Eukaryotic Genomes: Organization, Regulation, and Evolution

1. Eukaryotic Genomes: Organization, Regulation, and Evolution Chapter 19

2. Organization

3. Eukaryotic Cells • Genome is much larger than prokaryotic cells • Multicellular beings have very strict cell specialization • Controlled by regulation • DNA associates with proteins in a complex called chromatin in order to order the DNA and regulate it

4. Chromatin Structure • Based on successive levels of DNA packing • Eukaryotic DNA is precisely combined with a large amount of protein • Chromatin changes during the course of the cell cycle • Eukaryotic chromosomes contain an enormous amount of DNA relative to their condensed length • Helps to regulate gene expression, condense and release and form chromosomes

5. Nucleosomes • Proteins called histones • Are responsible for the first level of DNA packing in chromatin • Bind tightly to DNA • 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

6. 2 nm DNA double helix Histone tails His- tones 10 nm Histone H1 Linker DNA (“string”) Nucleosome (“bead”) Nucleosomes • Each “bead” is a nucleosome • The basic unit of DNA packing

7. 30 nm Nucleosome Higher Levels of DNA Packing • Interactions between the histone tails of the nucleosomes • Causes the nucleosomes to coil around each other • Forms a 30-nm chromatin fiber

8. Protein scaffold Loops Scaffold 300 nm (c) Looped domains (300-nm fiber) DNA Packing • The 30-nm fiber, in turn • Forms looped domains, making up a 300-nm fiber (euchromatin)

9. 700 nm 1,400 nm (d) Metaphase chromosome DNA Packing • In a mitotic chromosome • The looped domains themselves coil and fold forming the characteristic metaphase chromosome (heterochromatin)

10. Regulation

11. Gene Regulation • All organisms • Must regulate which genes are expressed at any given time • Each cell of a multi-cellular eukaryote • Expresses only a fraction of its genes • In each type of differentiated cell • A unique subset of genes is expressed • In interphase cells • Most chromatin is in the highly extended form called euchromatin • Genes within highly packed heterochromatin • Are usually not 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 Gene Regulation • Many key stages of gene expression • Can be regulated in eukaryotic cells • Each gene is regulated in its own particular way or ways

13. Chromatin changes Transcription RNA processing mRNA degradation Translation Protein processing and degradation Histone tails DNA double helix Amino acids available for chemical modification (a) Histone tails protrude outward from a nucleosome Unacetylated histones Acetylated histones Regulation of Chromatin Structure • Histone Modifications- • Chemical modification of histone tails • Affect the configuration of chromatin and thus gene expression • Histone acetylation • Seems to loosen chromatin structure and thereby enhance transcription • DNA Methylation- reduced transcription in some species

14. Regulation of Transcription Initiation • After chromatin modification, the next step in regulating gene expression is to stop initiation • Enzymes modify the chromatin to make the DNA more or less available to bind o transcription initiation

15. Organization of a Typical Gene

16. Poly-A signal sequence Termination region Proximal control elements Enhancer (distal control elements) 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 3 UTR (untranslated region) 5 Cap 5 UTR (untranslated region) • Associated with most eukaryotic genes are multiple control elements • Segments of noncoding DNA that help regulate transcription by binding certain proteins

17. Transcription Factors • To initiate transcription • Eukaryotic RNA polymerase requires the assistance of proteins called transcription factors • Only when the complete initiation complex is assembled can the polymerase produce the complimentary strand • Some specific transcription factors function as repressors • To inhibit expression of a particular gene

18. Chromatin changes Transcription RNA processing mRNA degradation Translation Protein processing and degradation Exons DNA Primary RNA transcript RNA splicing or mRNA Post- Transcriptional Regulation • 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

19. Post-Transcriptional Regulation • mRNA Degradation • The life span of mRNA molecules in the cytoplasm • Is an important factor in determining the protein synthesis in a cell • Determines how long the mRNA will last in the cytoplasm and how many times the mRNA will be read • Is determined in part by sequences in the leader and trailer regions

20. Initiation of Translation • The initiation of translation of selected mRNAs • Can be blocked by regulatory proteins that bind to specific sequences or structures of the mRNA • Prevent ribosome from attaching to the mRNA • Translation of all the mRNAs in a cell may be regulated simultaneously • Plays a role in the translation of mRNA’s stored in egg cells

21. Post- Translational Regulation • 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 • Mutations in proteasomes can lead to cancer

22. 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 Post- Translational Regulation

23. Cancer

24. Cancer • Cancer results from genetic changes that affect cell cycle control • The gene regulation systems that go wrong during cancer play important roles in embryonic development • The genes that normally regulate cell growth and division during the cell cycle • Include genes for growth factors, their receptors, and the intracellular molecules of signaling pathways

25. Cancer • Oncogenes • Are cancer-causing genes • Proto-oncogenes • Are normal cellular genes that code for proteins that stimulate normal cell growth and division • An oncogene arises from a genetic change in a proto-oncogene that either increases the amount of protein produced or in the activity of the protein molecule

26. Proto-oncogene DNA Translocation or transposition: gene moved to new locus, under new controls Point mutation within a control element 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 Mutations that change proto-oncogenes into oncogenes

27. Tumor Suppression • Tumor-suppressor genes • Encode proteins that inhibit abnormal cell division • Mutation in these may contribute to the onset of cancer • Code for proteins that: • Repair damaged DNA • Control adhesion of cells to each other • Inhibit the cell cycle

28. Colon 1 Loss of tumor- suppressor gene APC (or other) 4 Loss of tumor-suppressor gene p53 2 Activation of ras oncogene Colon wall 3 Loss of tumor- suppressor gene DCC 5 Additional mutations Normal colon epithelial cells Larger benign growth (adenoma) Small benign growth (polyp) Malignant tumor (carcinoma) Cancer Development • Normal cells are converted to cancer cells • By the accumulation of multiple mutations affecting proto-oncogenes and tumor-suppressor genes • A multistep model for the development of colorectal cancer

29. Other Promoters of Cancer • Certain viruses • Promote cancer by integration of viral DNA into a cell’s genome • Individuals who inherit a mutant oncogene or tumor-suppressor allele • Have an increased risk of developing certain types of cancer