1. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings PowerPoint Lectures for Biology, Seventh Edition Neil Campbell and Jane Reece Lectures by Chris Romero Chapter 19 Eukaryotic Genomes: Organization, Regulation, and Evolution

2. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Overview: How Eukaryotic Genomes Work and Evolve Two features of eukaryotic genomes are a major information-processing challenge: – First, the typical eukaryotic genome is much larger than that of a prokaryotic cell – Second, cell specialization limits the expression of many genes to specific cells The DNA-protein complex, called chromatin, is ordered into higher structural levels than the DNA- protein complex in prokaryotes

3. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings

4. Concept 19.1: Chromatin structure is based on successive levels of DNA packing Eukaryotic DNA is precisely combined with a large amount of protein Eukaryotic chromosomes contain an enormous amount of DNA relative to their condensed length

5. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Nucleosomes, or “Beads on a String” Proteins called histones are responsible for the first level of DNA packing in chromatin The association of DNA and histones seems to remain intact throughout the cell cycle In electron micrographs, unfolded chromatin has the appearance of beads on a string Each “bead” is a nucleosome, the basic unit of DNA packing Animation: DNA Packing Animation: DNA Packing

6. LE 19-2a DNA double helix Histone tails His- tones Linker DNA (“string”) Nucleosome (“bead”) 10 nm 2 nm Histone H1 Nucleosomes (10-nm fiber)

7. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Higher Levels of DNA Packing The next level of packing forms the 30-nm chromatin fiber

8. LE 19-2b 30 nm Nucleosome 30-nm fiber

9. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings In turn, the 30-nm fiber forms looped domains, making up a 300-nm fiber

10. LE 19-2c 300 nm Loops Scaffold Protein scaffold Looped domains (300-nm fiber)

11. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings In a mitotic chromosome, the looped domains coil and fold, forming the metaphase chromosome

12. LE 19-2d Metaphase chromosome 700 nm 1,400 nm

13. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Interphase chromatin is usually much less condensed than that of mitotic chromosomes Much of the interphase chromatin is present as a 10-nm fiber, and some is 30-nm fiber, which in some regions is folded into looped domains Interphase chromosomes have highly condensed areas, called heterochromatin, and less compacted areas, called euchromatin

14. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Concept 19.2: Gene expression can be regulated at any stage, but the key step is transcription All organisms must regulate which genes are expressed at any given time A multicellular organism’s cells undergo cell differentiation, specialization in form and function

15. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Differential Gene Expression Differences between cell types result from differential gene expression, the expression of different genes by cells within the same genome In each type of differentiated cell, a unique subset of genes is expressed Many key stages of gene expression can be regulated in eukaryotic cells

16. LE 19-3 Signal NUCLEUS DNA RNA Chromatin Gene available for transcription Gene Exon Intro Transcription Primary transcript RNA processing Cap Tail mRNA in nucleus Transport to cytoplasm CYTOPLASM mRNA in cytoplasm Translation Degradation of mRNA Polypeptide Cleavage Chemical modification Transport to cellular destination Degradation of protein Active protein Degraded protein

17. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Regulation of Chromatin Structure Genes within highly packed heterochromatin are usually not expressed Chemical modifications to histones and DNA of chromatin influence both chromatin structure and gene expression

18. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Histone Modification In histone acetylation, acetyl groups are attached to positively charged lysines in histone tails This process seems to loosen chromatin structure, thereby promoting the initiation of transcription

19. LE 19-4 Histone tails Amino acids available for chemical modification DNA double helix Histone tails protrude outward from a nucleosome Acetylation of histone tails promotes loose chromatin structure that permits transcription Unacetylated histones Acetylated histones

20. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings DNA Methylation DNA methylation, the addition of methyl groups to certain bases in DNA, is associated with reduced transcription in some species In some species, DNA methylation causes long- term inactivation of genes in cellular differentiation In genomic imprinting, methylation turns off either the maternal or paternal alleles of certain genes at the start of development

21. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Epigenetic Inheritance Although the chromatin modifications just discussed do not alter DNA sequence, they may be passed to future generations of cells The inheritance of traits transmitted by mechanisms not directly involving the nucleotide sequence is called epigenetic inheritance

22. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Regulation of Transcription Initiation Chromatin-modifying enzymes provide initial control of gene expression by making a region of DNA either more or less able to bind the transcription machinery

23. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Organization of a Typical Eukaryotic Gene Associated with most eukaryotic genes are control elements, segments of noncoding DNA that help regulate transcription by binding certain proteins Control elements and the proteins they bind are critical to the precise regulation of gene expression in different cell types

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

25. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The Roles of Transcription Factors To initiate transcription, eukaryotic RNA polymerase requires the assistance of proteins called transcription factors General transcription factors are essential for the transcription of all protein-coding genes In eukaryotes, high levels of transcription of particular genes depend on control elements interacting with specific transcription factors

26. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Enhancers and Specific Transcription Factors Proximal control elements are located close to the promoter Distal control elements, groups of which are called enhancers, may be far away from a gene or even in an intron An activator is a protein that binds to an enhancer and stimulates transcription of a gene Animation: Initiation of Transcription Animation: Initiation of Transcription

27. LE 19-6 Distal control element Activators Enhancer DNA DNA-bending protein TATA box Promoter Gene General transcription factors Group of mediator proteins RNA polymerase II RNA polymerase II RNA synthesis Transcription Initiation complex

28. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Some transcription factors function as repressors, inhibiting expression of a particular gene Some activators and repressors act indirectly by influencing chromatin structure

29. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Combinatorial Control of Gene Activation A particular combination of control elements can activate transcription only when the appropriate activator proteins are present

30. LE 19-7 Control elements EnhancerPromoter Albumin gene Crystallin gene Available activators Available activators Albumin gene not expressed Albumin gene expressed Liver cell Lens cell Crystallin gene not expressed Crystallin gene expressed Liver cell nucleus Lens cell nucleus

31. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Coordinately Controlled Genes Unlike the genes of a prokaryotic operon, coordinately controlled eukaryotic genes each have a promoter and control elements The same regulatory sequences are common to all the genes of a group, enabling recognition by the same specific transcription factors

32. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Mechanisms of Post-Transcriptional Regulation Transcription alone does not account for gene expression More and more examples are being found of regulatory mechanisms that operate at various stages after transcription Such mechanisms allow a cell to fine-tune gene expression rapidly in response to environmental changes

33. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings RNA Processing In alternative RNA splicing, different mRNA molecules are produced from the same primary transcript, depending on which RNA segments are treated as exons and which as introns Animation: RNA Processing Animation: RNA Processing

34. LE 19-8 Primary RNA transcript DNA or Exons RNA splicing mRNA

35. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings mRNA Degradation The life span of mRNA molecules in the cytoplasm is a key to determining the protein synthesis The mRNA life span is determined in part by sequences in the leader and trailer regions

36. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings RNA interference by single-stranded microRNAs (miRNAs) can lead to degradation of an mRNA or block its translation The phenomenon of inhibition of gene expression by RNA molecules is called RNA interference (RNAi) Animation: Blocking Translation Animation: Blocking Translation Animation: mRNA Degradation Animation: mRNA Degradation

37. LE 19-9 Dicer Hydrogen bond Protein complex miRNA Target mRNA Degradation of mRNA OR Blockage of translation

38. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Initiation of Translation The initiation of translation of selected mRNAs can be blocked by regulatory proteins that bind to sequences or structures of the mRNA Alternatively, translation of all mRNAs in a cell may be regulated simultaneously

39. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Protein Processing and Degradation After translation, various types of protein processing, including cleavage and the addition of chemical groups, are subject to control Proteasomes are giant protein complexes that bind protein molecules and degrade them Animation: Protein Processing Animation: Protein Processing Animation: Protein Degradation Animation: Protein Degradation

40. LE 19-10 Protein to be degraded Ubiquitinated protein Proteasome Protein entering a proteasome Protein fragments (peptides) Proteasome and ubiquitin to be recycled Ubiquitin

41. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Concept 19.3: Cancer results from genetic changes that affect cell cycle control The gene regulation systems that go wrong during cancer are very same systems that play important roles in embryonic development Thus, research into the molecular basis of cancer has benefited from and informed many other fields of biology

42. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Types of Genes Associated with Cancer Genes that normally regulate cell growth and division during the cell cycle include: – Genes for growth factors – Their receptors – Intracellular molecules of signaling pathways Mutations altering any of these genes in somatic cells can lead to cancer

43. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Oncogenes and Proto-Oncogenes Oncogenes are cancer-causing genes Proto-oncogenes are normal cellular genes that code for proteins that stimulate normal cell growth and division A DNA change that makes a proto-oncogene excessively active converts it to an oncogene, which may promote excessive cell division and cancer

44. LE 19-11 Proto-oncogene DNA Translocation or transposition: gene moved to new locus, under new controls New promoter Gene amplification: multiple copies of the gene Point mutation within a control element Oncogene Point mutation within the gene Normal growth-stimulating protein in excess Normal growth-stimulating protein in excess Normal growth-stimulating protein in excess Hyperactive or degradation- resistant protein

45. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Tumor-Suppressor Genes Tumor-suppressor genes encode proteins that inhibit abnormal cell division Any decrease in the normal activity of a tumor- suppressor protein may contribute to cancer

46. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Interference with Normal Cell-Signaling Pathways Many proto-oncogenes and tumor suppressor genes encode components of growth-stimulating and growth-inhibiting pathways, respectively We will focus on products of two genes, the ras proto-oncogene and p53 tumor-suppressor gene

47. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The Ras protein, encoded by the ras gene, is a G protein that relays a signal from a growth factor receptor to a cascade of protein kinases Many ras oncogenes have a mutation that leads to a hyperactive Ras protein that issues signals on its own, resulting in excessive cell division

48. LE 19-12_1 Cell cycle-stimulating pathway Growth factor G protein Receptor MUTATION Protein kinases (phosphorylation cascade) NUCLEUS Hyperactive Ras protein (product of oncogene issues signals on its own. Transcription factor (activator) DNA Gene expression Protein that stimulates the cell cycle

49. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The p53 gene encodes a tumor-suppressor protein that is a specific transcription factor that promotes synthesis of cell cycle–inhibiting proteins Named for its 53,000-dalton protein product, the p53 gene is often called the “guardian angel of the genome” Mutations that knock out the p53 gene can lead to excessive cell growth and cancer

50. LE 19-12_2 Active form of p53 DNA DNA damage in genome UV light Protein kinases MUTATION Defective or missing transcription factor, such as p53, cannot activate transcription Protein kinases (phosphorylation cascade) Cell cycle-inhibiting pathway Cell cycle-stimulating pathway Protein that stimulates the cell cycle NUCLEUS DNA Gene expression Transcription factor (activator) Receptor G protein Growth factor MUTATION Hyperactive Ras protein (product of oncogene) issues signals on its own Protein that inhibits the cell cycle

51. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Increased cell division, possibly leading to cancer, can result if the cell cycle is overstimulated (as in Figure 19.12a) or not inhibited when it normally would be (as in Figure 19.12b)

52. LE 19-12_3 Protein overexpressed EFFECTS OF MUTATIONS Protein absent Cell cycle not inhibited Increased cell division Cell cycle overstimulate Effects of mutations Active form of p53 DNA DNA damage in genome UV light Protein kinases MUTATION Defective or missing transcription factor, such as p53, cannot activate transcription Protein kinases (phosphorylation cascade) Cell cycle-inhibiting pathway Cell cycle-stimulating pathway Protein that inhibits the cell cycle NUCLEUS DNA Gene expression Transcription factor (activator) Receptor G protein Growth factor MUTATION Hyperactive Ras protein (product of oncogene) issues signals on its own Protein that stimulates the cell cycle

53. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The Multistep Model of Cancer Development More than one somatic mutation is generally needed to produce a full-fledged cancer cell About a half dozen DNA changes must occur for a cell to become fully cancerous These changes usually include at least one active oncogene and mutation or loss of several tumor- suppressor genes Colorectal cancer, with 135,000 new cases and 60,000 deaths in the United States each year, illustrates a multistep cancer path

54. LE 19-13 Colon Colon wall Loss of tumor- suppressor gene APC (or other) Normal colon epithelial cells Small benign growth (polyp) Larger benign growth (adenoma) Activation of ras oncogene Loss of tumor- suppressor gene DCC Loss of tumor- suppressor gene p53 Additional mutations Malignant tumor (carcinoma)

55. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Certain viruses promote cancer by integration of viral DNA into a cell’s genome By this process, a retrovirus may donate an oncogene to the cell Viruses seem to play a role in about 15% of human cancer cases worldwide

56. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Inherited Predisposition to Cancer The fact that multiple genetic changes are required to produce a cancer cell helps explain the predispositions to cancer that run in some families Individuals who inherit a mutant oncogene or tumor-suppressor allele have an increased risk of developing certain types of cancer

57. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Concept 19.4: Eukaryotic genomes can have many noncoding DNA sequences in addition to genes The bulk of most eukaryotic genomes consists of noncoding DNA sequences, often described in the past as “junk DNA” However, much evidence is accumulating that noncoding DNA plays important roles in the cell

58. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The Relationship Between Genomic Composition and Organismal Complexity Compared with prokaryotic genomes, the genomes of eukaryotes: – Generally are larger – Have longer genes – Contain much more noncoding DNA

59. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The sequencing of the human genome reveals what makes up most of the 98.5% of the genome that does not code for proteins, rRNAs, or tRNAs Most intergenic DNA is repetitive DNA, present in multiple copies in the genome About three-fourths of repetitive DNA is made up of transposable elements and sequences related to them

60. LE 19-14 Exons (regions of genes coding for protein, rRNA, or tRNA) (1.5%) Alu elements (10%) Simple sequence DNA (3%) Large-segment duplications (5–6%) Unique noncoding DNA (15%) Introns and regulatory sequences (24%) Repetitive DNA that includes transposable elements and related sequences (44%) Repetitive DNA unrelated to transposable elements (about 15%)

61. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Transposable Elements and Related Sequences The first evidence for wandering DNA segments came from geneticist Barbara McClintock’s breeding experiments with Indian corn McClintock identified changes in the color of corn kernels that made sense only by postulating that some genetic elements move from other genome locations into the genes for kernel color

62. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings

63. Movement of Transposons and Retrotransposons Eukaryotic transposable elements are of two types: – Transposons, which move within a genome by means of a DNA intermediate – Retrotransposons, which move by means of an RNA intermediate

64. LE 19-16 DNA of genome Transposon is copied Mobile transposon Transposon Insertion New copy of transposon Transposon movement (“copy-and-paste” mechanism) Retrotransposon movement DNA of genome Insertion RNA Reverse transcriptase Retrotransposon New copy of retrotransposon

65. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Sequences Related to Transposable Elements Multiple copies of transposable elements and related sequences are scattered throughout the eukaryotic genome In primates, a large portion of transposable element–related DNA consists of a family of similar sequences called Alu elements

66. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Other Repetitive DNA, Including Simple Sequence DNA Simple sequence DNA contains many copies of repeated short sequences Simple sequence DNA is common in centromeres and telomeres, where it probably plays structural roles in the chromosome

67. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Genes and Multigene Families Most eukaryotic genes are present in one copy per haploid set of chromosomes The rest of the genome occurs in multigene families, collections of identical or very similar genes

68. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Some multigene families consist of identical DNA sequences, usually clustered tandemly, such as those that code for RNA products

69. LE 19-17a DNA Non-transcribed spacer RNA transcripts Transcription unit DNA 18S 5.8S 28S rRNA 18S 5.8S 28S Part of the ribosomal RNA gene family

70. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The classic examples of multigene families of nonidentical genes are two related families of genes that encode globins Globin gene family clusters also include pseudogenes, nonfunctional nucleotide sequences that are similar to the functional genes

71. LE 19-17b Heme Hemoglobin  -Globin  -Globin  -Globin gene family  -Globin gene family Chromosome 11 Chromosome 16    11 11 22      AA Embryo Fetus Adult Fetus and adult The human  -globin and  -globin gene families

72. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Concept 19.5: Duplications, rearrangements, and mutations of DNA contribute to genome evolution The basis of change at the genomic level is mutation, underlying much of genome evolution The earliest forms of life likely had a minimal number of genes, including only those necessary for survival and reproduction The size of genomes has increased over evolutionary time, with the extra genetic material providing raw material for gene diversification

73. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Duplication of Chromosome Sets Accidents in meiosis can lead to one or more extra sets of chromosomes, a condition known as polyploidy The genes in one or more of the extra sets can diverge by accumulating mutations; these variations may persist if the organism carrying them survives and reproduces

74. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Duplication and Divergence of DNA Segments Unequal crossing over during prophase I of meiosis can result in one chromosome with a deletion and another with a duplication of a particular region

75. LE 19-18 Nonsister chromatids Transposable element Gene Crossover Incorrect pairing of two homologues during meiosis and

76. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Evolution of Genes with Related Functions: The Human Globin Genes The genes encoding the various globin proteins evolved from one common ancestral globin gene, which duplicated and diverged After the duplication events, differences between the genes in the globin family arose from mutations that accumulated in the gene copies over many generations

77. LE 19-19 Duplication of ancestral gene Mutation in both copies Transposition to different chromosomes Further duplications and mutations Ancestral globin gene    11 11 22     AA  -Globin gene family on chromosome 16   -Globin gene family on chromosome 11          Evolutionary time

78. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Subsequent duplications of these genes and random mutations gave rise to the present globin genes, which code for oxygen-binding proteins

79. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The similarity in the amino acid sequences of the various globin proteins supports this model of gene duplication and mutation

80. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings

81. Evolution of Genes with Novel Functions The copies of some duplicated genes have diverged so much in evolution that the functions of their encoded proteins are now very different

82. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Rearrangements of Parts of Genes: Exon Duplication and Exon Shuffling An exon can be duplicated on one chromosome and deleted from the homologous chromosome In exon shuffling, errors in meiotic recombination lead to some mixing and matching of exons, either within a gene or between two nonallelic genes

83. LE 19-20 Epidermal growth factor gene with multiple EGF exons (green) EGF FF F F Fibronectin gene with multiple “finger” exons (orange) K K KEGF F Plasminogen gene with a “kringle” exon (blue) Portions of ancestral genes TPA gene as it exists today Exon shuffling Exon shuffling Exon duplication

84. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings How Transposable Elements Contribute to Genome Evolution Movement of transposable elements or recombination between copies of the same element may generate beneficial new sequence combinations Some mechanisms can alter functions of genes or their patterns of expression and regulation