1. Dr. Faisal Al-AllafAssistant Professor of Geneticsand Molecular MedicineUmm Al-Qura University Faculty ofMedicine, Makkah, Saudi Arabiafallaf@uqu.edu.saTel/Fax: 5270000 Ext: 4198The Cellular andMolecular Basisof Inheritance1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 1

2. . . . . :http://el.uqu.edu.sa/jusur/index.php?un_id=uqu :http://www.uqu.edu.sa/faallaf 40/40/1341Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa2 3. Course contents (syllabus)GENOME, TRANSCRIPTOME, AND PROTEOMECell, DNA and RNAGene structure and genetic codesCell cycle and DNA replicationTranscription and post-transcriptional modificationRNA and regulation of gene expressionTranslation and post-translational modificationCHROMOSOMES AND CELL DIVISIONChromosomes morphology and classificationCell cycle divisionMitosisMeiosis1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 3 4. How genetic information is transmittedfrom one generation to the next?The body grow by increasing in cellsize and cell number (mitosis).Body growth involves individualcells replicating their components,dividing in half, expanding anddoing the same again.This sequence is called the cellcycle and it involves two criticalevents: Replication of chromosomal DNA Segregation of the duplicated chromosomes by mitosis1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 4 5. The cell cycleCell proliferate in response toextracellular factors (growth factors,hormones, and cell-cell interactions).The cell cycle is driven by activationand deactivation of enzymes knownas cyclin-dependent protein kinasesCell cycle has four major phases:1. Interphase (G1, S, G2)2. Mitosis (M) phase 1431/04/04Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 5 6. G1 phase is the Gap betweenM- and S-phasesDuring G0 the cell decides to divide,die, or stay differentiatedG1-phaseThe cytoplasm increases in volume(proteins, carbohydrates and lipidsynthesized)Damage to the DNA is repaired. If thenuclear DNA is damaged, a proteincalled p53 increases in activity andstimulate p21 to arrest the cells at G1phase. If the repair failed the cellcommitted suicide (apoptosis)The cell checks that its environmentis favourable before committing itselfto S-phase1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 6 7. S-phase is the DNA synthesisphase in which DNA copy itself S-phase DNAsynthetic enzymes activated, DNA replicated, and centriols begin to duplicate From the end of S-phase to the middle of M phase there are 23 pairs of chromosomes, each contain two sister chromatides (2C) At the middle of M, there are 23 pairs of chromosomes but 92 nuclear DNA double-helices (4C) G1 and G0 are the only phases of the cell cycle throughout which 46chromosomes correspond to 2C DNA molecules S-phase lasts about 8 hours1431/04/04Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 7 8. G2-phase is the gap between S-phase and mitosisG2-phaseCompletion of DNA synthesischeckedCell volume checkedM-phase cyclin dependentkinases activatedCentrioles duplication completeThe second checkpoint on cellsize occurs during G21431/04/04Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 8 9. Three checkpoints in the cell cycleG1/S checkpoint: at the end ofG1thecellbecomescommitted to completing acycle of division. The cell willnot proceed beyond this pointif thereareinadequatenutrients or growth factorsG2/M checkpoint: cells will notproceed beyond this point ifthere is DNA damage(s)M/G2 checkpoint: cells willbecome arrested at this stageif the mitotic spindle fails toassemble adequately1431/04/04Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 9 10. DNA replication is initiated atorigins of replicationHeterochromatin replicates later in S-phase than euchromatinReplication is initiated at multiplepoints known as Replication originsThe DNA double helix is split openby a DNA helicase to expose thebase sequencesThe resulting separations of the DNAstrand are called Replication bubblesTwo replication complexes form ateach origin, one at each end of thebubble and is a Y-shaped structurecalled Replication fork1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 10 11. Replication is mediated by enzymescalled DNA polymerasesDNA polymerases synthesized new DNA fromdeoxyribonucleotide triphosphates (ATP, GTP, CTP,and TTP) which are converted intomononphosphate nucleotides (AMP, GMP, CMPand TMP)The release and hydrolysis of phosphate from thetriphosphates provide energy for the reaction and itensures that it is irreversibleUnlike RNA polymerase, DNA polymerase has anabsolute requirement for a perfectly base-pairednucleotide which has important consequences:DNA polymerase requires a primer on which to initiateextensionIt will pause if an incorrect base is insertedThe primers are synthesized by RNA polymerasePrimers, energy, templates, enzymes (polymerases,helicase, ligase, topoisomerase, primase,telomearse)1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 11 12. DNA synthesis of both strandsoccurs in the 5` to 3` direction DNA can copy itself and each strand directing the synthesisof a complementary one The synthesis of both new complementary strands occurs in the 5` to 3` direction; meaningthat replication moving along the template strands from 3 to 5 There are three stages for DNA replication: 1. Initiation 2. Elongation 3. Termination1431/04/04Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 12 13. Initiation of DNA replication1. Helicase binds to origin ofreplicationand unwind(separates) parental strands2. Single-strand binding proteins(SSBs) keep strands apart3. Primase binds to the helicaseto form the primosome whichsynthesizes the RNA primer (ashort stretch of RNA on theDNA template)1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 13 14. Elongation of DNA1. DNA polymerase clamps to theleading strand and adds DNAnucleotides to the RNA primer2. DNA polymerase proofreadingactivity checks and replacesincorrect bases3. Continuous (leading) strandsynthesis produces newfragments on the 3 to 5template but in a 5 to 3direction4. Discontinuous (lagging) strandsynthesis produces Okazakifragments on the 5 to 3template but in a 5 to 3direction5. Replication proceeds along the single strands at about 40-50 nucleotides per second, simultaneously in both directions 1431/04/04Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 14 15. DNA replication fork The new sections of DNA along the lagging strand are typically 100-200 bases long and are known as Okazaki fragments Leading strand synthesized in a continuous process from a single RNA primer by DNA polymeraseand lagging strand synthesized in pieces called Okazaki fragment from multiple RNA primers by DNA polymerase The strand that is leading and the strand that is lagging depends on the direction the replication complexes is migrating Following synthesis of lagging strands, they are linked together by action of the enzyme DNA ligase Note that mitochonderial DNA replication occurs independently of that in the nucleus and utilizes a different set of enzymes1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 15 16. Replication fork1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 16 17. Termination of DNA replicationEnzymesremove RNAprimersDNA ligase joins upadjacent Okazakifragments and seals othernicks in the sugar-phosphate backbone1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 17 18. DNA replication is semi-conservative The process of DNA replication is termed semi-conservative, as only one strand of each serves as a template for synthesis of a new strand Therefore, only one strand is conserved in each double- stranded DNA molecule After the replication fork passes, chromatin structure is reformed by the addition of new histones1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 18 19. Post-replication repair enzymes andbase mismatch proofreading systemsUsing the template strandas a guide, the followingenzymes are responsiblefor repair mechanism:Endonucleasedetectsmismatch or errorExonuclease removesnucleotides from nick topast defectPolymerase ( and )replace the erroneouslyinserted basesDNA Ligase rejoin thedouble strands1431/04/04Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 19 20. 1431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 20 21. References and Private Reading These slides are only a handout and the students must read the text book (Emeryselement of medical genetics)1. Emerys Elements of Medical Genetics, 13th edition 2007, by Peter TURNPENNY and Sian ELLARD. Churchill Livingstone ELSEVIER. ISBN: 978-0-7020-2917-22. Medical Genetics at a Glance, 2nd edition 2008, by Dorian PRITCHARD and Bruce KORF. Blackwell Publishing. ISBN: 978-1-4051-4846-73. Genetics for Dummies, 2005, by Tara Robinson, Wiley Publishing, Inc. ISBN: 978- 0-7645-9554-74. Cell Biology and Genetics, Crash Course, 2nd edition 2006, by Manson, Jones, Morris, Michael STEEL and Dan HORTON-SZAR. MOSBY ELSEVIER. ISBN: 0- 7234-3248-15. Human Molecular Genetics, 3rd edition, 2003, by STRACHAN T. and A. READ. Garland science/Taylor and Francis group. ISBN: 978-0-8153-4182-66. Genomes, 3rd edition 2006, by T.A. BROWN. Garland science, ISBN: 978-0-8153- 4138-31431/04/04 Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa21 22. Acknowledgments For the providers of all the educational materials (video clips, pictures, diagrams and charts) including publishers, pharmaceutical companies or unknown internet users who made their material available for use, in this and other presentations, I offer heartfelt thanks and deep appreciation. I feel particularly grateful to faculty, staff, and our brilliant students who provided a unique intellectual and wonderful environment for work.1431/04/04Dr. Faisal Al-Allaf, fallaf@uqu.edu.sa 22