1. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E The Stability of the Genome Duplication, Deletion, Transposition

2. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Duplication of DNA sequences Could result from unequal crossover –chromosomes are not perfectly aligned during meiosis –One gets a duplication; the other gets a deletion –Process continues, to build tandem repeats After duplication, base substitutions (variation) Multiple gene family members from one ancestral copy - divergence required

3. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Figure 10.24

4. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Evolution of globin genes Hemoglobin is tetramer: 4 globin chains Globin genes from different vertebrates have common features –All globins have 3 exons & 2 introns Globin-like proteins are different –leghemoglobin from plants; muscle myoglobin –4 exons & 3 introns –ancestral? (4 exons to 3 exons)

5. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Evolution of globin genes fusion of 2 exons (~800 million years ago) Some primitive fish have only one globin gene perhaps fish diverged before the first duplication First duplication formed  &  (~500 million yrs ago)  &  separated transposition/translocation today's organization humans –  -globin genes clustered on chromosome 16, –  -globin genes clustered on chromosome 11

6. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Figure 10.26

7. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Evolution of globin genes Hemoglobin made of 2 pairs of chains –One pair always  family; the other always  -family –Combinations differ with stage of development –embryonic, fetal, adult Clusters also contain pseudogenes –have mutations that stop function –evolutionary relics –widespread in many gene families –function unknown (junk?)

8. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Evolution of globin genes Parts of globin genes evolve at very different rates –DNA of coding regions is highly conserved –Noncoding regions are much more variable –Coding area changes are most frequently one base substitutions –Noncoding - often include additions & deletions as well

9. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Evolution of gene families Evolving Sequence and Function –Growth hormone & prolactin are pituitary hormones Related amino acid sequences Evoke completely different responses from target cells –Number of changes only ROUGHLY corresponds to time –Redundant codons allow “silent” substitutions

10. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Transposons Described by Barbara McClintock (Cold Spring Harbor, late 1940s) –Nobel Prize – 1983 –Initially ignored: papers (and subject) complex –Studied patterns & markings in leaf & kernel coloration maize –Noticed instability in markers in both germ line and soma –Concluded movement of genetic elements affecting gene expression –Defined terms: transposition and transposable elements

11. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Transposons Eventually (late 1960s) verified transposons in bacteria –Encode transposase enzyme –Excision from donor site & insertion at target site –bind to specific sequences (IR’s) ends of transposon –complex binds to target DNA –catalyzes integration –Integration creates small duplication in target (DR’s) –“Footprints” left behind when transposon excises

12. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Figure 10.28

13. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Figure 10.29

14. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Transposons Eukaryotic genomes contain large numbers of transposable elements –~40% of DNA in human cell nucleus from transposable elements –Vast majority are crippled by mutation or suppressed –they insert almost randomly within target DNA sometimes cause mutations in genes ~1 in 500 human mutations caused by transposons

15. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Mechanisms of transposition are complex Some are excised from donor site and inserted into target –Example: mariner family of transposons –Found throughout plant & animal kingdoms Some are replicated leaving donor site unchanged (bacteria)

16. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Mechanisms of transposition are complex Often, transposition in eukaryotes involves RNA intermediate –DNA transcribed, producing an RNA –Converted to DNA by reverse transcriptase –dsDNA copy is integrated into target DNA site –Retrotransposons sometimes have gene for RVTase –Retroviruses, like the AIDS virus related to retrotransposons Acquired envelope protein genes Or, loss of envolope gene could make transposon

17. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Figure 10.30

18. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Mobile elements and evolution Most moderately repeated sequences of genome are –interspersed & arise by transposition –2 most common families: Alu & L1 families –both transpose by means of RNA intermediates

19. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Mobile elements and evolution L1 –~500,000 copies; ~15% of total nuclear DNA –Vast majority of these are incomplete & defective –Likely present in earliest eukaryotic cells (found everywhere) –Full length, human L1 sequence at least 6000 bp in length) encodes unique protein with 2 catalytic activities Endonuclease activity cleaves target DNA Reverse transcriptase activity makes DNA from RNA

20. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Mobile elements and evolution Alu –roughly 1 million copies –Family of short sequences ~300 bp in length –Related to 7S RNA present in signal recognition particles found with membrane-bound ribosomes –Alu origin: 7S RNA & reverse transcription? –Amplification aided by L1 endonuclease? –Seen only in higher primates –First appearing ~60 million years ago

21. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Mobile elements and evolution Proposed functions of transposable elements –No function - a genetic parasite –Fuels evolution

22. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Support for evolutionary role Found as essential parts of regulatory regions –Human parathyroid hormone gene expression controlled, in part, by relics of transposable element Can clearly reorder genomes –Domain shuffling using introns as breakpoints Transposable elements become useful genes –Telomerase derived from reverse transcriptase? –Enzymes involved in antibody gene rearrangement are derived from transposase encoded by ancient DNA transposon

23. Copyright, ©, 2002, John Wiley & Sons, Inc.,Karp/CELL & MOLECULAR BIOLOGY 3E Support for evolutionary role Transposable (P) element of Drosophila melanogaster –T. H. Morgan's flies from 1900s devoid of the P element –All members of the wild species have it today –Spread rapidly (80 years) Transmission likely mediated by parasites