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Sec$on 9 Evolu$onary Rela$onships Sec$on 9 Learning Goals •  Explain why the ribosomal 16S gene is a good “marker” for molecular phylogene$c comparisons. •  Be able to interpret a phylogene$c tree. •  Explain the origins of mitochondria and chloroplasts and the evidence that supports this theory. Evolu$on is the thread that connects all biology. What is useful is retained. What is not is subject to elimina$on While DNA muta$ons can cause harm, without them popula$ons would not evolve. Taxonomy places organisms into groups with shared features Phylogene$c trees can be constructed to represent a hypothesis about the evolu$onary rela$onships between taxonomic groups •  Branches link groups according to their presumed ancestry •  Groups for which less distance separates them share more features than groups that are separated by greater distance All life can be divided into three domains •  Certain features are shared across all life. For example: –  Core set of macromolecules –  DNA as the gene$c code –  ATP as an energy source –  Transfer of electrons to acquire energy •  Yet, random muta$on leads to differences in individuals; selec$ve pressure leads to reten$on of those differences within a popula$on. For example: –  Structural difference of conserved features (e.g. ribosome, RNA polymerase) –  Gain of features that are advantageous (or loss of those that are costly) (e.g. wings, gills, pep$doglycan, nucleus,tail, etc.) Classifica$on systems allow us to group organisms These grouping allow us to make hypotheses regarding the extent of their shared evolu$onary history •  Group by anatomical features •  Group by biochemical pathways (bacteria) •  Group by molecular traits (mainly DNA sequence) -  Scien$sts now rely heavily on comparison of molecular traits, but tradi$onal evolu$onary trees were built by comparing anatomical features. -  Trait used for comparison must be heritable With limited anatomical features, clinicians tradi$onally relied on biochemical tests for bacterial iden$fica$on Gram Posi$ve? No Gram Nega$ve Yes Bacillus? Coccus? Catalase nega$ve? Catalase posi$ve? Yes Yes Alpha hemoly$c? Yes Sensi$ve to optochin? Yes S. pneumoniae No Beta hemoly$c? Does it produce endospores?
No Yes Lactobacillus Clusters of cocci? Tetrads of cocci? Bacillus species species Yes Yes Staphylococcus species Yes Staphylococcus pyogenes Micrococcus species The advent of molecular techniques has allowed comparison using “markers” present in all organisms Gold standard is ribosomal small subunit rRNA gene, but others are also used Nucleo$de sequence within the conserved or variable region can be compared with that from another species 0 100 V1 200 V2 300 400 500 600 700 V3 Conserved regions Variable regions V4 800 900 0 V5 1000 1100 V6 1200 V7 1300 1400 V8 1500 V9 Why is the 16S gene a good phylogene$c “marker” •  The gene is universally conserved (all living organisms contain this gene). •  The sequence has both highly conserved (unchanging) regions and variable regions. •  The length (~1500 nucleo$des) provides a good amount of material for comparison but is small enough for easy sequencing. Isolate DNA from pure culture* SSU LSU 5S Amplify 16S using PCR Popula$on of iden$cal PCR products *culture does not need to be pure if PCR is followed by cloning step (see metagenomics lecture) Sequence and compare to known 16S sequences in database Sec$on 9 Learning Goals ü Explain why the ribosomal 16S gene is a good “marker” for molecular phylogene$c comparisons. •  Be able to interpret a phylogene$c tree. •  Explain the origins of mitochondria and chloroplasts and the evidence that supports this theory. Oversimplified view of molecular comparison to
build phylogenetic trees
•  The two sequences must be aligned to produce the most matches -  Some$mes gaps need to be introduced -  Whether and where to introduce gaps is subjective •  The number of matches is counted Organism 1: ACTGCATACCG -­‐AGGCTCT Organism 2: ACTGCTTACGGTAGGCTCT Adapted from Salyers and Whitt (2001) Microbiology: Diversity, Disease and the Environment
Oversimplified view of molecular
comparison to build phylogenetic trees
•  X is a common ancestor •  A and B share the most nucleo$des (10), so they diverged from each other more recently than from C •  B and C share 6 nucleo$des; A and C share 6 nucleo$des; we can’t tell which is more closely related to C, thus their distance of separa$on from C is the same. Adapted from Salyers and Whitt (2001) Microbiology: Diversity, Disease and the Environment
Building phylogene$c trees •  A phylogene$c tree is a structure in which species are arranged on branches that link them according to their rela$onship and/or evolu$onary descent. •  Gene$c distance is the number of muta$on/evolu$onary events between species since their divergence (but not all muta$on events are revealed in the current sequences). •  There are many methods that can be used to build trees, each resul$ng in differences in the final outcome. Computers can do much of the work, but human input is needed in much of the decision making. •  All methods group more similar sequences together. •  One must be careful not to over-­‐interpret taxonomic informa$on gained through less sophis$cated tree-­‐building methods. Of the choices listed, which group is the most divergent from its closest rela$ve? C
B
DE
F
G
A) Group G B) Group F C) Group C D) Group B A
Of the choices listed, which group is the most divergent from its closest rela$ve? C
B
DE
F
G
A) Group G B) Group F C) Group C D) Group B A
The branch length between groups is indica$ve of relatedness Of the choices listed, which two groups share the most evolu$onary history? C
B
DE
F
G
A)  Group C and D B)  Group B and A C) Group D and E D) Cannot determine A
Of the choices listed, which two groups share the most evolu$onary history? C
B
DE
F
G
A)  Group C and D B)  Group B and A C) Group D and E D) Cannot determine A
The distance between D and E is the same as between E and F; therefore, we cannot determine the answer from these data Sec$on 9 Learning Goals ü Explain why the ribosomal 16S gene is a good “marker” for molecular phylogene$c comparisons. ü Be able to interpret a phylogene$c tree. •  Explain the origins of mitochondria and chloroplasts and the evidence that supports this theory. Comparison of DNA sequences allows us to compare across all life But the eukaryotes were originally grouped based on phylogene$c traits What are the differences that dis$nguish the prokaryotes from the eukaryotes? What are the differences that dis$nguish the prokaryotes from the eukaryotes? Some possible answers: •  Prokaryotes do not have membrane-­‐bound organelles; eukaryotes do. •  Prokaryotes reproduce asexually, eukaryotes undergo sexual reproduc$on •  The prokaryotes are evolu$onarily more ancient and diverse What are the differences that dis$nguish the prokaryotes from the eukaryotes? Some possible answers: •  Prokaryotes do not have membrane-­‐
bound organelles; eukaryotes do. •  Prokaryotes reproduce asexually, eukaryotes undergo sexual reproduc$on •  The prokaryotes are evolu$onarily more ancient and diverse e.g. mitochondria The evolu$on of mitochondria allowed cells to take advantage of oxygen in cellular respira$on First cellular life (prokaryotes) Oxygen accumula$on Oldest cyanobacteria First fossils eukaryotes (photosynthesis) Earth’s crust cools Mul$cellular eukaryotes Early animals oxygena$on 4 3 2 1 Billion years from present Using oxygen as a terminal electron acceptor allows extrac$on of much more energy from the input molecule (e.g. glucose) 0 Models for the origin of the eukaryotic cell
Eukarya
Bacteria
animals
plants
Archaea
Ancestor
of chloroplast
Nucleus
formed
Ancestor of
mitochondrion
Eukarya
Bacteria
animals
plants
Ancestor
of chloroplast
Nucleus
formed
Ancestor of
mitochondrion
Adapted from Brock Biology of Organisms Fig. 14.10 (12th ed.) Archaea
Strong evidence indicates that mitochondria originated as a bacterium that was engulfed by or invaded another cell •  Mitochondria are the same size and shape of bacteria. •  Both have small, circular DNA genomes. •  Mitochondrial replica$on is autonomous from that of the “host” cell. •  Both have ribosomes of the eukaryo$c type. •  The electron transport chain is located in the cell membrane of bacteria (inner membrane of gram-­‐
nega$ves) and the inner membrane of mitochondria. Whole genome analysis supports the hypothesis
that the mitochondrial ancestor was most closely
related to the Alphaproteobacteria
Thrash et al. (2011) Sci Rep. 1:13 Sec$on 9 Learning Goals ü Explain why the ribosomal 16S gene is a good “marker” for molecular phylogene$c comparisons. ü Be able to interpret a phylogene$c tree. ü Explain the origins of mitochondria and chloroplasts and the evidence that supports this theory.