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Basic considerations in formulating a21st Century theory of the genome and
genome evolutionJames A. Shapiro
University of [email protected], http:shapiro.bsd.uchicago.edu
• 40 years of molecular deconstruction ==> no units, only systems
• Contact with physicists ==> emphasis on basic principles
• Experience with transposable elements ==> genetic change notstochastic
What are theBasic Functions
of Cells?
Fraser et al. 1997. Genomic sequence of a Lyme disease spirochaete, Borrelia burgdorferi.Nature 390: 580-586.
• Transport• Metabolism• Growth (includes RNA,protein synthesis, DNAreplication, membranebiogenesis, etc.)• Division (includesdistribution of essential cellstructures & molecules)• Movement (includestropisms and motility)• Communication with othercells (multicellularity,symbiosis & pathogenesis)• Cognition, signaltransduction & informationprocessing (all the above)
SignalTransduction
in YeastMating
Mary J. Cismowski et al. Genetic screens in yeast to identify mammalian nonreceptor modulators ofG-protein signaling. Nature Biotechnology 17, 878 - 883 (1999).
Pheromone ResponseFrank van Drogen et al. MAP kinasedynamics in response to pheromones inbudding yeast. Nature Cell Biology 3,1051 - 1059 (2001).
What are basic properties ofbiological macromolecules?
• Stereospecificity• Structural flexibility• Multivalency (multiple
ligands)• Allostery• Reversible covalent
modifications (-CH3, ~PO3)• Weak interactions• Cooperativity• Modularity• Combinatorics
What functions does the genome fulfill?
• Expression (Protein & RNA coding) – transcription,chromatin
• Maintenance – replication & repair• Transmission to progeny cells – chromosome
movement/segregation at division• Restructuring – natural genetic engineering
(genome as RW storage organelle)
• NB – all functions involve nucleoprotein complexes,not naked DNA
What is the functional organization ofthe genome?
• Formatting by generic signals for compaction, attachment,transcription, replication, transmission, repair
• Networking by repeat sequences• Higher order “indexing” by chromatin domains, spatial
organization of chromosomes
DNA formatting to execute a cognitive algorithm - E. coli discriminates glucose and lactose
1. The algorithm: IF lactose present AND glucose not present AND cell can synthesizeactive LacZ (beta-galactosidase) and LacY (lactose permease), THEN transcribe lacZYAfrom lacP
2. The formatted DNA:
DNA formatting for chromosome transmissionto daughter cells - centromeres
B A Sullivan, M DBlower & G HKarpenDETERMININGCENTROMEREIDENTITY:CYCLICALSTORIES ANDFORKINGPATHS NatureReviews Genetics2; 584-596 (2001)
What are the sources of novelty inevolution?
•New protein functions - domain shuffling andaccretion•New adaptive systems (Hox complexes,
olfactory sensing, early embryonic functions) –duplication, reverse transcription, multipletranspositions•New species (novel cereals, Muntjac,
Drosophila) – hybridization, changes in ploidy,chromosome fusions, inversions•New kinds of cells (e.g. Gram-negative bacteria,
eukaryotes, plants) - symbiosis
Protein evolution by domain accretion
International Human Genome Sequencing Consortium. Initial sequencing and analysis of the humangenome. Nature 409, 860 - 921 (2001)
Most pairs of similar OR gene clusters (labeled in black) fall into established syntenic chromosomalregions (http://www.ncbi.nlm.nih.gov/Homology/index.html), but some (labeled in red) do not.Young, J. M. et al. Different evolutionary processes shaped the mouse and human olfactory receptor genefamilies. Hum. Mol. Genet. 11, 535-546 (2002)
Protein Amplification: Olfactory Receptors
Segmental Duplications in Arabidopsis
Segmentally duplicated regions in the Arabidopsis genome. Individual chromosomes are depicted as horizontal grey bars (withchromosome 1 at the top), centromeres are marked black. Coloured bands connect corresponding duplicated segments. Similarity betweenthe rDNA repeats are excluded. Duplicated segments in reversed orientation are connected with twisted coloured bands. The scale is inmegabases. Analysis of the genome sequence of the flowering plant Arabidopsis thaliana. Nature 408, 796 - 815 (2000).
Evolution of imprinted heterochromatin by insertions
Lippman Z, Gendrel AV, Black M, Vaughn MW, DedhiaN, McCombie WR, Lavine K, Mittal V, May B,Kasschau KD, Carrington JC, Doerge RW, Colot V,Martienssen R. Role of transposable elements inheterochromatin and epigenetic control. Nature. 2004 Jul22;430(6998):471-6.
How does primary genome structure change? -Natural Genetic Engineering: the cellular
toolbox for genome restructuring
• Homologous recombination• Non-homologous end-joining• Site-specific recombination• DNA transposons (large-scale rearrangements)• Retrotransposons and reverse transcription (small-
scale rearrangements)• Homing and retrohoming introns and inteins• Mutator polymerases
The MammalianImmune System:
An EvolvedRapid Evolution
System
D. C. van Gent, J. H. Hoeijmakers, R. Kanaar,Chromosomal Stability And The DNA Double-Stranded Break Connection 2, 196 (2001)
Somatic hypermutation &class switch recombination- transcription directed
Tasuko Honjo, Kazuo Kinoshita, and Masamichi Muramatsu. 2001. Molecular Mechanism of Class Switch Recombination: Linkage withSomatic Hypermutation. Annu. Rev. Immunol.;
Temporal & metabolic regulation of naturalgenetic engineering
0 10 200
100
200
MCS2 (2 subclones)
MCS1366 (4 subclones)
Days/32
Tota
l fus
ion
colo
nies
lacZaraB
Derepression(42C, starvation)ClpPX, Lon RpoS
MuA, HU, IHF
Strand transfer
Replication (exponential growth)
ClpX
DNA processing(RpoS-, Crp-dependent functions?)
lacZaraB
araB-lacZ fusion
STC = strand transfer complex
araB lacZ
U118
U118
lacZaraBCDC/Target complex
U118araB lacZ
Adjacent inversion (precludes fusion)
U118
Transposasome formation
MuB for replication(Crp-dependent starvation-induced functions inhibitand/or replace MuB?)
ClpX
Shapiro, J.A. 1997b. Genome organization, natural genetic engineering, and adaptive mutation. Trends in Genetics 13, 98-104
Leaf wounding and retrotransposon transcription
http://www-biocel.versailles.inra.fr/Anglais/03Transposon.html
The expression of the tobacco Tnt1 retrotransposon is induced bywounding : the expression of the LTR-GUS construct is detected by a bluestaining surrounding injury points in transgenic tomato (A), tobacco (B)and Arabidopsis (C) plants.
M.-A. Grandbastien et al. Stress activation and genomic impact of Tnt1 retrotransposons in Solanaceae.Cytogenetic and Genome Research 2005;110:229-241
Targeting of natural genetic engineering
Shapiro, JA. 2005. A 21st Century View Of Evolution: Genome System Architecture, Repetitive DNA, And Natural Genetic Engineering. Gene345: 91-100
Known molecular mechanisms:
• Sequence recognition by proteins (yeast mating-type switching,ribosomal LINE elements, homing introns, VDJ joining);• Protein-protein interaction wth transcription factors or chromatinproteins (Ty retrotransposon targeting);• Sequence recognition by RNA (reverse splicing of group IIretrohoming introns);• Transcriptional activation of target DNA (somatic hypermutation;class-switch recombination).
Unknown mechanisms:
• Telomere targeting of certain LINE elements in insects;• HIV & MLV targeting upstream of transcribed regions;• P factor homing directed by transcription, chromatin signals.
Yeast Ty5 targeting
S. Sandmeyer. Integration by design. PNAS, May 13, 2003; 100(10): 5586 - 5588.
Advantages of non-random searches ofgenome space at evolutionary crises
• Genome changes occur under stress or other conditions, when they aremost likely to prove beneficial;
• Multiple related changes can occur when a particular natural geneticengineering system is activated;
• Rearrangement of proven genomic components increases the chancethat novel combinations will be functional;
• Targeting can increase the probability of functional integration andreduce the risk of system damage;
• Rearrangements followed by localized changes provide opportunitiesfor fine tuning once novel function has been achieved.
What general principles operate in genomefunction and genome reorganization?
• All genome functions are interactive (no Cartesiandualism, genome always in communication with restof cell);
• Every genome component operates as part of acomplex information-processing system (no “onegene-one trait” correlation);
• Genome systems are organized and integrated intocell networks by repetitive DNA;
• Genome change is a regulated biological function;• Natural genetic engineering processes are subject to
biological feedback at multiple levels.