UC Davis EVE161 Lecture 13 by @phylogenomics

Lecture 13:

EVE 161: 
Microbial Phylogenomics
!

Lecture #13:
Era III: Genome Sequencing and
Phylogenomic Analysis
!
UC Davis, Winter 2014
Instructor: Jonathan Eisen

Slides for UC Davis EVE161 Course Taught by Jonathan Eisen Winter 2014

!1

Where we are going and where we have been

• Previous lecture:
! 12: Guest Lecture
• Current Lecture:
! 13: Genome Sequencing III
• Next Lecture:
! 14: Metagenomics


!2

Phylogenomics I:Major Evolutionary Transitions


Phylogenomics I:Major Evolutionary Transitions

• Analysis of S. pombe genome by Wood et al
2002

• Compared the genomes of eukaryotes to
those of prokaryotes

• “Are there genes found in all eukaryotes
with no obvious homologs in any
prokaryote?”


Evolutionary Model

S. pombe

Eukaryotes

S. cerevisiae

Encephalatozoon

Archaea
Bacteria

Worm
Fly
Humans
Dictyostelium

Arabidopsis
Chlamydomonas
Phytophthora
Tetrahymena
Plasmodium
Trypanosoma
Euglena
Naegleria
Trichomonas

Giardia


Eukaryotic Specific Genes
• >200 genes found including:

– Cytoskeleton components: tubulin,
ankyrin, myosin

– Protein degradation: ubiquitin, proteases

– Chromatin and DNA packaging

• Of the 200 many had no known function:
could encode novel eukaryotic wide
processes

Multi- vs. Single-Cellular Eukaryotes
• Further analysis of S. pombe genome
• Compared multi-cellular vs. single-cellular eukaryotes
(animals and plants vs. yeast)
• “Are there genes in all multi-cellular and not in any singlecellular?”
• Found only 3
• Concluded that the genetic basis of multi-cellularity was
likely to be gene regulation and not invention of new genes


Multiple Origins of Multicellularity


Phylogenomics II: 
Endosymbiont Evolution


Endosymbiont Evolution

• Compared to free-living relatives

–
–
–
–

Smaller genomes

Lower GC content

Higher pIs

Higher rates of sequence evolution

• Baumannia shows ALL of these


Uses of Whole Genome Trees


Wolbachia Evolution


Variation Between Endosymbionts and Free Living

• Repair hypothesis

!

• Population genetics hypothesis

!


Variation Between Endosymbionts and Free Living


!


!

• PopGen explanations favored


Variation Among Endosymbionts



MutS

MutL

+

+

+

+

+

+

+

+

_

_

_

_




!


!




!


!

• Repair explanations favored


Phylogenomics III: 
Lateral Gene Transfer


Vertical Evolution

From C. Darwin, origin of species, via W. F. Doolittle

Vertical Inheritance - Binary Bission


Lateral inheritance I: Competence


Lateral inheritance II: Conjugation


Agrobacterium conjugation


Figure 7.19 - Ab transfer


Lateral inheritance III: Transduction


Pathogenicity Island


Steps in Lateral Gene Transfer (LGT)
A

B

C


D

A

B

C

1

D

Gene acquires host features


A

B

C

D

2

Transfer

1



A

B

C

3-5

D

Integration, selection, spread
2

Transfer

1



A

B

C

D

Amelioration
Integration, selection, spread
6

3-5

2

Transfer

1



How to Infer Gene Transfers

• Unusual distribution patterns

!

• Unusual nucleotide composition

!

• High sequence similarity to supposedly
distantly related species

!

• Unusual gene trees

!

• Observe transfer events


Case Study I: Aphids

ig. 1 Coloration and carotenoids in the pea aphid. Typical green (A) and red (B) aphid clones, (C) 5AY, a green mutant clone arising from the red clone 5A. (D)
roﬁles of carotenoids in red (5A, LSR1), mutant redgreen (5AY, two samples), and green (8-10-1, 7-2-1) pea aphid clones. Torulene and a related red compound
re restricted to red clones; the mutant 5AY clone lacks these and displays an elevation in their predicted precursor, -carotene.



Table 1 Genes in the A. pisum genome with closest homology to carotenoid biosynthetic enzymes, including scaffold of origin and matching EST sequences.
Similar color indicates that the gene is on the same scaffold. The 3' end of scaffold NW_001925130 overlaps with the 5' end of NW_001923501 for 5400 base
pairs, and PCR demonstrated continuity of these scaffolds. Pink row is the gene corresponding to torR and conferring red color (see text). Protein length, amino
acids; ESTs are those present in GenBank, mostly from clone LSR1.



Fig. 2 Phylogenetic relations of inferred carotenoid biosynthetic enzymes from the pea aphid genome. (A) Carotenoid desaturases and (B) carotenoid cyclase–
carotenoid synthases. Sequences are from aphids, bacteria, plants, and fungi; no homologs were detectable in other sequenced animal genomes. Bootstrap
support greater than 50% is indicated on branches.

!


Case Study II: GEBA


Tree of Life

Figure from Barton, Eisen et al. “Evolution”, CSHL Press based on Baldauf et al Tree

Genomes Poorly Sampled


TIGR Tree of Life Project



!41

Genomes Still Poorly Sampled


Genomic Encyclopedia of Bacteria & Archaea
Wu et al. 2009 Nature 462, 1056-1060


GEBA Lesson 1: rRNA utility in IDing novel genomes

From Wu et al. 2009 Nature 462, 1056-1060


!45

GEBA Lesson 2: rRNA Tree is not perfect

16s

WGT, 23S

Badger et al. 2005 Int J System Evol Microbiol 55: 1021-1026.

!46

GEBA Lesson 3: Phylogenetic sampling improves annotation

• Took 56 GEBA genomes and compared results vs. 56
randomly sampled new genomes
• Better definition of protein family sequence “patterns”
• Greatly improves “comparative” and “evolutionary”
based predictions
• Conversion of hypothetical into conserved hypotheticals
• Linking distantly related members of protein families
• Improved non-homology prediction


!47

GEBA Lesson 4 : Metadata Important


!48

GEBA Lesson 5:Improves discovering new genetic diversity


!49

Protein Family Rarefaction Curves
• Take data set of multiple complete genomes
• Identify all protein families using MCL
• Plot # of genomes vs. # of protein families


!50

Wu et al. 2009 Nature 462, 1056-1060

!51

Synapomorphies exist

Wu et al. 2009 Nature 462, 1056-1060

!52

Phylogenetic Distribution Novelty:
Bacterial Actin Related Protein

87

C. boidinii gi57157304
S. cerevisiae gi14318479
L. starkeyi gi166080363
S. japonicus gi213407080
ACTIN
A. cliftonii gi14269497
U. pertusa gi50355609
99
H. sapiens gi4501889
M. cerebralis gi46326807
67
C. cinerea gi169844021
ARP1
N. crassa gi85101929
100
I. scapularis gi215507378
100 H. sapiens gi5031569
51
65
100
ARP2
D. melanogaster gi24642545
100 G. gallus gi45382569
75
C. neoformans gi58266690
ARP3
100
100 H. sapiens gi5031573
H. ochraceum gi227395998
BARP
P. patens gi168051992
ARP4
73
99
A. thaliana gi18394608
94
100
ARP5
D. discoideum gi66802418
74
97
ARP6
100
D. hansenii gi21851 1921
100
O. sativa gi182657420
ARP7
A. thaliana gi1841 1737
D. melanogater gi19920358
100
M. musculus gi226246593
ARP10

0.5

Haliangium ochraceum DSM 14365

Patrik D’haeseleer, Adam Zemla,
Victor Kunin

See also Guljamow et al. 2007 Current Biology.


GEBA Cyanobacteria

A
B1

B2
B3

D

Paulinella
C1

C2
C3
A
E

Glaucophyte
Green
Red

G

Chromalveolates

F
0.3

B

Shih et al. 2013. PNAS 10.1073/pnas.1217107110

Fig.
mum
noba

Haloarchaeal GEBA-like

Lynch et al. (2012) PLoS ONE 7(7): e41389. doi:10.1371/journal.pone.0041389

The Dark Matter of Biology

From Wu et al. 2009 Nature 462, 1056-1060


GEBA Uncultured
SAR

A: Hydrothermal vent
B: Gold Mine
C: Tropical gyres (Mesopelagic)
D: Tropical gyres (Photic zone)

OP3

Site
Site
Site
Site

OP1

406

OD1

1

Number of SAGs from Candidate Phyla

4
6
1

1
13
-

2
-

2
-

Sample collections at 4 additional sites are underway.

Phil Hugenholtz

!57


JGI Dark Matter Project
brackish/freshwater

TG

HSM
SM

GBS
GBS

HOT
OT

SAK
AK

hydrothermal

sediment
ETL
E

BACTERIA

ARCHAEA

UGA recoded for Gly (Gracilibacteria)

seawater

HGT from Eukaryotes (Nanoarchaea)

bioreactor

EPR
EPR

T
TA

G
GOM
OM

Growing
AA chain

U

oxidoretucase
Ribo

A

P51$

environmental
samples (n=9)
draft genomes
(n=201)

W51$*O

recognizes
UGA

G

isolation of single
cells (n=9,600)

SSU rRNA gene
based identification
(n=2,000)

whole genome
amplification (n=3,300)

U

genome sequencing,
assembly and QC (n=201)

1

H

H
1

1

$,$5

adenine

+2+2

+2+2

OH

2+3

Woyke et al. Nature 2013.

limiting
phosphate,
fatty acids,
carbon, iron SpotT

1+ 2

1$'+

51$ SROPHUDVH

ı3

ı2
-10

ı1

GTP or GDP
+ATP

limiting
amino acids
RelA

ppGpp
(GTP or GDP)
+ PPi

H

DksA

Expression of components
for stress response
O

OH

+2+2

O
O

O

1+

1+

2+3

2+3

tetrapeptide

e- acceptor

stringent response
(Diapherotrites, Nanoarchaea)

H

+2+2
O

IMP

1

1

O

O
1+

ı4
-35

)$,$5

1

guanine

O

PurP

O

+2 1

H

H H

+

ȕ ȕ¶

Į7'

?
H 1

+

1+2

O
Oxidation

Archaea

PurF
PurD
3XU1
PurL/Q
PurM
PurK
PurE
3XU
PurB

1+2

1

O

Į17'

archaeal type purine synthesis
(Microgenomates)

1

Eukaryota
ADP

sigma factor (Diapherotrites, Nanoarchaea)

ribosome

PRPP

Reduction

1$'+ + + H-

A U
A U
G U
A A U G A U

Ribo

1+

H

Korarchaeota
Cren Thermoprotei
Thaumarchaeota
Cren MCG
Cren pISA7
Cren C2
Aigarchaeota
Nanoarchaea
Micrarchaea
pMC2A384 (Diapherotrites)
DSEG (Aenigmarchaea)
Nanohaloarchaea
Euryarchaeota
:6
OP11 (Microgenomates)
OD1 (Parcubacteria)
SR1
BH1
TM7
GN02 (Gracilibacteria)
Bacteriodetes
OP1 (Acetothermia)
'HLQRFRFFXVí7KHUPXV
093í
70
ZB3
)LEUREDFWHUHV
TG3
Spirochaetes
WWE1 (Cloacamonetes)
Proteobacteria
)LUPLFXWHV
Tenericutes
)XVREDFWHULD
Chrysiogenetes
Chlorobi
6$5 0DULQLPLFURELD

Caldithrix
GOUTA4
Acidobacteria
Elusimicrobia
Nitrospirae
49S1 2B
Chloroflexi
Caldiserica
AD3
OP9 (Atribacteria)
:36í2
Synergistetes
Thermodesulfobacteria
Deferribacteres
CD12 (Aerophobetes)
OP8 (Aminicenantes)
AC1
SBR1093
SPAM
GAL15
Dictyoglomi
EM3
Thermotogae
Aquificae
GAL35
EM19 (Calescamantes)
2FW6SDí )HUYLGLEDFWHULD

Deltaproteobacteria
Cyanobacteria
:36í2
Actinobacteria
Gemmatimonadetes
NC10
SC4
WS2
NKB19 (Hydrogenedentes)
WYO
Armatimonadetes
WS4
Planctomycetes
Chlamydiae
OP3 (Omnitrophica)
Lentisphaerae
Verrucomicrobia
BRC1
Poribacteria
WS1
+Gí
LD1
GN01
WS3 (Latescibacteria)
GN04

+

e- donor

ADP

archaeal toxins (Nanoarchaea)

1+
2+3
tetrapeptide

murein (peptido-glycan)

lytic murein transglycosylase


A Genomic Encyclopedia of Microbes (GEM)


GEBA Lesson 6: Improves analysis of metagenomic data


!61

Other Markers
Sargasso Phylotypes
0.500

GEBA Project improves
metagenomic analysis
EFG
EFTu
HSP70
RecA
RpoB
rRNA

0.250

0.125

us
ar
ch
ae
ot
C
a
re
na
rc
ha
eo
ta

er
m

er
ia
ct

Th

ba

Eu

cc
u
in
o

co

ry

s-

so

iro
c

ha
et

es

ex
i
Sp

hl
or
oﬂ

C

FB
C

Fu
De

ap

ro
t

eo
ba
Be
ct
ta
er
pr
ia
ot
eo
ba
G
am
ct
er
m
ia
ap
ro
te
ob
Ep
ac
si
lo
te
np
ria
ro
te
ob
De
ac
lta
te
pr
ria
ot
eo
ba
ct
C
er
ya
ia
no
ba
ct
er
ia
Fi
rm
ic
ut
es
Ac
tin
ob
ac
te
ria
C
hl
or
ob
i

0.000

Al
ph

Weighted % of Clones

0.375

Major Phylogenetic Group

Venter et Eisen Winter 304:
Slides for UC Davis EVE161 Course Taught by Jonathan al., Science2014

66-74. 2004

!62

Venter et Eisen Winter 304:
Major Phylogenetic Group
Slides for UC Davis EVE161 Course Taught by Jonathan al., Science2014
ar
ch

re
n

C

ot
a

ae

a

ot

us

rm

ria

s

te

te

ae

ry
ar
ch

Eu

Th
e

s-

cu

oc

De
in
oc

ac

so
b

Fu

ae

ch

Sp
iro

xi

or
oﬂ
e

hl

C

FB

C

i

or
ob

hl

C

ria

te

ob
ac

tin

Ac

es

ut

ic

rm

Fi

ria

ria

te

ac

ob

ya
n

C

te

ia

er

ia

er

ct

ba
c

eo

pr
ot

lta

De

ct

eo
ba

ro
t

np

si
lo

ria

te

0.375

Ep

eo
ba

ro
t

ap

am
m

G

ba
c

eo

ia

er

ct

ba

eo

pr
ot

pr
ot

ta

ph
a

Be

Al

Weighted % of Clones

Other Markers

0.500

Sargasso Phylotypes

But not a lot
EFG
rRNA

66-74. 2004

EFTu

0.250

0.125

0.000

!63

rRNA Tree of Life

Bacteria

Archaea

Eukaryotes
Figure from Barton, Eisen et al. “Evolution”, CSHL Press.
2007.
Based on tree from Pace 1997 Science 276:734-740

!64

PD: Genomes 

From Wu et
al. 2009
Nature 462,
1056-1060


!65

PD: Genomes + GEBA 

From Wu et
al. 2009
Nature 462,
1056-1060


!66

PD: Isolates

From Wu et al. 2009 Nature 462,
Slides for UC Davis EVE161 Course Taught by Jonathan Eisen Winter 2014 1056-1060

!67

UC Davis EVE161 Lecture 13 by @phylogenomics

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