Cell Biology

S. Rahgozar,PhD

University of Isfahan

Faculty of Science

3. The nucleus

3.2. Internal organization and rRNA processing

1392-93

Internal organization of the nucleus

Lamin-dependent complexes form nuclear bodies that have roles in

DNA repair, chromatin organization, gene regulation and

signal transduction

Heterochromatin in interphase nuclei

Chromosomes and higher-order chromatin structure

Types of heterochromatins

Constitutive

Facultative

The amount of facultative heterochromatin

varies depending on the transcriptional activity

of the cell

o Interphase chromatin is not randomly distributed within the nucleus

1885, Carl Rabl: Chromosome organization in

salamander. A) Complete chromosome, B)

Telomeres only

1984, Chromosome organization in Drosophila.

Rather than randomly winding around one another,

each chromosome occupies a discrete region of

the nucleus. B) Topological separation between

chromosomes.

o Individual chromosomes occupy distinct

territories within the nuclei of mammalian cells

Location of actively transcribed genes, where newly

transcribed RNAs will be released.

Location of heterochromatin bound to the matrix of

the nuclear lamina

the chromosome location differ upon the

species and tissues.

chromatin reorganizes during cell differentiation

and gene expression by means of nuclear actin and

myosin.

Looped domains contain

50-100kb of DNA. They

appear to represent

discrete functional units,

which independently

regulate gene expression. Lampbrush chromosome, amphibian oocyte

From 30nm fiber (0.1 cm) to μm

Structure of a lampbrush chromosome

Sub-compartments within the nucleus

Early and late distribution in DNA synthesis. Newly synthesised

DNA is labeled with bromodeoxyuridine, a thymidine analog,

then detected by staining with a fluorescent antibody.

DNA synthesis starts at:

1- Perinucleolar sites ; 20 clusters distributed around

the nucleolus and associated with nuclear lamina

2- Origins of replication; about 4000 sites in a mammalian

cell, organized into distinct functional bodies called

replication factories.

Nuclear speckles

Storage sites of mRNA

splicing components (20-50

within a nucleous).

PML bodies

Sites of accumulation of

transcription factors and chromatin-

modifying proteins (such as histone

deacetylases)

Cajal bodies

Sites of small RNPs assembly

and processing.

Contain Coilin protein.

Green: Coilin

Red: Fibrillarin

Nucleolus and rRNA processing

Nucleolus is the site of

rRNA transcription and processing and ribosome assembly

RNA modification

Actively growing mammalian cells contain 5-10 million

ribosomes that must be synthesized each time the cell

divides.

Structure of the nucleolus

FC: fibrillar center

DFC: dense fibrillar component

G: granular component

Nucleoli in amphibian oocyte

Amlified rRNAs are

clustered in multiple nucleoli

o Ribosomal RNA genes and the organization of the nucleolus

RNA polymerase I

(200 copies)

Ch13,14,15,21,22

o Following each cell division,

nucleoli become associated

with the related chromosomal

regions. Therefore called

nucleolar organizing regions.

o 5S is present only on ch1

o 5S is transcript by RNA

polymerase III elsewhere

o Transcription and processing of rRNA

Electron micrograph of nucleolar chromatin showing 3 rRNA genes surrounded by growing RNA chains

Processing of pre-rRNA

ETS: external transcribed

spacers

ITS: internal transcribed spacers

snoRNPs U3

U8

U22

Pre-rRNA Processing (cont)

~300 proteins and 200 snoRNAs are located

in nucleoli and involved in pre-RNA

processing.

Role of snoRNAs in base modification of pre-RNA

snoRNAs contains short sequences of ~15

nucleotides that are complementary to 18S or 28S

rRNAs

o Ribosome assembly