II. INTERNAL ORGANIZATION OF EUKARYOTIC CELLS

Structure of a „typical” animal cell

Structure of a „typical” plant cell

II.1.4. Nucleus Biological significance: storage, expression and transmission of the genetic information.

Segmented nuclei of leukocytes neutrophils

Structural components

II Nuclear membrane (envelope) cytoplasm nuclear membrane nuclear lamina chromatin nuclear pore Biological significance: limiting layer between the inner part of the nucleus and the cytoplasm. Specific transport of micro- and macromolecules.

nuclear envelope perinuclear space The nuclear envelope is formed from two concentric membranes that are continuous with the endoplasmic reticulum (ER). The space between the two membranes is the perinuclear space which is continuous with the lumen of the ER. nuclear lamina The inner nuclear membrane contains proteins which enable it to bind to the nuclear lamina (network of intermediate filaments), which binds the chromatin. nuclear pores The double-membrane envelope is penetrated by nuclear pores.

Nuclear pore EM image Construction surface wiev side wiev

Relationship of the nucleus and the cytosol

II Chromatin The chromatin is composed of deoxyribonucleic acids and proteins. In interphase they are arranged in a relatively loose, „network” form. EM shows that there are two different forms of the chromatin, one appearing as a light element (euchromatin), the other one being dark (heterochromatin). Heterochromatin Heterochromatin: Closely packed chromatin. The DNA is organized by histon and non-histone proteins. Euchromatin: Here the chromatin is loosely arranged. The DNA double helices are partially split to single strands by RNA polymerases.

Euchromatic nucleus of neuron

H2A, H2B, H3 and H4 (two molecules of each) are building the Histone-Octamer. The DNA double helix makes two turns around this octamer (166 base pairs). H1 binds to theis complex. (packing level: 6-7x) H1 Octamer Nucleosome 1. Level of organisation: Nucleosome Structure The chromatin is a DNA-protein complex About half of the chromatin is protein: they are either histones or non-histone proteins. Histones Histones are small, positively charged proteins, which bind non-specifically to DNA end compensate the negative charge of phosphate groups of the DNA. Histone classes: H1, H2A, H2B, H3 and H4.

30 nm chromatin fiber 2. Level of organisation : 30 nm chromatin fiber The nucleosomes are spirally organized around the H1 core (packing level: ~ 40 x) This structure is broken to segments by binding of non-histone proteins (e.g. gene regulator proteins).

300 nm chromatin fibers 3. Level of organisation: 300 nm chromatin fibers 30 nm chromatin fiber loop domains base pairs 300 nm

Chromosomes W. Waldeyer, 1888: " …thick chromatin structures which can be seen during mitosis and meiosis...„ Name: chroma: color and soma: body Boveri and Sutton, Chromosome theory: the chromoses are the carriers of the genetic information.. Chromosomes exists only during cell division as microscopically detecteble units. At the start of the mitosis (M phase) each chromose has replicated and consists of two sister chromatids joined together

Fine structure Chromatids: 300 nm chromatin fiber form spirals (packing level: x) 300 nm 700 nm

Staining of the chromosomes Q-bands Fluorescent stains (e.g. Quinacrin): Q-bands specific for AT-rich DNA regions G-bands R-bands Giemsa-staining:according to the technique applied, either G-bands ("Giemsa-positiv") specific for AT-rich DNA-regions or R-bands ("Giemsa-negativ") specific for GC-rich DNA-regions

Chromosomes The human karyotype

fruitfly 8 house fly 12 ant 48 frog 26 carp 104 dog 38 cat 64 mouse40 rat42 cattle60 rhesus monkey 42 human 46 baker’s yeast32 corn20 rice 24 wheat 42 tobacco48 Chromosomal organization of different species

Variability in chromosomal organization. Chromosomal organization (karyotype) can show high variability even in closely related species. The number of genes is equal! Reeves deer Indian deer

Scanning-EM, human chromosomes Schematic view of chromosomal regions X45 T e l o m er shorter arm „p” longer arm „q” centromer sister chromatids Structure of the chromosomes

Telomers, aging, cancer (and Dolly) Human telomer: base pairs of repeated TTAGGG DNA sequences. Shortening: base pairs/division. In germline cells: telomere terminal transferase (telomerase): ribonucleoprotein reverse transcriptase.

Binding of the chromosomes to the mitotic spindle Attachment site: kinetochor kinetochor at the centromer

Preparation of chromosomes, karyotype determination Blood culture (phytohaemagglutinin) colchicin separation of erythrocytes and white blood cells fixation staining microphoto arrange according to size, centromer position and banding pattern