Lecture 4 Genome structure & Repair.pptx

Genome structure and DNA replication

1. Molecular Biology Lecture 4 Human genome structure & DNA Repair D r . Emad Ibrahim Osman

2. Structural organization of eukaryotic DNA A typical human cell contains 46 chromosomes, whose total DNA is approximately 1m long. That is a lot to pack into a little nucleus, so Eukaryotic DNA is highly packaged

3. Eukaryotic DNA p a c k i n g Eukaryotic DNA exhibits many levels of packaging The fundamental unit is the nucleosome, DNA wound around histone proteins Nucleosomes arrange themselves together to form higher and higher levels of packaging.

4. Nucleosomes The lowest DNA packaging level Can be thought of as like a length of thread wound around a spool, the thread representing DNA and the spool being histone proteins

5. Nucleosome Structure Approximately 200 bp of DNA: Core DNA – (140-150) bp associated with the histone octomer 19 bases complete the two turns around the histone octomer Linker DNA – ( 30-50) bp linking nucleosomes together

6. The Histone Octomer Four proteins: H2A, H2B, H3, and H4 H3 and H4 are arginine rich and highly conserved H2A and H2B are slightly enriched in lysine Both arginine and lysine are basic amino acids making the histone proteins both basic and positively charged The octomer is made of two copies of each protein

7. The Fifth Histone, H1 A fifth protein, H1, is part of the nucleosome, but seems to be outside the octomer H1 varies between tissue and organisms and seems to stick to the 19 bases attached to the end of the core sequence Ausio J

8. Packaging DNA G C A T Protein scaffold Metaphase Chromosome 700 nm 11 nm 30 nm 200 nm 2 nm Looped Domains Nucleosomes B DNA Helix Tight helical fiber

9. The Packaged DNA Chromatin=DNA+RNA+ histone & nonhistone protein. Transcriptionally inactive chromatin is densely packed and is referred to as heterochromatin; Transcriptionally active chromatin stains less densely and is referred to as euchromatin. Most highly packaged form of DNA is “heterochromatin” Heterochromatin cannot be transcribed, therefore expression of genes is prevented

10. DNA D NA is either 1. Genes (3%)as segments of DNA that are transcribed may code for both polypeptides or RNAs 2. Non gene “ Junk DNA ”(97%) is DNA that does not code for proteins. - Nine different types of DNA were listed as junk DNA - These nine types can be grouped into three larger groups: A) Repetitive DNA sequences B) Untranslated parts of RNA transcripts (pre-mRNA) C) Other non-coding sequences

11. A) Repetitive DNA Repeated sequences seem too short to code for proteins and are not known to be transcribed. The major classes of repetitive DNA: Satellites - Up to 10 5 tandem repeated short DNA sequences, concentrated in heterochromatin at the ends (Telomeres) and centers (centromeres) of chromosomes. ( used in DNA finger printing) Minisatellites - Similar to satellites, but found in clusters of fewer repeats, scattered throughout the genome Microsatellites - Shorter still than minisatellites.

12. B) Untranslated Parts of mRNA Not all of the pre-mRNA transcribed from DNA actually codes for the protein. These non-coding parts are never translated. Three non-coding parts of eukaryotic mRNA: 1) 5' untranslated region 2) Introns - Segments of DNA that are transcribed into RNA, but are removed from the RNA transcript before the RNA leaves the nucleus as mRNA 3) 3' untranslated region

13. A “Simple” Eukaryotic Gene Transcription Start Site Terminator Sequence 3’ Promoter/ Control Region 5’ Introns RNA Transcript 5’ Untranslated Region 3’ Untranslated Region Exon 2 Exon 3 Int. 2 Exon 1 Int. 1 Exons

14. Other Non-coding Sequences Pseudogenes - DNA that resembles functional genes, but is not known to produce functional proteins. Two types: Unprocessed pseudogenes Processed pseudogenes Heterogeneous Nuclear RNA - A mixture of RNAs of varying lengths found in the nucleus. Approximately 25 % of the hnRNA is pre-mRNA that is being processed, the source and role of the remainder is unknown.

15. DNA REPAIR

16. DNA REPAIR Despite the proofreading system during DNA synthesis, errors can occur. In addition, DNA is constantly being subjected to environmental insults that cause the alteration or removal of nucleotide bases. The damaging agents can be either chemicals, for example, nitrous acid, or radiation (e.g. UV) which make pyrimidines dimer

17. Bases are also altered or lost spontaneously from mammalian DNA at a rate of many thousands per cell per day. If the damage is not repaired, a permanent change (mutation) is introduced that can result in any of a number of deleterious effects, including loss of control over the proliferation of the mutated cell, leading to cancer. Luckily, cells are remarkably efficient at repairing damage done to their DNA.

18. Most of the repair systems involve recognition of the damage (lesion) on the DNA, removal or excision of the damage, replacement or filling the gap left by excision using the sister strand as a template for DNA synthesis, and ligation. These repair systems thus perform excision repair, with the removal of one to tens of nucleotides. Repair reduces the error rate from one in ten million bases to one in a billion.

19. Types of DNA repair mechanisms 1- Base excision repair - Correction of base alterations 2- Repair of damage caused by ultraviolet (UV) light 3- Methyl-directed mismatch repair 4- Repair of double-strand breaks

20. Correction of base alterations (base excision repair) The bases of DNA can be altered, undergoes deamination to form uracil either spontaneously or by the action of alkylating compounds Most DNA damage can be corrected by excision repair involving recognition and removal of the damage by repair proteins, followed by replacement by DNA polymerase and joining by ligase.

21. Repair of damage caused by ultraviolet (UV) light Ultraviolet light can cause thymine dimers that are recognized and removed by uvrABC proteins of nucleotide excision repair. Defects in the XP proteins needed for thymine dimer repair in humans result in xeroderma pigmentosum.

22. Methyl-directed mismatch repair Mismatched bases are repaired by a similar process of recognition and removal by Mut proteins in E. coli. The extent of methylation is used for strand identification in prokaryotes. Defective mismatch repair by homologous proteins in humans is associated with hereditary nonpolyposis colorectal cancer HNPCC .

23. Repair of double-strand breaks Abnormal bases (such as uracil) are removed by glycosylases in base excision repair, and the sugar phosphate at the AP site is cut out. Double-strand breaks in DNA are repaired by nonhomologous end-joining (error prone) and homologous recombination.

24. T h e E n d T h a n k s