Cell and Molecular Biology Chromosomes and Chromatin Behrouz Mahmoudi

The Problem Human genome (in diploid cells) = 6 x 109 bp 6 x 109 bp X 0.34 nm/bp = 2.04 x 109 nm = 2 m/cell Very thin (2.0 nm), extremely fragile Diameter of nucleus = 5-10 mm DNA must be packaged to protect it, but must still be accessible to allow gene expression and cellular responsiveness

Chromosomes Single DNA Molecule and associated proteins Karyotype: The characterization of the chromosome complement of a species (such as the shape, type, number, etc. of chromosomes). Chromatin vs. Chromosomes Single DNA Molecule and associated proteins Evident during metaphase of Mitosis and Meiosis Karyotype---A karyotype is the complete set of all chromosomes of a cell of any living organism. The chromosomes are arranged and displayed (often on a photo) in a standard format: in pairs, ordered by size. Karyotypes are examined in searches for chromosomal aberrations, and may be used to determine other macroscopically visible aspects of an individual's genotype, such as sex (XX vs. XY pair). Technically, chromosomes are the duplicated, condensed form of chromatin, but the terms are used somewhat interchangeably.

HISTONES Main packaging proteins 5 classes: H1, H2A, H2B, H3, H4. proteins that DNA tightly coils around to form chromosomes. Main packaging proteins 5 classes: H1, H2A, H2B, H3, H4. Rich in Lysine and Arginine Main packaging proteins---found in eukaryotes and some Archaes Mass of histones in nucleus = mass of DNA The (+) charges are instrumental in the recognition process, binding by electrostatic interactions to the phosphate backbone. HOW CAN THEY BE REMOVED? Sequence independent contacts mediate interactions between core histones and DNA. DNA is bent around histone core. Basis for bending is H-bonding between histones and contacts in minor groove (140 H-bonds, primarily between histone and O of phosphates in backbone in minor groove, 7 between R-groups and bases in minor groove). Note: average DNA interacting protein has 20 H-bonds. Basic nature of histones masks negative phosphates, allows bending. binding is strong but in areas with little info, is non-specific.

Compacting DNA strands Functions Compacting DNA strands Histones act as spools around which DNA winds. This enables the compaction necessary to fit the large genomes of eukaryotes inside cell nuclei: the compacted molecule is 40,000 times shorter than an unpacked molecule. Chromatin regulation Histones undergo posttranslational modifications that alter their interaction with DNA and nuclear proteins. Modifications of the tail include methylation, acetylation, phosphorylation, ubiquitination. Histone modifications act in diverse biological processes such as gene regulation, DNA repair, chromosome condensation (mitosis) and spermatogenesis (meiosis).

DNA condensation The double helix of DNA is highly negatively charged due to all the negatively charged phosphates in the backbone. All that negative charge must be counterbalanced by a positive charge, and the cell makes histones that bind DNA and aid in DNA's packaging Histones are positively charged proteins that wrap up DNA through interactions between their positive charges and the negative charges of DNA

Fig. 9

The Nucleosome: The Unit of Chromatin The basic repeating structural (and functional) unit of chromatin is the nucleosome, which contains nine histone proteins and about 166 base pairs of DNA Two each of the histones H2A, H2B, H3, and H4 come together to form a histone octamer, which binds and wraps about 1.7 turns of DNA, or about 146 base pairs. The addition of one H1 protein wraps another 20 base pairs, resulting in two full turns around the octamer

Nucleosomes, with about 20 to 60 base pairs of linker DNA, can form, an approximately 10 nm "beads-on-a-string" fibre. The solenoid defines the packing of DNA as a 30 nm fiber of chromatin and results from the helical winding of at least five nucleosome strands DNA packed into solenoids, unlike DNA in nucleosome form, is not transcriptionally active solenoids coil around each other to form a loop (anywhere from 20 to 80,000 base pairs), followed by a rosette (consisting of six connected loops), then a coil, and at last, two chromatids solenoids are able to become increasingly more packed, forming chromosomes

30 nm Fiber 30 nm fiber is coil of nucleosomes with 6/turn Figure: 20.19 The next level of compaction Title: The 30 nm thread is a coiled coil Caption: The 30 nm fiber has a coiled structure. Photograph kindly provided by Barbara Hamkalo. 30 nm fiber is coil of nucleosomes with 6/turn

E H Euchromatin (E) vs Heterochromatin (H) Heterochromatin is a tightly packed form of DNA, which comes in different varieties. These varieties lie on a continuum between the two extremes of constitutive and facultative heterochromatin Constitutive heterochromatin is usually repetitive and forms structural functions such as centromeres or telomeres Facultative heterochromatin is not repetitive

Euchromatin is a lightly packed form of chromatin (DNA, RNA and protein) that is rich in gene concentration, and is often (but not always) under active transcription. Euchromatin comprises the most active portion of the genome within the cell nucleus. 92% of the human genome is euchromatic.

The centromere is the part of a chromosome that links sister chromatids. During mitosis, spindle fibers attach to the centromere via the kinetochore kinetochore - a highly complex multiprotein structure that is responsible for the actual events of chromosome segregation

A telomere is a region of repetitive nucleotide sequences at each end of a chromatid, which protects the end of the chromosome from deterioration or from fusion with neighboring chromosomes

NOTE: if histones from different species are added to any eukaryotic DNA sample, chromatin is reconstituted. Implication? NOTE: if histones from different species are added to any eukaryotic DNA sample, chromatin is reconstituted. Implication? Very highly conserved in eukaryotes in both Structure—very similar aa composition, most substitutions are conservative. Function Protection Must allow function QUESTION: CAN THEY AFFECT GENE EXPRESSION?