consisting of genes that are organized as heterochromatin only in certain cell types or at certain stages of development, and constitutive heterochromatin that consists of chromosome structural components such as telomeres and centromeres. During interphase the chromatin organizes itself into discrete individual patches, called chromosome territories.
Active genes, which are generally found in the euchromatic region of
the chromosome, tend to be located towards the chromosome's territory
boundary.
The nucleolus is the largest of the discrete densely stained, membraneless structures known as nuclear bodies found in the nucleus. It forms around tandem repeats of rDNA, DNA coding for ribosomal RNA (rRNA). These regions are called nucleolar organizer regions (NOR). The main roles of the nucleolus are to synthesize rRNA and assemble ribosomes.
The structural cohesion of the nucleolus depends on its activity, as
ribosomal assembly in the nucleolus results in the transient association
of nucleolar components, facilitating further ribosomal assembly, and
hence further association. This model is supported by observations that
inactivation of rDNA results in intermingling of nucleolar structures.
In the first step of ribosome assembly, a protein called RNA polymerase I transcribes rDNA, which forms a large pre-rRNA precursor. This is cleaved into two large rRNA subunits – 5.8S, and 28S, and a small rRNA subunit18S. The transcription, post-transcriptional processing, and assembly of rRNA occurs in the nucleolus, aided by small nucleolar RNA (snoRNA) molecules, some of which are derived from spliced introns from messenger RNAs encoding genes related to ribosomal function. The assembled ribosomal subunits are the largest structures passed through the nuclear pores.
When observed under the electron microscope, the nucleolus can be seen to consist of three distinguishable regions: the innermost fibrillar centers (FCs), surrounded by the dense fibrillar component (DFC) (that contains fibrillarin and nucleolin), which in turn is bordered by the granular component (GC) (that contains the protein nucleophosmin).
Transcription of the rDNA occurs either in the FC or at the FC-DFC
boundary, and, therefore, when rDNA transcription in the cell is
increased, more FCs are detected. Most of the cleavage and modification
of rRNAs occurs in the DFC, while the latter steps involving protein
assembly onto the ribosomal subunits occur in the GC.
Besides the nucleolus, the nucleus contains a number of other nuclear bodies. These include Cajal bodies, gemini of Cajal bodies, polymorphic interphase karyosomal association (PIKA), promyelocytic leukaemia (PML) bodies, paraspeckles,
and splicing speckles. Although little is known about a number of these
domains, they are significant in that they show that the nucleoplasm is
not a uniform mixture, but rather contains organized functional
subdomains.
Other subnuclear structures appear as part of abnormal disease
processes. For example, the presence of small intranuclear rods has been
reported in some cases of nemaline myopathy. This condition typically results from mutations in actin, and the rods themselves consist of mutant actin as well as other cytoskeletal proteins.
Cajal bodies and gems
Cajal body
A nucleus typically contains between one and ten compact structures called Cajal bodies or coiled bodies (CB), whose diameter measures between 0.2 μm and 2.0 μm depending on the cell type and species. When seen under an electron microscope, they resemble balls of tangled thread and are dense foci of distribution for the protein coilin. CBs are involved in a number of different roles relating to RNA processing, specifically small nucleolar RNA (snoRNA) and small nuclear RNA (snRNA) maturation, and histone mRNA modification.
Similar to Cajal bodies are Gemini of Cajal bodies, or gems, whose name is derived from the Gemini constellation
in reference to their close "twin" relationship with CBs. Gems are
similar in size and shape to CBs, and in fact are virtually
indistinguishable under the microscope. Unlike CBs, gems do not contain small nuclear ribonucleoproteins (snRNPs), but do contain a protein called survival of motor neuron (SMN) whose function relates to snRNP biogenesis. Gems are believed to assist CBs in snRNP biogenesis,
though it has also been suggested from microscopy evidence that CBs and
gems are different manifestations of the same structure.
Later ultrastructural studies have shown gems to be twins of Cajal
bodies with the difference being in the coilin component; Cajal bodies
are SMN positive and coilin positive, and gems are SMN positive and
coilin negative.
PIKA and PTF domains
PIKA domains, or polymorphic interphase karyosomal associations, were
first described in microscopy studies in 1991. Their function remains
unclear, though they were not thought to be associated with active DNA
replication, transcription, or RNA processing.
They have been found to often associate with discrete domains defined
by dense localization of the transcription factor PTF, which promotes
transcription of small nuclear RNA (snRNA).
PML-nuclear bodies
Promyelocytic leukemia protein
(PML-nuclear bodies) are spherical bodies found scattered throughout
the nucleoplasm, measuring around 0.1–1.0 μm. They are known by a number
of other names, including nuclear domain 10 (ND10), Kremer bodies, and
PML oncogenic domains.
PML-nuclear bodies are named after one of their major components, the
promyelocytic leukemia protein (PML). They are often seen in the nucleus
in association with Cajal bodies and cleavage bodies.
Pml-/- mice, which are unable to create PML-nuclear bodies, develop
normally without obvious ill effects, showing that PML-nuclear bodies
are not required for most essential biological processes.
Splicing speckles
Speckles are subnuclear structures that are enriched in pre-messenger
RNA splicing factors and are located in the interchromatin regions of
the nucleoplasm of mammalian cells.
At the fluorescence-microscope level they appear as irregular, punctate
structures, which vary in size and shape, and when examined by electron
microscopy they are seen as clusters of interchromatin granules.
Speckles are dynamic structures, and both their protein and RNA-protein
components can cycle continuously between speckles and other nuclear
locations, including active transcription sites. Speckles can work with p53
as enhancers of gene activity to directly enhance the activity of
certain genes. Moreover, speckle-associating and non-associating p53
gene targets are functionally distinct.
Studies on the composition, structure and behaviour of speckles
have provided a model for understanding the functional
compartmentalization of the nucleus and the organization of the
gene-expression machinery splicing snRNPs and other splicing proteins necessary for pre-mRNA processing.
Because of a cell's changing requirements, the composition and location
of these bodies changes according to mRNA transcription and regulation
via phosphorylation of specific proteins.
The splicing speckles are also known as nuclear speckles (nuclear
specks), splicing factor compartments (SF compartments), interchromatin
granule clusters (IGCs), and B snurposomes.
B snurposomes are found in the amphibian oocyte nuclei and in Drosophila melanogaster embryos. B snurposomes appear alone or attached to the Cajal bodies in the electron micrographs of the amphibian nuclei. IGCs function as storage sites for the splicing factors.
Discovered by Fox et al. in 2002, paraspeckles are irregularly shaped compartments in the interchromatin space of the nucleus. First documented in HeLa cells, where there are generally 10–30 per nucleus, paraspeckles are now known to also exist in all human primary cells, transformed cell lines, and tissue sections.
Their name is derived from their distribution in the nucleus; the
"para" is short for parallel and the "speckles" refers to the splicing
speckles to which they are always in close proximity.
Paraspeckles sequester nuclear proteins and RNA and thus appear to function as a molecular sponge that is involved in the regulation of gene expression.
Furthermore, paraspeckles are dynamic structures that are altered in
response to changes in cellular metabolic activity. They are
transcription dependent
and in the absence of RNA Pol II transcription, the paraspeckle
disappears and all of its associated protein components (PSP1, p54nrb,
PSP2, CFI(m)68, and PSF) form a crescent shaped perinucleolar cap in the
nucleolus. This phenomenon is demonstrated during the cell cycle. In
the cell cycle, paraspeckles are present during interphase and during all of mitosis except for telophase. During telophase, when the two daughter nuclei are formed, there is no RNA Pol II transcription so the protein components instead form a perinucleolar cap.
Perichromatin fibrils
Perichromatin fibrils are visible only under electron microscope.
They are located next to the transcriptionally active chromatin and are
hypothesized to be the sites of active pre-mRNA processing.
Clastosomes
Clastosomes are small nuclear bodies (0.2–0.5 μm) described as having
a thick ring-shape due to the peripheral capsule around these bodies. This name is derived from the Greek klastos, broken and soma, body. Clastosomes are not typically present in normal cells, making them hard to detect. They form under high proteolytic conditions within the nucleus and degrade once there is a decrease in activity or if cells are treated with proteasome inhibitors. The scarcity of clastosomes in cells indicates that they are not required for proteasome function. Osmotic stress has also been shown to cause the formation of clastosomes.
These nuclear bodies contain catalytic and regulatory subunits of the
proteasome and its substrates, indicating that clastosomes are sites for
degrading proteins.
The nuclear envelope
allows control of the nuclear contents, and separates them from the
rest of the cytoplasm where necessary. This is important for controlling
processes on either side of the nuclear membrane: In most cases where a
cytoplasmic process needs to be restricted, a key participant is
removed to the nucleus, where it interacts with transcription factors to
downregulate the production of certain enzymes in the pathway. This
regulatory mechanism occurs in the case of glycolysis, a cellular pathway for breaking down glucose to produce energy. Hexokinase is an enzyme responsible for the first step of glycolysis, forming glucose-6-phosphate from glucose. At high concentrations of fructose-6-phosphate, a molecule made later from glucose-6-phosphate, a regulator protein removes hexokinase to the nucleus,
where it forms a transcriptional repressor complex with nuclear
proteins to reduce the expression of genes involved in glycolysis.
In order to control which genes are being transcribed, the cell
separates some transcription factor proteins responsible for regulating
gene expression from physical access to the DNA until they are activated
by other signaling pathways. This prevents even low levels of
inappropriate gene expression. For example, in the case of NF-κB-controlled genes, which are involved in most inflammatory responses, transcription is induced in response to a signal pathway such as that initiated by the signaling molecule TNF-α,
binds to a cell membrane receptor, resulting in the recruitment of
signalling proteins, and eventually activating the transcription factor
NF-κB. A nuclear localisation signal
on the NF-κB protein allows it to be transported through the nuclear
pore and into the nucleus, where it stimulates the transcription of the
target genes.
The compartmentalization allows the cell to prevent translation of unspliced mRNA.
Eukaryotic mRNA contains introns that must be removed before being
translated to produce functional proteins. The splicing is done inside
the nucleus before the mRNA can be accessed by ribosomes for
translation. Without the nucleus, ribosomes would translate newly
transcribed (unprocessed) mRNA, resulting in malformed and nonfunctional
proteins.
The main function of the cell nucleus is to control gene expression and mediate the replication of DNA during the cell cycle.
It has been found that replication happens in a localised way in the
cell nucleus. In the S phase of interphase of the cell cycle;
replication takes place. Contrary to the traditional view of moving
replication forks along stagnant DNA, a concept of replication factories
emerged, which means replication forks are concentrated towards some
immobilised 'factory' regions through which the template DNA strands
pass like conveyor belts.
A generic transcription factory
during transcription, highlighting the possibility of transcribing more
than one gene at a time. The diagram includes 8 RNA polymerases however
the number can vary depending on cell type. The image also includes
transcription factors and a porous, protein core.
Gene expression first involves transcription, in which DNA is used as
a template to produce RNA. In the case of genes encoding proteins, that
RNA produced from this process is messenger RNA (mRNA), which then
needs to be translated by ribosomes to form a protein. As ribosomes are
located outside the nucleus, mRNA produced needs to be exported.
Since the nucleus is the site of transcription, it also contains a
variety of proteins that either directly mediate transcription or are
involved in regulating the process. These proteins include helicases, which unwind the double-stranded DNA molecule to facilitate access to it, RNA polymerases, which bind to the DNA promoter to synthesize the growing RNA molecule, topoisomerases, which change the amount of supercoiling in DNA, helping it wind and unwind, as well as a large variety of transcription factors that regulate expression.
Newly synthesized mRNA molecules are known as primary transcripts or pre-mRNA. They must undergo post-transcriptional modification
in the nucleus before being exported to the cytoplasm; mRNA that
appears in the cytoplasm without these modifications is degraded rather
than used for protein translation. The three main modifications are 5' capping, 3' polyadenylation, and RNA splicing. While in the nucleus, pre-mRNA is associated with a variety of proteins in complexes known as heterogeneous ribonucleoprotein particles
(hnRNPs). Addition of the 5' cap occurs co-transcriptionally and is the
first step in post-transcriptional modification. The 3' poly-adenine tail is only added after transcription is complete.
RNA splicing, carried out by a complex called the spliceosome,
is the process by which introns, or regions of DNA that do not code for
protein, are removed from the pre-mRNA and the remaining exons
connected to re-form a single continuous molecule. This process
normally occurs after 5' capping and 3' polyadenylation but can begin
before synthesis is complete in transcripts with many exons. Many pre-mRNAs can be spliced in multiple ways to produce different mature mRNAs that encode different protein sequences. This process is known as alternative splicing, and allows production of a large variety of proteins from a limited amount of DNA.
The entry and exit of large molecules from the nucleus is tightly
controlled by the nuclear pore complexes. Although small molecules can
enter the nucleus without regulation, macromolecules such as RNA and proteins require association karyopherins called importins to enter the nucleus and exportins
to exit. "Cargo" proteins that must be translocated from the cytoplasm
to the nucleus contain short amino acid sequences known as nuclear localization signals, which are bound by importins, while those transported from the nucleus to the cytoplasm carry nuclear export signals bound by exportins. The ability of importins and exportins to transport their cargo is regulated by GTPases, enzymes that hydrolyze the molecule guanosine triphosphate (GTP) to release energy. The key GTPase in nuclear transport is Ran,
which is bound to either GTP or GDP (guanosine diphosphate), depending
on whether it is located in the nucleus or the cytoplasm. Whereas
importins depend on RanGTP to dissociate from their cargo, exportins
require RanGTP in order to bind to their cargo.
Nuclear import depends on the importin binding its cargo in the
cytoplasm and carrying it through the nuclear pore into the nucleus.
Inside the nucleus, RanGTP acts to separate the cargo from the importin,
allowing the importin to exit the nucleus and be reused. Nuclear export
is similar, as the exportin binds the cargo inside the nucleus in a
process facilitated by RanGTP, exits through the nuclear pore, and
separates from its cargo in the cytoplasm.
Specialized export proteins exist for translocation of mature
mRNA and tRNA to the cytoplasm after post-transcriptional modification
is complete. This quality-control mechanism is important due to these
molecules' central role in protein translation. Mis-expression of a
protein due to incomplete excision of exons or mis-incorporation of
amino acids could have negative consequences for the cell; thus,
incompletely modified RNA that reaches the cytoplasm is degraded rather
than used in translation.
During its lifetime, a nucleus may be broken down or destroyed, either in the process of cell division or as a consequence of apoptosis (the process of programmed cell death). During these events, the structural components of the nucleus — the envelope and lamina — can be systematically degraded.
In most cells, the disassembly of the nuclear envelope marks the end of the prophase
of mitosis. However, this disassembly of the nucleus is not a universal
feature of mitosis and does not occur in all cells. Some unicellular
eukaryotes (e.g., yeasts) undergo so-called closed mitosis,
in which the nuclear envelope remains intact. In closed mitosis, the
daughter chromosomes migrate to opposite poles of the nucleus, which
then divides in two. The cells of higher eukaryotes, however, usually
undergo open mitosis,
which is characterized by breakdown of the nuclear envelope. The
daughter chromosomes then migrate to opposite poles of the mitotic
spindle, and new nuclei reassemble around them.
At a certain point during the cell cycle in open mitosis, the
cell divides to form two cells. In order for this process to be
possible, each of the new daughter cells must have a full set of genes, a
process requiring replication of the chromosomes as well as segregation
of the separate sets. This occurs by the replicated chromosomes, the sister chromatids, attaching to microtubules, which in turn are attached to different centrosomes.
The sister chromatids can then be pulled to separate locations in the
cell. In many cells, the centrosome is located in the cytoplasm, outside
the nucleus; the microtubules would be unable to attach to the
chromatids in the presence of the nuclear envelope. Therefore, the early stages in the cell cycle, beginning in prophase and until around prometaphase, the nuclear membrane is dismantled.
Likewise, during the same period, the nuclear lamina is also
disassembled, a process regulated by phosphorylation of the lamins by
protein kinases such as the CDC2 protein kinase.
Towards the end of the cell cycle, the nuclear membrane is reformed,
and around the same time, the nuclear lamina are reassembled by
dephosphorylating the lamins.
However, in dinoflagellates,
the nuclear envelope remains intact, the centrosomes are located in the
cytoplasm, and the microtubules come in contact with chromosomes, whose
centromeric regions are incorporated into the nuclear envelope (the
so-called closed mitosis with extranuclear spindle). In many other
protists (e.g., ciliates, sporozoans) and fungi, the centrosomes are intranuclear, and their nuclear envelope also does not disassemble during cell division.
Apoptosis is a controlled process in which the cell's structural
components are destroyed, resulting in death of the cell. Changes
associated with apoptosis directly affect the nucleus and its contents,
for example, in the condensation of chromatin and the disintegration of
the nuclear envelope and lamina. The destruction of the lamin networks
is controlled by specialized apoptotic proteases called caspases,
which cleave the lamin proteins and, thus, degrade the nucleus'
structural integrity. Lamin cleavage is sometimes used as a laboratory
indicator of caspase activity in assays for early apoptotic activity.
Cells that express mutant caspase-resistant lamins are deficient in
nuclear changes related to apoptosis, suggesting that lamins play a role
in initiating the events that lead to apoptotic degradation of the
nucleus. Inhibition of lamin assembly itself is an inducer of apoptosis.
The nuclear envelope acts as a barrier that prevents both DNA and
RNA viruses from entering the nucleus. Some viruses require access to
proteins inside the nucleus in order to replicate and/or assemble. DNA
viruses, such as herpesvirus
replicate and assemble in the cell nucleus, and exit by budding through
the inner nuclear membrane. This process is accompanied by disassembly
of the lamina on the nuclear face of the inner membrane.
Disease-related dynamics
Initially, it has been suspected that immunoglobulins in general and autoantibodies in particular do not enter the nucleus. Now there is a body of evidence that under pathological conditions (e.g. lupus erythematosus) IgG can enter the nucleus.
Nuclei per cell
Most eukaryotic
cell types usually have a single nucleus, but some have no nuclei,
while others have several. This can result from normal development, as
in the maturation of mammalian red blood cells, or from faulty cell division.
Anucleated cells
Human red blood cells, like those of other mammals, lack nuclei. This occurs as a normal part of the cells' development.
An anucleated cell contains no nucleus and is, therefore, incapable
of dividing to produce daughter cells. The best-known anucleated cell is
the mammalian red blood cell, or erythrocyte, which also lacks other organelles such as mitochondria, and serves primarily as a transport vessel to ferry oxygen from the lungs to the body's tissues. Erythrocytes mature through erythropoiesis in the bone marrow,
where they lose their nuclei, organelles, and ribosomes. The nucleus is
expelled during the process of differentiation from an erythroblast to a reticulocyte, which is the immediate precursor of the mature erythrocyte. The presence of mutagens may induce the release of some immature "micronucleated" erythrocytes into the bloodstream. Anucleated cells can also arise from flawed cell division in which one daughter lacks a nucleus and the other has two nuclei.
A number of dinoflagellates
are known to have two nuclei. Unlike other multinucleated cells these
nuclei contain two distinct lineages of DNA: one from the dinoflagellate
and the other from a symbiotic diatom.
Evolution
As the major defining characteristic of the eukaryotic cell, the nucleus's evolutionary
origin has been the subject of much speculation. Four major hypotheses
have been proposed to explain the existence of the nucleus, although
none have yet earned widespread support.
The first model known as the "syntrophic model" proposes that a symbiotic relationship between the archaea and bacteria created the nucleus-containing eukaryotic cell. (Organisms of the Archaeal and Bacterial domains have no cell nucleus.) It is hypothesized that the symbiosis originated when ancient archaea similar to modern methanogenic archaea, invaded and lived within bacteria similar to modern myxobacteria,
eventually forming the early nucleus. This theory is analogous to the
accepted theory for the origin of eukaryotic mitochondria and chloroplasts, which are thought to have developed from a similar endosymbiotic relationship between proto-eukaryotes and aerobic bacteria. One possibility is that the nuclear membrane arose as a new membrane system following the origin of mitochondria in an archaebacterial host. The nuclear membrane may have served to protect the genome from damaging reactive oxygen species produced by the protomitochondria.
The archaeal origin of the nucleus is supported by observations that
archaea and eukarya have similar genes for certain proteins, including histones. Observations that myxobacteria are motile, can form multicellular complexes, and possess kinases and G proteins similar to eukarya, support a bacterial origin for the eukaryotic cell.
A second model proposes that proto-eukaryotic cells evolved from
bacteria without an endosymbiotic stage. This model is based on the
existence of modern Planctomycetota bacteria that possess a nuclear structure with primitive pores and other compartmentalized membrane structures. A similar proposal states that a eukaryote-like cell, the chronocyte, evolved first and phagocytosed archaea and bacteria to generate the nucleus and the eukaryotic cell.
The most controversial model, known as viral eukaryogenesis,
posits that the membrane-bound nucleus, along with other eukaryotic
features, originated from the infection of a prokaryote by a virus. The
suggestion is based on similarities between eukaryotes and viruses such
as linear DNA strands, mRNA capping, and tight binding to proteins
(analogizing histones to viral envelopes). One version of the proposal suggests that the nucleus evolved in concert with phagocytosis to form an early cellular "predator". Another variant proposes that eukaryotes originated from early archaea infected by poxviruses, on the basis of observed similarity between the DNA polymerases in modern poxviruses and eukaryotes. It has been suggested that the unresolved question of the evolution of sex could be related to the viral eukaryogenesis hypothesis.
A more recent proposal, the exomembrane hypothesis,
suggests that the nucleus instead originated from a single ancestral
cell that evolved a second exterior cell membrane; the interior membrane
enclosing the original cell then became the nuclear membrane and
evolved increasingly elaborate pore structures for passage of internally
synthesized cellular components such as ribosomal subunits.
The nucleus was the first organelle to be discovered. What is most
likely the oldest preserved drawing dates back to the early microscopist
Antonie van Leeuwenhoek (1632–1723). He observed a "lumen", the
nucleus, in the red blood cells of salmon. Unlike mammalian red blood cells, those of other vertebrates still contain nuclei.
The nucleus was also described by Franz Bauer in 1804 and in more detail in 1831 by Scottish botanistRobert Brown in a talk at the Linnean Society of London. Brown was studying orchids
under the microscope when he observed an opaque area, which he called
the "areola" or "nucleus", in the cells of the flower's outer layer. He did not suggest a potential function.
In 1838, Matthias Schleiden proposed that the nucleus plays a role in generating cells, thus he introduced the name "cytoblast" ("cell builder"). He believed that he had observed new cells assembling around "cytoblasts". Franz Meyen
was a strong opponent of this view, having already described cells
multiplying by division and believing that many cells would have no
nuclei. The idea that cells can be generated de novo, by the "cytoblast"
or otherwise, contradicted work by Robert Remak (1852) and Rudolf Virchow (1855) who decisively propagated the new paradigm that cells are generated solely by cells ("Omnis cellula e cellula"). The function of the nucleus remained unclear.
Between 1877 and 1878, Oscar Hertwig published several studies on the fertilization of sea urchin eggs, showing that the nucleus of the sperm enters the oocyte
and fuses with its nucleus. This was the first time it was suggested
that an individual develops from a (single) nucleated cell. This was in
contradiction to Ernst Haeckel's theory that the complete phylogeny
of a species would be repeated during embryonic development, including
generation of the first nucleated cell from a "monerula", a
structureless mass of primordial protoplasm ("Urschleim").
Therefore, the necessity of the sperm nucleus for fertilization was
discussed for quite some time. However, Hertwig confirmed his
observation in other animal groups, including amphibians and molluscs. Eduard Strasburger produced the same results for plants in 1884. This paved the way to assign the nucleus an important role in heredity. In 1873, August Weismann postulated the equivalence of the maternal and paternal germ cells
for heredity. The function of the nucleus as carrier of genetic
information became clear only later, after mitosis was discovered and
the Mendelian rules were rediscovered at the beginning of the 20th century; the chromosome theory of heredity was therefore developed.
