Eukaryotic Chromatin Structure and Gene Expression Regulation

1. Chapter 19 • Eukaryotic Genomes

2. Figure 19.1 • Eukaryotes DNA-protein complex,  chromatin • More complex structural levels than prokaryotes

3. Both prokaryotes and eukaryotes • Must alter patterns of gene expression in response to changes in environment

4. Chromatin structure based on successive levels of DNA packing • Eukaryotic DNA • Combined w/ protein • Eukaryotic chromosomes • Contain an enormous amount of DNA relative to their condensed length

5. 2 nm DNA double helix Histone tails His- tones 10 nm Histone H1 Nucleosome (“bead”) Linker DNA (“string”) (a) Nucleosomes (10-nm fiber) Nucleosomes, or “Beads on a String” • Basic packing unit • DNA wrapped around histone protein Figure 19.2 a

6. 30 nm Nucleosome (b) 30-nm fiber • Next level of packing • Forms 30-nm chromatin fiber Figure 19.2 b

7. Protein scaffold Loops Scaffold 300 nm (c) Looped domains (300-nm fiber) • The 30-nm fiber, in turn • Forms looped domains, making up a 300-nm fiber Figure 19.2 c

8. 700 nm 1,400 nm (d) Metaphase chromosome • Mitotic chromosome • Looped domains coil and fold forming the metaphase chromosome Figure 19.2 d

9. Interphase cells • Most chromatin is highly extended (euchromatin)

10. Gene expression can be regulated at any stage, but the key step is transcription • All organisms • regulate which genes are expressed at any given time • During development of a multicellular organism cell specialization in form and function (cell differentiation)

11. Each cell of a multicellular eukaryote • Expresses only a fraction of its genes • In each type of differentiated cell • Unique subset of genes is expressed

12. Signal NUCLEUS Chromatin Chromatin modification: DNA unpacking involving histone acetylation and DNA demethlation DNA Gene available for transcription Gene Transcription Exon RNA Primary transcript Intron RNA processing Tail mRNA in nucleus Cap Transport to cytoplasm CYTOPLASM mRNA in cytoplasm Degradation of mRNA Translation Polypetide Cleavage Chemical modification Transport to cellular destination Active protein Degradation of protein Degraded protein Figure 19.3 • Many key stages of gene expression (regulation) In eukaryotic cells

13. Genes within highly packed heterochromatin usually not expressed

14. Acetylated histones Unacetylated histones (b) Acetylation of histone tails promotes loose chromatin structure that permits transcription Figure 19.4 b • Histone acetylation • Loosens chromatin structure enhances transcription

15. Addition of methyl groups to DNA bases • Associated w/ reduced transcription

16. Proximal control elements Enhancer (distal control elements) Poly-A signal sequence Termination region Exon Intron Intron Exon Exon DNA Downstream Upstream Promoter Transcription Poly-A signal Exon Exon Intron Intron Exon Cleared 3 end of primary transport Primary RNA transcript (pre-mRNA) 5 Chromatin changes RNA processing: Cap and tail added; introns excised and exons spliced together Transcription Intron RNA RNA processing Coding segment mRNA degradation Translation mRNA P G Protein processing and degradation P P Start codon Poly-A tail Stop codon 5 Cap 3 UTR (untranslated region) 5 UTR (untranslated region) • Multiple control elements • Noncoding DNA that regulatetranscription Figure 19.5

17. Chromatin changes Transcription RNA processing mRNA degradation Translation Protein processing and degradation Exons DNA Primary RNA transcript RNA splicing or mRNA Alternate RNA Processing • Different mRNA molecules produced from same primary transcript, depending on which RNA segments are treated as exons and which as introns Figure 19.8

18. mRNA Degradation • Life span of mRNA molecules in the cytoplasm • Important in protein synthesis

19. The micro- RNA (miRNA) precursor folds back on itself, held together by hydrogen bonds. The bound miRNA can base-pair with any target mRNA that contains the complementary sequence. One strand of each short double- stranded RNA is degraded; the other strand (miRNA) then associates with a complex of proteins. 3 The miRNA-protein complex prevents gene expression either by degrading the target mRNA or by blocking its translation. An enzyme called Dicer moves along the double- stranded RNA, cutting it into shorter segments. 5 4 2 1 2 Chromatin changes Transcription RNA processing mRNA degradation Translation Protein complex Protein processing and degradation Dicer Degradation of mRNA OR miRNA Target mRNA Blockage of translation Hydrogen bond • RNA interference(RNAi) by single-stranded microRNAs (miRNAs) • degradation of mRNA or block its translation 5 Figure 19.9

20. Enzymatic components of the proteasome cut the protein into small peptides, which can be further degraded by other enzymes in the cytosol. 3 The ubiquitin-tagged protein is recognized by a proteasome, which unfolds the protein and sequesters it within a central cavity. 2 1 Multiple ubiquitin mol- ecules are attached to a protein by enzymes in the cytosol. Chromatin changes Transcription RNA processing Proteasome and ubiquitin to be recycled Ubiquitin Translation mRNA degradation Proteasome Protein processing and degradation Protein fragments (peptides) Protein to be degraded Ubiquinated protein Protein entering a proteasome • Proteasomes • Giant protein complexes that degrade molecules Figure 19.10

21. Cancer results from genetic changes that affect cell cycle control • The gene regulation systems that go wrong during cancer are same systems found in embryonic development

22. Oncogenes • Cancer-causing genes • Proto-oncogenes • Normal genes that code for proteins that stimulate normal cell growth and division

23. Proto-oncogene DNA Point mutation within a control element Translocation or transposition: gene moved to new locus, under new controls Point mutation within the gene Gene amplification: multiple copies of the gene Oncogene Oncogene New promoter Normal growth-stimulating protein in excess Hyperactive or degradation- resistant protein Normal growth-stimulating protein in excess Normal growth-stimulating protein in excess • DNA changeproto-oncogene excessively active oncogeneexcessive cell division cancer Figure 19.11

24. Tumor-suppressor genes • Code f/ proteins that inhibit abnormal cell division

25. (b) Cell cycle–inhibiting pathway. In this pathway, DNA damage is an intracellular signal that is passed via protein kinases and leads to activation of p53. Activated p53 promotes transcription of the gene for a protein that inhibits the cell cycle. The resulting suppression of cell division ensures that the damaged DNA is not replicated. Mutations causing deficiencies in any pathway component can contribute to the development of cancer. 1 Protein kinases 2 2 MUTATION 3 Defective or missing transcription factor, such as p53, cannot activate transcription UV light Active form of p53 3 DNA damage in genome 1 DNA Protein that inhibits the cell cycle • p53gene encodes a tumor-suppressor protein • (cell cycle–inhibiting proteins) • ‘Guardian angel’ of the genome Figure 19.12b

26. Mutations that knock out the p53 gene  excessive cell growth and cancer (c) Effects of mutations. Increased cell division, possibly leading to cancer, can result if the cell cycle is overstimulated, as in (a), or not inhibited when it normally would be, as in (b). EFFECTS OF MUTATIONS Protein overexpressed Protein absent Cell cycle not inhibited Cell cycle overstimulated Increased cell division Figure 19.12c

27. Normal cells are converted to cancer cells • By the accumulation of multiple mutations affecting proto-oncogenes and tumor-suppressor genes

28. Colon 1Loss of tumor- suppressor gene APC (or other) 4Loss of tumor-suppressor gene p53 2Activation of ras oncogene Colon wall 3Loss of tumor- suppressor gene DCC 5 Additional mutations Normal colon epithelial cells Larger benign growth (adenoma) Small benign growth (polyp) Malignant tumor (carcinoma) • A multistep model for the development of colorectal cancer Figure 19.13

29. Certain viruses • Promote cancer by integration of viral DNA into a cell’s genome

30. Inheritance of a mutant oncogene  increased risk of developing cancer

31. Noncoding DNA sequences • Bulk of eukaryotic genomes • In the past called “junk DNA” • Evidence is accumulating • noncoding DNA plays important roles in the cell

32. Genomes of eukaryotes (v. prokaryotic) • Larger • Longer genes • Much greater amount of noncoding DNA

33. Exons (regions of genes coding for protein, rRNA, tRNA) (1.5%) Repetitive DNA that includes transposable elements and related sequences (44%) Introns and regulatory sequences (24%) Unique noncoding DNA (15%) Repetitive DNA unrelated to transposable elements (about 15%) Alu elements (10%) Simple sequence DNA (3%) Large-segment duplications (5-6%) • Most of the 98.5% that does not code for proteins rRNAs, or tRNAs Figure 19.14

34. Transposable Elements and Related Sequences • Wandering DNA segments • Barbara McClintock’s breeding experiments with Indian corn Figure 19.15

35. New copy of transposon Transposon DNA of genome Transposon is copied Insertion Mobile transposon (a) Transposon movement (“copy-and-paste” mechanism) New copy of retrotransposon Retrotransposon DNA of genome RNA Insertion Reverse transcriptase (b) Retrotransposon movement Movement of Transposons and Retrotransposons • Transposons: move by means of a DNA intermediate • Retrotransposons: move by means of an RNA intermediate Figure 19.16a, b

36. Multiple copies of transposable elements • scattered throughout genome • In humans and other primates • are called Alu elements

37. Other Repetitive DNA • Simple sequence DNA • Copies of tandemly repeated short sequences • Common in centromeres and telomeres (structural roles in chromosome)

38. Genes and Multigene Families • Most eukaryotic genes • 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

39. Heme Hemoglobin -Globin -Globin gene family -Globin gene family Chromosome 16 Chromosome 11  A    G    2 1   1 2 Fetus and adult Embryo Fetus Adult Embryo • e.g. related families of genes that encode globins -Globin Figure 19.17b The human -globin and -globin gene families

40. Duplications, rearrangements, and mutations of DNA contribute to genome evolution • The basis of change at the genomic level is mutation which underlies much of genome evolution

41. Duplication of Chromosome Sets • Accidents in cell division •  extra copies of all or part of a genome, which may then diverge if one set accumulates sequence changes

42. Transposable element Gene Nonsister chromatids Crossover Incorrect pairing of two homologues during meiosis and • Unequal crossing over •  one chromosome with a deletion and another with a duplication Figure 19.18

43. Ancestral globin gene Duplication of ancestral gene Mutation in both copies   Transposition to different chromosomes Evolutionary time   Further duplications and mutations               1 2 A G 2 1 -Globin gene family on chromosome 16  -Globin gene family on chromosome 11 • Genes for various globin proteins • Evolved from one common ancestral globin gene, which duplicated and diverged Figure 19.19

44. Similarity in the amino acid sequences of the various globin proteins • Supports this model of gene duplication and mutation Table 19.1

45. The copies of some duplicated genes • Have diverged so much during evolutionary time that the functions of their proteins are now substantially different

46. Rearrangements of Parts of Genes: Exon Duplication and Exon Shuffling

47. EGF EGF EGF EGF Epidermal growth factor gene with multiple EGF exons (green) Exon shuffling Exon duplication F F F F Fibronectin gene with multiple “finger” exons (orange) F EGF K K K Exon shuffling Plasminogen gene with a “kfingle” exon (blue) Portions of ancestral genes TPA gene as it exists today • Exon shuffling • Mixing and matching of different exons either within a gene or between two nonallelic genes Figure 19.20

48. How Transposable Elements Contribute to Genome Evolution • Movement of transposable elements • Can generates new sequence combinations that are beneficial to the organism • Some mechanisms • Alter functions of genes or their patterns of expression and regulation