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What do genes do?

What do genes do?

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What do genes do?

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  1. What do genes do? 15% • Chapter 17 - Protein Synthesis 21% 96% 7% 36%

  2. MOUSE EMBRYO GROWN FROM SKIN CELLS!

  3. Human Skin cells can be converted to Pluripotent (make 220 types of cells!) Embryonic Stem cells - Nov 2007! • How did they do it? • Skin cell and Embryonic stm cell - what’ the difference? • Which genes are active - many genes are silenced in the differentiated skin cell. • Turn on those genes needed to dedifferentiate the skin cells. • OOPS can it lead to problems? • Cancers! Virus vector used to deliver the genes can cause problems.

  4. What makes a plant short? T t Mutation in gene coding for Gibberellin – a plant growth hormone T t

  5. Mutation in gene coding for fibroblast growth factor– one of many factors needed for cell division What causes dwarfism? T t T t

  6. Early clues- The study of metabolic defects provided evidence that genes specify proteins Alkaptonuria –hereditary disease - change in metabolism due to lack of a particular enzyme (1909 - Garrod)

  7. Phenylthiocarbamide –PTC paper taste test Taster = dominant allele Nontaster = recessive allele Presence of gene = presence of taste receptor (protein) on tongue!

  8. 2amino –4hydroxypteridine à xanthopterin (blue) (green-blue) â tetrahydroiopterin (violet-blue) biopterin drosopterin sepiapterin(blue) (orange) (yellow) â isosepiapterin (yellow) • Mutation in eye color = synthesis of specific protein is blocked in a metabolic pathway (1930s)

  9. Beadle and Edward Tatum - link between genes and enzymes in bread mold, Neurospora crassa. • complete growth medium includes all 20 amino acids.

  10. ONE GENE ONE ENZYME HYPOTHESIS: A gene makes an enzyme (1941) Not always - gene products can be nonenzymatic proteins Also, a protein may have several POLYPEPTIDE chains (remember quarternary structure?) - each has its own gene. Changed hypothesis to one gene one polypeptide. BUT….

  11. ONE GENE ONE POLYPEPTIDE HYPOTHESIS 1 polypeptide

  12. Gene Therapy Within hours after doctors shot the normal OTC gene attached to a therapeutic virus into his liver, Jesse developed a high fever. His immune system began raging out of control, his blood began clotting, ammonia levels climbed, his liver hemorrhaged and a flood of white blood cells shut down his lungs.

  13. Transcription and translation are two main processes linking gene to protein • Genes provide the instructions for making specific proteins. • The bridge between DNA and protein synthesis is RNA. • The molecular chain of command in a cell is : DNA -> RNA -> protein.

  14. What are the differences in protein synthesis between prokaryotic and eukaryotic cells?

  15. RNA – Ribonucleic acid • RNA = Bases are adenine, guanine, cytosine, uracil (NO THYMINE) • Sugar is Ribose (not deoxyribose) • Mostly single stranded • Many types – mRNA, tRNA, rRNA

  16. Transcription • DNA strand provides a template for the synthesis of a complementary RNA strand. • Transcription of a gene produces a messenger RNA (mRNA) molecule.

  17. Transcription is the DNA-directed synthesis of RNA: a closer look • Messenger RNA is transcribed from the template strand of a gene. • Genes are read 3’->5’, creating a 5’->3’ mRNA molecule. Template Strand What will the mRNA code be? 5’ …A U G G C C U G G A C U U C A …. 3’

  18. Transcriptioncan beseparatedinto threestages:initiation, elongation, andtermination. Fig. 17.6a

  19. Use the HW assignment link for protein synthesis flash animation

  20. 1) Initiation (Promotor + transcription factors + RNA Polymerase) • RNA polymerase attaches at PROMOTOR SITE upstream of the gene/transcription unit

  21. 1) Initiation (unwinding of DNA strands + RNA synthesis begins) • RNA polymerase scans and recognizes the promotor sequence (TATA BOX) with the aid of proteins that can bind to DNA called - ‘transcription factors’. RNA polymerase separates the DNA strands and bonds the RNA nucleotides as they base-pair along the DNA template.

  22. 2)Elongation (RNA transcript elongates in 5’ -> 3’ direcion) • RNA polymerase moves downstream adding nucleotides to 3’ end of RNA transcript according to base paring rules => A-U; G-C.

  23. Fig. 17.7

  24. 3)Termination (RNA Polymerase reaches termination sequence) • RNA Polymerase and RNA transcript are released. Remember - this is premRNA in eukaryotes.

  25. Eukaryotic cells modify RNA after transcription • At the 5’ end of the pre-mRNA molecule, a modified form of guanine is added, the 5’ cap. (prevents breakdown and attch to ribosomes) • At the 3’ end, an enzyme adds 50 to 250 adenine nucleotides, the poly(A) tail. (protection from hydrolysis/breakdown of mRNA; helps export it) In eukaryotes this is still only pre-mRNA - the preview before the show!

  26. RNA splicing: Removal of Introns and attachment of exons • Noncoding segments, introns, lie between coding regions. • The final mRNA transcript includes coding regions, exons, that are translated into amino acid sequences, plus the leader and trailer sequences.

  27. This splicing is accomplished by a spliceosome. • spliceosomes consist of a variety of proteins and RNA (snRNPs). Important - the RNA acts as an enzyme in the spliceosome! Specific sequences are recognized on the DNA and introns cut out; exons joined. • Now m-RNA is READY for export to cytoplasm in eukaryotes - about time!

  28. All this to make mRNA… LE 19-5 Enhancer (distal control elements) Proximal control elements Poly-A signal sequence Termination region Exon Intron Exon Exon Intron DNA Upstream Downstream Promoter Transcription Poly-A signal Exon Intron Exon Exon Intron Primary RNA transcript (pre-mRNA) Cleaved 3¢ end of primary transcript 5¢ RNA processing: Cap and tail added; introns excised and exons spliced together Intron RNA Coding segment mRNA 3¢ Start codon Stop codon 5¢ UTR (untranslated region) 5¢ Cap Poly-A tail 3¢ UTR (untranslated region)

  29. Exon shuffling/Alternate splicing - makes different mRNA from a single transcription event

  30. WHY SPLICE mRNA (remove introns and join exons) in eukaryotes? • mRNA splicing appears to have several functions. • First, at least some introns contain sequences that control gene activity in some way. • Splicing itself may regulate the passage of mRNA from the nucleus to the cytoplasm. • One clear benefit of split genes is to enable one gene to encode for more than one polypeptide (EXON SHFFLING or ALTERNATE RNA SPLICING). (So what happened to the one gene one polypeptide hypothesis?) • Alternative RNA splicing gives rise to two or more different polypeptides, depending on which segments are treated as exons. • Split genes may also facilitate the evolution of new proteins/genes (important) - how?

  31. Thalassemia is due to splicing errors

  32. Translation • During translation, the information contained in the order of nucleotides in mRNA is used to determine the amino acid sequence of a polypeptide. • Translation occurs at ribosomes in both prokaryotes and eukaryotes.

  33. Transcription Translation

  34. Central Dogma • Transcription = DNA → RNA • Translation = RNA → protein • Taken together, they make up the "central dogma" of biology: DNA → RNA → protein.

  35. In the genetic code, nucleotide triplets specify amino acids • In the triplet code, three consecutive bases specify an amino acid, creating 43 (64) possible code words. Why not use 2 bases? • Genetic code is ‘triplet and degenerate’ - why? AAAAAAAAAAAAAAAAAA - artificial mRNA Phe – Phe – Phe – Phe – Phe – Phe Khorana and Nirenberg (1960) - which triplet code for which aminoacid. {Crick, Brenner - “ do you realize - we are the only two people in the world who know it is a triplet code?”} Make your own protein: http://gslc.genetics.utah.edu/units/basics/transcribe/

  36. 61 of 64 triplets code for 20 amino acids.

  37. How many nucleotides will it take to code for a polypeptide that is 100 amino acids long? • It would take more than 300 nucleotides to code for a polypeptide that is 100 amino acids long. • Why not ‘exactly 300 nucleotides’? • Some codes specify start (AUG) and stop signals (UGA, UAA, UAG) • Some specify other sequences needed to get transcription rolling (promotor)

  38. The genetic code is redundant/degenerate but not ambiguous. • There are typically several different codons that would indicate a specific amino acid. • However, any one codon indicates only one amino acid. • [If you have a specific codon, you can be sure of the corresponding amino acid, but if you know only the amino acid, there may be several possible codons.] • Both GAA and GAG specify glutamate, but no other amino acid. • Codons synonymous for the same amino acid often differ only in the third codon position.

  39. To extract the message from the genetic code requires specifying the correct starting point. • This establishes the reading frame and subsequent codons are read in groups of three nucleotides. • The cell’s protein-synthesizing machinery reads the message as a series of nonoverlapping three-letter words.

  40. The genetic code must have evolved very early in the history of life • The genetic code is nearly universal, shared by organisms from the simplest bacteria to the most complex plants and animals.

  41. What Have They Done To Our Food? – (60 Minutes- 2001) Up to 70 percent of processed food in the American market contains products of genetic engineering, including soft drinks, catsup, potato chips, cookies, ice cream and corn flakes.

  42. Translation is the RNA-directed synthesis of a polypeptide: a closer look Characters: • mRNA – messenger carrying code for amino acid sequence • tRNA – transfer RNA reads RNA code and transcribes it into aminoacid sequence • Ribosomes – docking station for mRNA and tRNA tRNA mRNA Fig. 17.12

  43. Each tRNA has 2 parts: • A code reading part: called ANTICODON • An amino acid attachment site How many nucleotides will each tRNA use in its ANTICODON? Why? How many different tRNA molecules will be needed? Why? One for every codon dictates 61, but there are 45 tRNAs