Molecular Biology 2024 - BIOL10221 Lecture Notes

BIOL10221 Molecular Biology Module 3 - Lecture 2: Genomes II Ray O’Keefe [email protected] What will we cover? Higher order packaging of DNA – chromatin, histones, 30-nm fibres Euchromatin and Heterochromatin Centromeres and telomeres Why is it important? Without DNA packa...

BIOL10221 Molecular Biology Module 3 - Lecture 2: Genomes II Ray O’Keefe [email protected] What will we cover? Higher order packaging of DNA – chromatin, histones, 30-nm fibres Euchromatin and Heterochromatin Centromeres and telomeres Why is it important? Without DNA packaging the cell will never be able to accommodate the large amount of DNA needed in a cell Different states of DNA packaging allow proper expression of genes Unique features of chromosomes allow their proper maintenance and segregation during cell division BIOL10221 Module 3 - Lecture 2 2 The human genome comprises 3200 Mb DNA split into 24 linear DNA molecules shortest is 48 Mb longest is 250 Mb each one in a different chromosome In a normal diploid cell there are 46 chromosomes (Two sets of Chromosomes 1-22 and either XX or XY) 6400 Mb DNA! BIOL10221 Module 3 - Lecture 3 2 This is human chromosome 8 It is about 3.5 µm in length = 0.0035 mm It contains a single DNA molecule of 145 Mb This molecule is 4.9 cm in length How is the DNA packaged into the chromosome? BIOL10221 Module 3 - Lecture 4 2 DNA extracted from the nucleus is called Chromatin digestion with an endonuclease shows that chromatin is a DNA-protein complex with the proteins spaced at regular intervals along the DNA BIOL10221 Module 3 - Lecture 5 2 This endonuclease digestion result agrees with electron microscopy of chromatin which shows a beads-on-a-string structure BIOL10221 Module 3 - Lecture 6 2 The proteins in chromatin are called histones BIOL10221 Module 3 - Lecture 7 2 The beads are called nucleosomes each nucleosome is an octamer or 8 proteins two each of histones H2A, H2B, H3 and H4 BIOL10221 Module 3 - Lecture 8 2 Histone H1 is the linker histone it attaches outside the nucleosome nucleosome + DNA + linker histone = chromatosome BIOL10221 Module 3 - Lecture Cryo-EM 9 2 Beads on a string reduces the length of a DNA molecule to one-sixth there must be higher levels of packaging The next level is the 30 nm chromatin fibre the 4.9 cm molecule in chromosome 8 would now be about 1.2 mm in length side view top view BIOL10221 Module 3 - Lecture 10 2 Cryo-EM structure of 30nm chromatin fibres Nucleosomes are stacked only with non-neighbors and zig-zag back and forth connected by a straight DNA linker Tetranucleosomal units highlighted with different colours BIOL10221 Module 3 - Lecture 11 2 Cryo-EM of linker histone H1 and interactions between the nucleosome units in the 30 nm chromatin fibre Off-axis asymmetric binding of histone H1 stabilises 30nm fibre Interaction between the histone H4 acidic N-terminal tail and H2A/H2B acidic patch important for 30nm fibre formation BIOL10221 Module 3 - Lecture 12 2  The Importance of Histones/Nucleosomes Histones allow the vast amount of DNA to be packed into nucleosomes and other higher order DNA structures Chromatin structure can be changed through the reversible chemical modification of histones DNA packaging into nucleosomes is not permanent but dynamic Nucleosomes detach or shift to allow transcription of DNA Nucleosomes must detach for replication of DNA Nucleosome presence or modification can control gene expression BIOL10221 Module 3 - Lecture 13 2  Guinea pig plasma cell – non-dividing nucleolus contains ribosomal RNA genes, euchromatin (light areas) site of ribosome biogenesis contains active genes probably as 30 nm fibre heterochromatin (dark areas) contains inactive genes more densely packed constitutive heterochromatin contains DNA that is always tightly packed – in all cells facultative heterochromatin contains DNA that is tightly packed only in some cells BIOL10221 Module 3 - Lecture 14 2 In euchromatin, DNA is in the form of the 30 nm fiber or less compact attached to the nuclear matrix (inside nucleus) and nuclear lamina (around edge of nucleus) so the DNA does not get tangled up maintains the shape of the nucleus nuclear matrix and nuclear lamina associated diseases Progeria – premature aging Down’s Syndrome Huntington disease BIOL10221 Module 3 - Lecture 15 2 Mechanisms of higher levels of packaging are not understood the highest level - the metaphase chromosome – is only found in dividing cells BIOL10221 Module 3 - Lecture 2 16 Dividing bluebell cells BIOL10221 Module 3 - Lecture 17 2 The metaphase chromosome is the highest level of packaging The centromeres hold the daughter chromosomes together contains special histones – CENP-A instead of H3 attachment points for microtubules that pull chromosomes apart The telomeres protect the ends from exonuclease attack from being mistaken for chromosomes breaks and joined together by DNA repair mechanisms BIOL10221 Module 3 - Lecture 18 2 Metaphase chromosomes can be stained to give a banding pattern The complete set is the called the karyogram the human karyogram All bands have names used for mapping positions of genes Centromeres have variable positions Defines long and short arms Constitutive heterochromatin blue No active genes Ribosomal DNA (rDNA) repeats together BIOL10221 Module 3 - Lecture 19 2 Resources Reading BROWN, Chapter 11 pages 211–223 ALBERTS (5th ed), Chapter 5 pages 178-192 ALBERTS (6th ed), pages 184-200 Review paper https://iubmb.onlinelibrary.wiley.com/doi/epdf/10.100 2/iub.1568 PDF on Blackboard Blackboard PDF files with Objectives, Terms, Questions and Answers BIOL10221 Module 3 - Lecture 20 2  BIOL10221 Molecular Biology Module 3 - Lecture 2: Genomes II Ray O’Keefe [email protected]