Download Evolution of colour vision in primates

STUDENT’S GUIDE
Case Study
Evolution of colour
vision in primates
Jarosław Bryk
Max Planck Institute for
Evolutionary Biology, Plön
Version 1.3
Case Stu
Case Stud
evolution of colour vision in primates
Evolution of colour
vision in primates
Colour vision evolved relatively early: most bony fish, reptiles and birds
have excellent colour vision. The first mammals, however, are thought to
have been largely nocturnal species. For them, the ability to see in lowlight conditions was at a premium, rather than the ability to see in colour.
The ancestors of modern mammals consequently lost colour vision at the
time of the dinosaurs. Some primates, including humans of course, have
re-evolved colour vision. This ability applies mainly to Old World species.
In contrast, with the exception of Howler monkeys, all male and some
female New World primates lack colour vision.
Most — 80% according to some estimates — of the information our brains
receive about their surroundings is transmitted via the sense of sight.
Seeing is such a good way of experiencing the environment that cameralike eyes have evolved independently more than 50 times. In mammals,
light is focused by the lens on a layer of specialised cells (cones and rods) in
the retina that contain visual pigments. These pigments are proteins called
opsins or rhodopsins. Each cone cell contains one of three sorts of opsins.
The different opsins are each sensitive to a different range of wavelengths
of light. Sometimes these are referred to as red, green and blue receptors,
although their peak sensitivities are violet, green and yellow-green.
Steven Walling, Wikimedia Commons.
In this activity you will explore the molecular basis and evolutionary
origin of trichromatic (red/green/blue) colour vision in humans and
our close evolutionary relatives using nucleic acid sequences of opsins,
key proteins involved in the process.
Nocturnal South American ‘night
monkeys’ (Autus spp.), like most
New World primates, lack fullcolour vision.
Section through the retina.
Notice how badly ‘designed’
the mammalian eye is: the
photoreceptors (the rods and cones)
face away from the light source.
Rod
DRAWINGS FROM: Wellcome Images.
Cone
Pigmented
cell
Interneurone
Copyright © Jarosław Bryk, 2011
Ganglion
cell
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evolution of colour vision in primates
Opsins are evolutionarily ancient proteins that take part in the process
of colour vision, while rhodopsins are involved in dim light vision. In this
exercise, we will concentrate on the opsins.
Humans are trichromats: we have three distinct opsins encoded by three
genes, named after the range of wavelengths they absorb: SWS absorbs
short-wavelength light, MWS, medium-wavelength and LWS, longwavelength light. The SWS opsin is encoded by a gene on chromosome 7 and
is sensitive to blue/violet light of 430 nm. The MWS and LWS opsins, subjects
of the following exercises, are both located on the X chromosome and are
sensitive to 530 nm (green) and 560 nm (green/yellow) light, respectively.
1.0
SWS
MWS
3PXO from the Protein Data Bank. Rendered using VMD software.
Each opsin is bound to 11-cis-retinal, a derivative of vitamin A. The 11-cisretinal is the light-sensitive molecule, but the way it reacts to light depends
on the opsin molecule it is attached to. Different opsins absorb different
wavelengths of light, and each type has evolved so that it is sensitive to
a specific, narrow range of wavelengths. This reaction can be studied
experimentally: opsin proteins can be synthesised and/or mutated in cells
on a Petri dish, isolated and then mixed with 11-cis-retinal to obtain visual
pigments. The absorption spectra of these pigments can be then measured
in a spectrophotometer to identify the wavelengths of light they absorb.
Thus it is possible to compare properties of different visual pigments and
identify features responsible for their fine-tuning.
N
C
Structure of an opsin. Opsins are
trans-membrane proteins. There
are seven helices in the protein
which are anchored in the cell
membrane. The 11-cis-retinal
(shown in red) lies near the centre,
within the helices.
LWS
DATA FROM: Stockman, et al (1993).
Absorbance
0.8
0.6
0.4
0.2
0.0
400
450
500
550
600
650
700
Wavelength (nm)
Simplified human cone response
curves. SWS= short-wavelengthsensitive opsin; MWS= mediumwavelength-sensitive opsin; LWS =
low-wavelength-sensitive opsin.
Let’s start by comparing the nucleic acid* sequences of human MWS and
LWS opsins to see exactly how different they are.
* the sequences are mRNA sequences in which the uracil (U) of RNA has been
replaced by thymine (T). Hence they look like DNA. RNA sequences are stored in
GenBank (an online database of sequence data) in this way.
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evolution of colour vision in primates
Comparing human MWS or LWS opsins
1. Double-click on the human_green_red.geneious document. This will
start the Geneious software and load two files of nucleic acid sequence
data into the programme. Note: If a box appears over the Geneious startup screen, saying that your trial of the ‘Pro’ version has ended, click on Use
Geneious Basic.
2. The main Geneious window will now show the protein-coding part
of the mRNA sequences of human MWS (green-light-sensitive) and
LWS (red-light-sensitive) opsins. Remember, these will look like DNA
because the ‘U’ (uracil) of RNA has been replaced by a ‘T’ (thymine).
Zoom buttons
Selecting ‘Wrap’ here means
you can scroll down through
the sequences.
Names
RNA sequences
3. Ensure that the two files are highlighted in the upper Geneious window,
then click the Alignment button. Click OK to accept the default Geneious
settings. The programme will now align both of the sequences, and a
new file of the aligned sequences will appear in the top panel and be
displayed in the main Geneious window:
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Alignment button
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evolution of colour vision in primates
4. Use the magnifying glass buttons to zoom in on the data. Quickly
scroll through the data, looking for any differences between the two
sequences. Don’t spend too long on this — you just want to obtain a
general impression of the data.
Questions
a. Do you think there are many differences between the two sequences?
What is the sequence similarity between the two genes? Hint: use the
Statistics panel in Geneious to obtain this information (the % tab on the
lower right of the screen).
b. Which of the differences between the sequences might be responsible
for the different properties of the proteins? Can you tell from the RNA
sequences themselves? Do you think that looking at the amino acid
sequences might be more helpful? Hint: click on the Translate button to
see the amino acids encoded by the mRNA.
The Geneious statistics panel
Translate button
c. How might a scientist be able to test whether the differences in the
proteins influence their sensitivity to different wavelengths?
The MWS and LWS opsin genes are very similar. In fact, their DNA sequences
are almost 98 % identical, even including the non-coding sequences of their
introns. There are only 24 nucleotides that differentiate the two coding
sequences, which corresponds to 15 amino acid changes between the
proteins encoded by these genes (why not 24 amino acid changes?).
Each of these amino acid changes influences the sensitivity of an opsin
to certain wavelengths of light. Thanks to experimental work on opsins
in vitro we now know that the most critical changes are in amino acid
positions 180, 277 and 285. If you know which amino acids are in these three
positions, it is possible to predict which wavelength of light a particular
opsin will be sensitive to.
Amino acid position
Opsin
Max. absorption (nm)
180
277
285
Human MWS (green)
561
S (serine)
Y (tyrosine)
T (threonine)
Human LWS (red)
530
A (alanine)
F (phenylalanine)
A (alanine)
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evolution of colour vision in primates
Both the MWS and LWS genes are positioned very close to each other and
in a head-to-tail fashion on chromosome X in all species of Old World
monkeys, which, in addition to the high sequence similarity, suggests that
they originated by duplication. This is one of the major ways new genes can
emerge, and we can use their sequences to explore when the duplication
occurred and to work out which sequence is likely to be the ancestral one.
Chromosome 7
with SWS gene
X chromosome
with MWS and
LWS genes
Question
d. How might you check whether the MWS and LWS duplications occurred
independently within humans and other species of Old World primates
or whether they have had a common origin before the Old World
monkeys evolved?
Multiple
MWS genes
Single
The origin of MWS and LWS opsins
LWS gene
We can answer the question above by comparing pairs of sequences (of
MWS and LWS opsin genes) from several species of Old World primates.
If all of the MWS and LWS sequences are descendants of an ancient
duplication, then the MWS sequences should be more similar to one
another than they are to the LWS sequences. This would give a tree similar
to Possibility One, below.
Location of the opsin genes in
humans. The genes on the X
chromosome are arranged head-totail.
If, however, the duplications occurred independently in each of the species,
the MWS and LWS sequences should be most similar within each species,
corresponding to Possibility Two, below.
Let’s investigate these two possibilities using mRNA sequences from a few
species of Old World monkeys.
MWS
MWS
MWS
MWS
MWS
LWS
LWS
LWS
LWS
LWS
MWS
LWS
MWS
LWS
MWS
LWS
POSSIBILITY ONE
Ancient duplication of opsin gene
(Duplication events marked with a blue dot)
Copyright © Jarosław Bryk, 2011
SPECIES SPECIES SPECIES SPECIES
1
2
3
4
Outgroup
SPECIES SPECIES SPECIES SPECIES
1
2
3
4
Outgroup
POSSIBILITY TWO
Independent duplication of opsin gene
(Duplication events marked with a blue dot)
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evolution of colour vision in primates
Chimpanzee
Julie Langford, Wikimedia Commons.
6. The main Geneious window will show the mRNA sequences of
fragments of the MWS and LWS opsins from a human, a chimpanzee,
a gorilla, a macaque, a diana monkey and a talapoin monkey. There is
also sequence of chicken iodopsin to be used as an outgroup, to which
all other sequences will be related to in a phylogenetic tree.
Thomas Lersch, Wikimedia Commons.
5. Double click on the origin_green_red.geneious document. This will
load a file of 13 ready-aligned mRNA sequences into the programme.
Gorilla
Eric Bajart, Wikimedia Commons.
7. Click the Tree button to open tree settings panel. Select TamuraNei as the Genetic Distance Model, Neighbor-Joining as the Tree build
Method and use Chicken iodopsin as an outgroup. Select the Resample
tree option and change the Number of replicates to 1000 and Support
Threshold % to 5. Ensure that Create Consensus Tree is selected. Leave
all the other options at the default values and click OK to build the tree.
Tony Hisgett, Wikimedia Commons.
Macaque
G. Drange, www.biopix.com
Diana monkey
Talapoin monkey
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evolution of colour vision in primates
Questions
e. How are the MWS opsins and LWS opsins grouped on the tree?
f. Which of the two possible patterns of evolution suggested in the two
diagrams on page 6 does your tree support? (Independent duplication
the opsin in each species, or duplication in an ancestor of all the
species?)
Now let’s see what the tree would look like if we add MWS and LWS opsins
from a more distantly-related, New World species — a howler monkey.
8. Double-click on the origin_green_red_howler.geneious document.
This will load some aligned mRNA sequence data into the programme.
10. Click the Tree button to open tree settings panel. Select TamuraNei as the Genetic Distance Model, Neighbor-Joining as the Tree build
Method and use Chicken iodopsin as an outgroup. Select the Resample
tree option and change the Number of replicates to 1000 and Support
Threshold % to 5. Ensure that Create Consensus Tree is selected. Leave
all the other options at the default values and click OK to build the tree.
Copyright © Jarosław Bryk, 2011
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Alessandro Catenazzi, Wikimedia Commons.
9. The main Geneious window will now show the mRNA sequences of part
of the MWS and LWS opsins from a human, a chimpanzee, a gorilla, a
diana monkey, a talapoin monkey, a macaque (all Old World primates)
plus a New World howler monkey. As before, there is also the sequence
of chicken iodopsin, to act as an outgroup.
Red howler monkey
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evolution of colour vision in primates
Questions
Study the tree you have just produced.
g. Howler monkeys can see in full colour: they are, like most humans and
Old World monkeys, trichromatic. What can you tell about the origin of
trichromatic colour vision in the howler monkey from this tree?
h. Can you tell which of the sequences — MWS or LWS — is ancestral (in
other words, which gene was duplicated: MWS or LWS?)
i. How do we know that howler monkeys did have a gene duplication and
not just different alleles of the same gene?
The howler monkey is a unique New World species that has acquired
trichromatic colour vision by gene duplication, similar to the one that
happened in Old World monkeys. All other New World monkey species that
have evolved trichromatic vision have done so without gene duplication.
Dario Sanches, Wikimedia Commons.
Different ways of evolving trichromatic
colour vision
Brown howler monkey
Question
j. Could you suggest how it would be possible to achieve trichromatic
colour vision with just two genes? Would it be possible with just one?
Almost all species of New World monkeys have trichromatic colour vision
based on only two genes. One of these genes (short-wavelength sensitive,
which is sensitive to blue light), similarly to Old World monkeys, is located
on an autosome. The other opsin gene is on the X chromosome. In contrast
to the Old World species, the gene on the X chromosome did not undergo a
duplication but it exists as different alleles in the population. This means
that alleles of MWS gene located on each of the X chromosomes may have
different sequences and thus, the proteins encoded by these alleles may
have different absorption spectra. And because one of the genes is on the
X chromosome, it also implies that the colour-sensing abilities of the two
sexes may be different.
Questions
k. What might the differences be between male and female New World
monkeys? Compare and contrast the situation in New World monkeys
with humans suffering from colour blindness.
l. Trick question: Do you think that different alleles of MWS and LWS
genes exist in humans? If you think they do, how might this affect the
colour vision abilities of humans? If you think we don’t have different
alleles, why not?
Copyright © Jarosław Bryk, 2011
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Colour blindness
Colour blindness, sometimes called
‘Daltonism’ after the English
chemist who first described it
in detail, is most commonly
inherited from mutations on the X
chromosome. The human genome
project has shown, however, that
mutations capable of causing
colour blindness originate from at
least 19 different chromosomes and
56 different genes.
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evolution of colour vision in primates
In New World monkeys, with the exception of the howler monkey, males
are dichromatic: they only have an SWS (blue-light-sensitive) opsin on their
autosome and an MWS (green-light-sensitive) opsin on their X chromosome.
But because the MWS opsin gene is very polymorphic (there are many
alleles in the population) and proteins encoded by the different variants
of the gene may be sensitive different wavelengths, individual males may
see the world differently. In fact, in New World monkeys scientists do not
call the opsins MWS or LWS, but simply give them numbers indicating the
wavelength with the maximum absorption of light for that variant of the
opsin. In any case, New World monkey males are all colour blind, but each
individual may be blind to a different colour.
Male New World monkey
Autosome
with SWS gene
X chromosome
with opsin allele
Dichromat
Colour blind
Female New World monkeys
New World monkey females, in contrast, can be trichromats if the two
alleles on their two X chromosomes produce proteins sensitive to different
wavelengths. If the females have identical alleles on both X chromosomes,
then they are dichromats and, just like the males, they can be colour blind
to different colours.
Colour blindness
Dichromat
Colour blind
The inability to distinguish red and green colours is the most common
form of colour blindness in humans, affecting up to 10% of population. It is
caused by missing or mutated MWS or LWS opsin genes.
Trichromat
Colour vision
Question
James King-Holmes, Science Photo Library.
m. Why are men colour blind much more often than women are?
Copyright © Jarosław Bryk, 2011
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The chemist John Dalton was the
first person to describe red-green
colour blindness (his brothers’ and
his own) in detail. He requested
that his eyes were dissected after
his death because he suspected that
the reason for his condition was
blue fluid in his eyeballs. After his
death this was shown to be false,
but some 150 years later DNA from
his preserved eyeballs (shown on
the left) was analysed and it showed
that Dalton had had deuteranopia:
his MWS gene was missing.
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evolution of colour vision in primates
12. Click the Tree button to open tree settings panel. Select TamuraNei as the Genetic Distance Model, Neighbor-Joining as the Tree build
Method and use Chicken iodopsin as an outgroup. Select the Resample
tree option and change the Number of replicates to 1000 and Support
Threshold % to 5. Ensure that Create Consensus Tree is selected. Leave
all the other options at the default values and click OK to produce the
tree.
Spider monkey
Anonymous, Wikimedia Commons.
11. Double click on the origin_new_old_world.geneious document. This
will load a file of 26 aligned mRNA sequences into the programme. The
main Geneious window will show the mRNA sequences of part of the
MWS and LWS opsins from the Old World primates we used before and,
in addition, some New World monkeys: a spider monkey, a squirrel
monkey, a weeper capuchin and a marmoset. As before, there is also
a sequence of chicken iodopsin to be used as an outgroup, to which all
other sequences will be related in a phylogenetic tree.
Lea Maimone, Wikimedia Commons.
Let’s now compare opsin sequences from the Old and New World primates
to see how they are related.
Squirrel monkey
Convergent evolution of colour vision in Old
and New World monkeys
Weeper capuchins
Manfred Werner, Wikimedia Commons.
n. Identify which species on your tree are Old World primates, and which
are New World primates.
o. Describe how the MWS and LWS opsins of the Old World primates are
distributed on the tree.
p. Some of the New World monkeys have their opsins described as a
number. The number is the wavelength the opsin is sensitive to.
Describe how the New World monkey’s opsins are arranged on the tree.
q. Do you think the New World opsins are more closely related to the
MWS or the LWS opsins of Old World primates?
Pacman, Wikimedia Commons.
Questions
Marmoset
One of the possible explanations for this pattern of variation is that is it a
result of convergent evolution. In many different species, a similar sensitivity
of opsins is achieved with the same mutations, thus making even unrelated
sequences quite similar, confusing the phylogenetic relationships between
species. As you have seen in the first exercise, there are 24 mutations
differentiating MWS from LWS opsin genes in humans, but only three of
these mutations are responsible for majority of the sensitivity shift. Let’s
compare sequences from other species that have similar sensitivity to
human MWS and LWS opsins to see what mutations are responsible for
their properties.
Copyright © Jarosław Bryk, 2011
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evolution of colour vision in primates
13. Double click on the convergence561.geneious document. This will load
the file of ready-aligned mRNA sequences into the programme.
14. The main Geneious window will now show part of the protein-coding
mRNA sequences of pairs opsins from a chimpanzee, howler monkey,
a weeper capuchin monkey and a squirrel monkey.
15. Use the Translate button to translate the nucleotide sequence into
protein sequence.
16. Ensure that the sequences are selected in the top window, then click
on the Alignment button. When the panel appears, click OK to accept
the Geneious default settings. The programme will now align the amino
acid sequences.
17. Use the magnifying glass buttons to zoom in on the data.
18. Scroll through the data, and look at positions 34, 89 and 97 (they
are equivalent to positions 180, 277 and 285 in the full-length opsin
sequence). Compare what you find with the data in the table on page 5.
All of these species achieve the 561 sensitivity of their opsins by identical
key mutations as in Old World monkeys, even though these sequences are
distantly related. Indeed, if we compared the whole sequences of many
opsins sensitive to certain wavelengths, we would find out that apart from
those key mutations they are otherwise different, thus emphasising the
importance and uniqueness of the key amino acid positions in the opsins.
Questions
r. Can you think of the reasons why there seems to be a limited number
of ways to evolve opsins sensitive to certain wavelengths?
s. Last but not least: why do you think colour vision evolved in the first
place?
Further reading
The making of the fittest. DNA and the ultimate forensic record of evolution by
Sean B. Carroll (2009) Quercus Books (Paperback) ISBN: 978 1847247247.
A popular account of some of the molecular evidence for evolution. Chapters four
and six cover the evolution of colour vision in primates.
Additional references are given in the Teacher’s guide.
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