• The cell which we have already described is a typical cell in which the nucleus and cytoplasm with its organized are found.

• Such cell which contains a true nucleus is referred to as eukaryotic cell (Gr.: Karyon = nucleus),

• But if the nuclear envelope is lacking, and thus, the nuclear substance is in direct contact with the rest of the protoplasm the cell is called =

prokaryotic cell (e.g., bacteria, some algae and most viruses).

What is the nucleus?

• The nucleus is the brain of eukaryotic cells.

• It is only present in eukaryotic cells (which are eukaryotic because they have a nucleus) and there is only one of these organelles in each cell.

• It is a membrane bound structure that contains the cell's hereditary information and controls the cell's growth and reproduction.

The nucleus has three primary responsibilities:

• execution and regulation of cytoplasmic events through its gene products, storage of information in the form of the DNA of genes, and retrieval of information from these genes in the form of RNA.

• The nucleus is the site for three important processes:

• replication of chromatin (DNA + protein), transcription of genes into RNA,

• and processing of those RNAs into mature forms, many of which are exported to the cytosol for translation.

• The cell nucleus is a remarkable organelle because it forms the package for our genes and their controlling factors. It functions to:

1. Store genes on chromosomes.2. Organize genes into chromosomes to allow cell division.3. Transport regulatory factors & gene products via

nuclear pores 4. Produce messages (

messenger Ribonucleic acid or mRNA) that code for proteins .

5. Produce ribosomes in the nucleolus .6. Organize the uncoiling of DNA to replicate  key genes.7. The nucleus also controls the synthesis of proteins in

the cytoplasm through the use of messenger RNA.

1) Nuclear envelope. (2) Nuclear pore (3)Nucleolus.( 4) Chromatin. (5)) Nucleus(6. ( Nucleoplasm.

2) The whole structure is surrounded by cytoplasm. 3) Similar to the cytoplasm of a cell, the nucleus contains nucleoplasm - a

highly viscous liquid containing the chromosomes and nucleoli.4) Chromosones contain information encoded in DNA attached to proteins

called histones and are usually arranged in to a dense network called chromatin.

5) Nucleoli are granular structures which make ribonucleic DNA (rDNA) and assemble it with proteins.

• The nucleus is the most prominent organelle in the cell, taking up about 10% of the total volume of the cell.

• Usually it is round and is the largest organelle in the cell. The nucleus varies in diameter from 10 to 20 micrometres

• The cell nucleus is a remarkable organelle because it forms the package for our genes and their controlling factors

• The nucleus is surrounded by a double membrane (each with the typical unit membrane structure) forming the nuclear envelope, . (nuclear membrane), which is similar to the cell membrane, about 30 nm wide.

• This membrane separates the contents of the nucleus from the cytoplasm (cyto-).

• The envelope helps to maintain the shape of the nucleus and assists in regulating the flow of molecules into and out of the nucleus through nuclear pores.

The nuclear envelope is supported from the exterior by the intermediate filament network and microtubules,

• while the nuclear lamina, which is located adjacent to • the inner surface of the nuclear envelope, provides internal

support.

• The space between the outer and inner membranes is also continuous with rough endoplasmic reticulum space.

• It can fill with newly synthesized proteins just as the rough endoplasmic reticulum does.

1. The inner and outer membrane fuse at regular spaces, forming nuclear pores, and which is continuous with the endoplasmic reticulum.

2. The nuclear pore complexes are the ports through which certain proteins enter and RNAs exit the nucleus.

• nuclear pores allow specific materials to pass in and out of the nucleus, just like proteins in the cell membrane regulate the movement of molecules in and out of the cell itself. 

• These pores permit the movement of selected molecules, like mRNA, from the nucleus to the rest of the cell.

• The protoplasm of the cell-body is usually known as cytoplasm or cytosome, while that of the nucleus is referred to as

karyoplasms.

““Karyoplasmic ratioKaryoplasmic ratio””• It has been found that in any species there is a relationship between

the size of nucleus and the size of the cytoplasm, this is known as the “Karyoplasmic ratio” This has been first originated protozoa and then extended to other animals.

• The karyoplasmic ration has considerable importance in the life of the cell, and is probably one of the factors which brings about cell-division

• The relation usually holds in cells which are homologous (i.e., of similar origin and function) as shown in sea-urchin larvae either from fertilized or unfertilized eggs. However, under different conditions this ration may vary.

The interphase nucleusThe interphase nucleus

• The nucleus in the interphase (the phase between two successive divisions) contains the chromatin substance; some of the larger flakes of chromatin are called chromocentres, karyosomes or false nucleoli because they are morphologically similar to some nucleoli.

• Chromatin substance is distributed throughout the nuclear sap (nucleoplasm).

• In addition, there are one or more spherical bodies, the nucleoli which differ from the karyosomes in some staining properties and in chemical composition.

The The nucleolusnucleolus

The nucleus also contains the nucleolus, Each cell nucleus usually contains at least one nucleolus which helps to synthesize ribosomes.

It is very densely concentrated and appears as a dark, almost perfectly circular dot in the middle of the unstained nuclear region.

The nucleolus contains ribosomes, RNA, DNA, and proteins. The nucleolus has some of the ribosomes that synthesize proteins (others are in the endoplasmic reticulum).

The mRNA content in this region is very high: it's being produced in large amounts to facilitate the protein-making activity. Copious amounts of RNA and proteins can be found in the nucleolus as well. r RNA is produced in the nucleolus of the cell and travels to the cytoplasm through the pores of the nuclear envelope.

• The nucleolus contains nucleolar organizers which are parts of chromosomes with the genes for ribosome synthesis on them. A number of chromosomes get together and transcribe ribosomal RNA at this site.

• The above figure shows electron micrograph of a nucleolus with the nuclear organizing region. The nuclear organizing (NO) regions are seen as circular areas (pale) surrounded by a rim of electron dense filaments. These filaments collectively are called the pars fibrosa (PF).

• This is formed from newly transcribed ribosomal RNA.

• Nucleoli increase in number and enlarge when the cell is stimulated to produce proteins. This is a sign that the cell has been stimulated or is actively involved in protein synthesis. Nucleoli disappear during cell division and then reform at the chromosomal nucleolar organizing centers.

The chromatinThe chromatin • the chromosomes are organized into long structures called

chromatin and not into individual chromosomes as we typically think.

• The chromatin (meaning "colored substance") contains DNA and proteins formed into packets of code called chromosomes. When the cell divides, the chromosomes fold up on themselves, getting wider.

• Chromatin substance is distributed throughout the nuclear sap (nucleoplasm).

• Chromatin has been divided into: • euchromatin and • heterochromatin. •

Euchromatin and HeterochromatinEuchromatin and Heterochromatin• Several different chemical techniques are used to

identify certain chromosomal regions by staining then so that they form chromosomal bands.

• For example, darker bands are generally found near the centromeres or on the ends (telomeres) of the chromosome, while other regions do not stain as strongly.

• The position of the dark-staining are heterochromatic region or heterochromatin.

• Light staining are euchromatic region or euchromatin.

Eurochromatin / HeterochromatinEurochromatin / Heterochromatin

The chromatinThe chromatin The chromatin substance; Heterochromatin was defined as a structure that does not alter in its condensation throughout the cell cycle , Heterochromatin is localized principally on the periphery of the nucleus

Euchromatin is decondensed during interphase.and is localized in the interior of the nucleoplasm.

• Light staining are euchromatic region or euchromatin.• Experiments of pulse labeling with 3H-uridine and

autoradiography showed that most transcription occurs in regions of euchromatin and the nucleolus

• dark-staining are heterochromatic region or heterochromatin. Heterchromatin: A region of the chromatin that does not uncoil after mitosis.

• In general, heterochromatic regions are sites of inactive genes; however some transcribed genes have been located in regions of heterchromatin.

Euchromatin and HeterochromatinEuchromatin and Heterochromatin

HeterochromatinHeterochromatin• Heterochromatin is classified into two groups: • (i) Constitutive and (ii) Facultative. • Constitutive heterochromatin remains permanently in

the heterochromatic stage, i.e., it does not revert to the euchromatic stage.

• In contrast, facultative heterochromatin consists of euchromatin that takes on the staining and compactness characteristics of heterochromatin during some phase of development.

• In mammalian cells, heterochromatin appears as darkly staining regions of the nucleus, often associated with the nuclear envelope

• Heterochromatin is classified into two groups: • (i) Constitutive and (ii) Facultative.• • Constitutive heterochromatin remains

permanently in the heterochromatic stage, i.e., it does not revert to the euchromatic stage.

• In contrast, facultative heterochromatin consists of euchromatin that takes on the staining and compactness characteristics of heterochromatin during some phase of development.

The chromatin substanceThe chromatin substance;;

constitutive heterochromatin, containing few genes and formed principally of repetitive sequences located in large regions with centromeres and telomeres.

facultative heterochromatin composed of transcriptionally active regions that can adopt the structural and functional characteristics of heterochromatin, such as the inactive X chromosome of mammals.

ChromatinChromatin • A)Heterochromatin • This is the condensed form of

chromatin organization.

• It is seen as dense patches of chromatin. Sometimes it lines the nuclear membrane, however, it is broken by clear areas at the pores so that transport is allowed.

• Abundant heterochromatin is seen in resting, or reserve cells such as small lymphocytes (memory cells) waiting for exposure to a foreign antigen.

• Heterochromatin is considered transcriptionally inactive.

• B) Euchromatin • Euchromatin is

threadlike, delicate.

• the presence of euchromatin is significant because the regions of DNA to be transcribed or duplicated must uncoil before the genetic code can be read.

• It is most abundant in active, transcribing cells.

Proteins associated with Chromatin• The complexes between eukaryotic DNA and

proteins are called Chromatin, which typically contains about twice as much protein as DNA.

• The major proteins of chromatin are the histones – small proteins containing a high proportion of basic aminoacids (arginine and lysine) that facilitate binding negatively charged DNA molecule .

• There are 5 major types of histones: H1, H2A, H2B, H3, and H4 – which are very similar among different sp of eukaryotes.

The major histone proteins:

Histone Mol. Wt No. of Percentage

Amino acid Lys + Arg

H1 22,500 244 30.8

H2A 13,960 129 20.2

H2B 13,774 125 22.4

H3 15,273 135 22.9

H4 11,236 102 24.5

The DNA double helix is bound to proteins called histones.  The histones have  positively charged (basic) amino acids to bind the negatively charged (acidic) DNA. 

Here is an SDS gel of histone proteins, separated by size

• In addition, chromatin contains an approximately equal mass of a wide variety of non-histone chromosomal proteins.

• There are more than a thousand different types of these proteins, which are involved in a range of activities, including DNA replication and gene expression.

• The DNA of prokaryotes is similarly associated with proteins, some of which presumably function as histones do, packing the DNA within the bacterial cell.

Histones, however are unique feature of eukaryotic cells and are responsible for distinct structural organization of eukaryotic chromatin

Nucleosome,

• The fundamental unit of chromatin, termed the nucleosome, is composed of DNA and histone proteins. This structure provides the first level of compaction of DNA into the nucleus.

The nucleosome is the fundamental unit of chromatin. It is composed of:

a core particle and a linker region (or internucleosomal region) that joins adjacent core particles .

• The the nucleosome, was described by Roger Kornberg in 1974.

• Two types of experiments led to Kornberg’s proposal of the nucleosome model.

• First, partial digestion of chromatin with micrococcal nuclease (an enzyme that degrades DNA) was found to yield DNA fragments approximately 200 base pairs long.

• In contrast, a similar digestion of naked DNA (not associated with protein) yielded a continuous smear randomly sized fragments.

• These results suggest that the binding of proteins to DNA in chromatin protects the regions of DNA from

from nuclease digestion, so that enzyme can

attack DNA only at sites separated by

approximately 200 base pairs.

• Second: Electron microscopy revealed that chromatin fibers have a beaded appearance, with the beads spaced at intervals of approximately 200 base pairs.

• Thus, both nuclease digestion and the electron microscopic studies suggest that chromatin is composed of repeating 200 base pair unit, which were called

nucleosome.

• Detailed analysis of these nucleosome core particles has shown that they contain 146 base pairs of DNA wrapped 1.75 times around a histone core consisting of two molecules each of H2A, H2B, H3, and H4 (the core histones).

• One molecule of the fifth histone H1, is bound to the DNA as it enters and exists each nucleosome core particle. This forms a chromatin subunit known as chromatosome.

• The length of the linker region, however, varies between species and cell type. It is within this region that the variable linker histones are incorporated. Therefore, the total length of DNA in the nucleosome can vary with species from 160 to 241 base pairs.

IV- General steps in chromatin assemblyIV- General steps in chromatin assembly • The assembly of DNA into chromatin involves a range of events,

beginning with the formation of the basic unit, the nucleosome, and ultimately giving rise to a complex organization of specific domains within the nucleus.

• The first step is the deposition onto the DNA of a tetramer of newly synthesized (H3-H4)2 to form a sub-nucleosomal particle, which is followed by the addition of two H2A-H2B dimers. This produces a nucleosomal core particle consisting of 146 base pairs of DNA wound around the histone octamer.

• This core particle and the linker DNA together form the nucleosome. Newly synthesized histones are specifically modified (e.g.the acetylation of histone H4).

• • The next step is the maturation step that requires ATP to establish

regular spacing of the nucleosome cores to form the nucleofilament. During this step the newly incorporated histones are de-acetylated.

• ·

• • During the maturation step, incorporation of linker histones,

non-histone chromatin associated proteins, called HMG (High Mobility Group), and other specific DNA-binding factors help to space and fold the nucleofilament. Therefore the early steps in assembly can have a great impact on the final characteristics of chromatin in specific nuclear domains.

The endThe end