Nucleohyaloplasm

Editor-In-Chief: Henry A. Hoff

Overview
A hyaloplasm is the clear, structureless, apparently homogeneous fluid of the cytoplasm. Similar to the hyaloplasm of a cell, the nucleus contains nucleohyaloplasm. It is a highly viscous liquid. This liquid contains enzymes (which direct activities that take place in the nucleus), intermediate metabolites, and many substances such as nucleotides (necessary for purposes as the replication of DNA and production of mRNA). All are dissolved in the nucleohyaloplasm. It is part of the nucleoplasm and is partly made up of nucleosol.

Introduction
Nucleohyaloplasm is the cytosol within the nucleus, without the microfilaments and the microtubules, also known as nucleosol, vis à vis mitosol and cytosol.

As a cytosol, it consists mostly of water, dissolved ions, small molecules, and large water-soluble molecules (such as protein). It contains about 20% to 30% protein. It has a high concentration of K⁺ ions and a low concentration of Na⁺ ions. Normal human cytosolic pH ranges between 7.3 - 7.5, depending on the cell type involved.

As a plasm it contains formative material, portions of the nucleoskeleton as it is being shaped or reshaped, macromolecules with limited mobility, and portions of the nuclear envelope as it is recycled.

Small particles
Small particles (< 40 kDa, <50 kDa , <~70 kDa , ≤ 70 kDa ) are able to pass through the nuclear pore complex by passive transport. Larger proteins require a nuclear localization signal (NLS). The pores are 100 nm in total diameter, with an opening diameter of about 50 nm; however, the gap through which molecules freely diffuse is only about 9-10 nm wide, due to the presence of regulatory systems within the center of the pore. The 10 nm diameter corresponds to an upper mass limit of 70 kDa. The majority of the non-protein molecules have a molecular mass of less than 300 Da.

This mixture of small molecules is extraordinarily complex, as the variety of molecules that are involved in metabolism (the metabolites) is immense. For example up to 200,000 different small molecules might be made in plants, although not all these will be present in the same species, or in a single cell. Estimates of the number of metabolites in a single cell of E. coli or baker's yeast predict that under 1,000 are made.

Polymerases
Many of the subunits of RNA polymerase II are small polymerases. RNA polymerase IIC (PolR2C) has a mass of 33 kDa, no NLS, is intracellular to the nucleus and is part of the transcription complex. Of the others, PolR2D-L are 19 kDa or less without NLS, except RNA polymerase IIE (PolR2E) 23 kDa has a NLS and is the only one that localizes to the nucleolus. Although PolR2K and PolR2L are small enough in mass to be considered oligopeptides, their numbers of aa are over the usual limit: 58 aa and 67 aa, respectively, in humans.

Large particles
Larger particles are also able to pass through the large diameter of a nuclear pore but at almost negligible rates. However, the nucleohyaloplasm does contain large amounts of macromolecules, which can alter how molecules behave, through macromolecular crowding. Since some of these macromolecules have less volume to move in, their effective concentration is increased. This crowding effect can produce large changes in both the rates and chemical equilibrium for reactions in the nucleohyaloplasm. It is particularly important in its ability to alter dissociation constants by favoring the association of macromolecules, such as when multiple proteins come together to form protein complexes, or when DNA-binding proteins bind to their targets in the genome.

Proteins
Proteins larger than those allowed through a nuclear pore by passive transport require a nuclear localization signal (NLS). This is an amino acid sequence that targets the cytosolic nuclear transport receptors of the nuclear pore complex. A nuclear import NLS will bind strongly to importin, while an export NLS (nuclear export signal, NES) binds to an exportin. For example, RNA polymerase IIA (Rbp1) 220kDa has a NLS.

Nuclear localization
The subcellular distribution of a substance to or within the nucleus is often referred to as nuclear localization. Many mechanisms have been found that produce nuclear localization in addition to a NLS.

Zac1 is a seven-zinc-finger transcription factor that preferentially binds GC-rich DNA elements and has intrinsic transactivation activity. The zinc-finger motif is of a Cys2His2-type. This motif is involved in DNA binding, dimerization, transactivation activity, and nuclear localization of Zac1 through interacting with importin α1. Zac1 has no typical NLS. Any two or more zinc-finger motifs act in concert to facilitate nuclear localization. Apparently, as with importin α transport of CaMKIV to the nucleus, importin α1 may mediate transport of Zac1 to the nucleus without the involvement of importin β. But, some other factors are involved, perhaps Ran-binding proteins such as RanBPM and Mog1, which play roles in nucleocytoplasmic transport and transcription factor recruitment.

Transcription factories
Active transcription units are clustered in the nucleus, in discrete sites called ‘transcription factories’. Such sites can be visualized after allowing engaged polymerases to extend their transcripts in tagged precursors (Br-UTP or Br-U), and immuno-labeling the tagged nascent RNA. Transcription factories can also be localized using fluorescence in situ hybridization, or marked by antibodies directed against polymerases. There are ~8,000 polymerase II factories in the nucleoplasm of a HeLa cell. Each polymerase II factory contains ~8 polymerases. As most active transcription units are associated with only one polymerase, each factory can be associated with ~8 different transcription units. That's ~64,000 polymerase II active transcription units. These units might be associated through promoters and/or enhancers, with loops forming a ‘cloud’ around the factory.

RNA
Of the many different types of RNA that can occur within a cell, most also can occur dissolved in the nucleohyaloplasm. In addition to mRNA, which is constructed during gene transcription to produce protein, there are a variety of RNAs that are transcripted from genes for their own sake into the nucleohyaloplasm: ribosomal RNA (rRNA), transfer RNA (tRNA), small nuclear RNA (snRNA), small nucleolar RNA (snRNA), non-coding RNA (ncRNA), miscRNA, microRNA, piwi-interacting RNA (piRNA), small interfering RNA (siRNA), signal recognition particle RNA (SRP RNA), and guide RNA (gRNA).

Each has to be transcribed from the applicable portion of DNA in the euchromatin. The unfolded structure of euchromatin allows gene regulatory proteins and RNA polymerase (RNAP) complexes to bind to the DNA sequence, which can then initiate the transcription process. Control of the process of gene transcription affects patterns of gene expression and thereby allows a cell to adapt to a changing environment, perform specialized roles within an organism, and maintain basic metabolic processes necessary for survival. RNAP can initiate transcription at specific DNA sequences known as promoters. It then produces an RNA chain which is complementary to the template DNA strand.

Ribozymes are transcripted into the nucleohyaloplasm. The functional part of the ribosome, the molecular machine that translates RNA into proteins, is fundamentally a ribozyme. Ribozymes often have divalent metal ions such as Mg2+ as cofactors. Ribozyme RNase P 30kDa has a NLS. But, RNase P subunit p25 (25 kDa), which is also localized to the nucleolus does not.

Probably the largest mRNA transcripted into the nucleohyaloplasm is from the gene for dystrophin (427 kDA protein). The primary transcript measures 2.4 megabases (thus the gene comprises 0.008% of the human genome), and takes 16 hours to transcribe. The 79 exons code for a protein of 3685 amino acid residues. Its mRNA is 14 kb or ~550 kDa.

Chromatin
Euchromatin is the less compact DNA form, and contains genes that are frequently expressed by the cell. Active genes, which are generally found in the euchromatic region of the chromosome, tend to be located towards the chromosome's territory boundary.

Heterochromatin is usually localized to the periphery of the nucleus along the nuclear envelope. It mainly consists of genetically inactive satellite sequences, and many genes are repressed to various extents, although some cannot be expressed in euchromatin at all.

Mobile chromatin
During interphase euchromatin is known to be attached to the nucleolus or nucleoli and heterochromatin is attached to the nuclear envelope. Further, in some cell types interphase euchromatin and heterochromatin are translationally immobile over distances ≥400 nm. Mobile chromatin can move with velocities averaging 50-70 nm/sec.

Nucleolus
The nucleolus is roughly spherical, and is surrounded by the euchromatin. No membrane separates the nucleolus from the nucleohyaloplasm. Nucleoli carry out the production and maturation of ribosomes. Large numbers of ribosomes are found inside.

Direct contact between the nucleolus and the nuclear envelope is frequently observed but is not dependent on nucleolar activity.

Although the size of the nucleolus is highly variable in any particular cell nucleus, there is in some cells a correlation with cell diameter: increasing cell size to increasing rounded diameter of the nucleolus. Based on this correlation, for an average mammalian cell of 6000 nm, the nucleolus would be ~300 nm in diameter. Interferometric analysis of nucleolus mass for mesothelial cells in culture places its mass average at 40 x 10-12 gm (40 pgm) or approximately 24 TDa (teradaltons). In addition, each cell may have approximately the same total nucleolar mass regardless of the number of nucleoli.

Structures
Of the structures local to the nucleohyaloplasm, some serve to confine it such as the inner membrane of the nuclear envelope. While others are completely suspended within it, for example, the nucleolus. Still others such as the nuclear matrix and nuclear lamina are found throughout the inside of the nucleus, some as part of the nucleoskeleton.

Besides the nucleolus, the nucleus contains a number of other non-membrane delineated bodies. These include Cajal bodies, Gemini of coiled 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 nucleohyaloplasm is not a uniform mixture, but rather contains organized functional subdomains.

Intra-nuclear transport
The lateral speed of a water molecule is ~35 µm/sec. The lateral speed of larger biological molecules in passive diffusion in water is on the order of 500 - 50 nm/sec. But in cytosol such as the nucleohyaloplasm: ~120 - 10 nm/sec due to crowding and collisions with large molecules. In mammalian cells, the average diameter of the nucleus is approximately 6 μm. The large amount of DNA and RNA should hinder the migration of nuclear proteins, but a protein could traverse the entire diameter of a nucleus in a matter of minutes.

Proteins are frequently transported across the cytosol, along well-defined routes, and delivered to particular addresses. Passive diffusion cannot account for the rate, directionality, or destination of such transport. Microtubules function as tracks and the movement is propelled by motor proteins. Movement can occur in both directions and at velocities between ~5 and 3000 nm/sec. One motor protein that localizes intracellularly to the nucleus is myosin 1F 125 kDa. It does not have a NLS. Its N terminal motor domain uses ATP and has actin binding sites.

All Cajal bodies move through the nucleohyaloplasm. These movements include translocations and moving to or from the nucleolus at velocities of ~10-15 nm/sec.

Constrained microstructures (~40-100 nm in size, on the order of 0.5-5 MDa) move with velocities averaging 50-70 nm/sec (comparable to that of mobile chromatin), but free-moving microstructures (in chromatin-free channels) can move at speeds of up to 500 nm/sec. The movement of PML-containing microstructures is energy-independent. Such mobility is characteristic of constrained diffusion.

The movement of elongated chromosomes throughout the chromatin filled nucleus may be associated with intranuclear motor protein action.

Human nucleohyaloplasm
The human genome contains many of the genes discussed in the sections above regarding the structure, composition, and physiology of the nucleohyaloplasm. These genes and comparable ones in similar species help to understand human evolutionary genetics.

Mature monocytes circulating in human peripheral blood contain multiple nucleoli of various sizes in one and the same nucleus. The nucleolar RNA content is apparently related to the nucleolar size.

Increases in the number of nucleoli, their size, and their activity reflect the proliferating activity of exponentially growing cells.

Acknowledgements
The content on this page was first contributed by: Henry A. Hoff.

Initial content for this page in some instances came from Wikipedia.