On the
Propagation of Mitochondrial Mutations
Abstract
How does
a new mutation get to appear in a mitochondrial
Address for correspondence:
Received:
Introduction
How does a new mutation get to appear in a
mitochondrial
Although the mechanisms of the production and
propagation of chromosomal mutations can be explained, the behaviour of
mitochondria and the propagation of mutations in the mtDNA are still very
poorly understood. This paper, in no
way, manages to resolve all the issues, but the steps involved in going from an
initial mutational event to the final state of homoplasmy
are discussed.
Background
The study of mitochondrial
The Phylogenetic Tree
The phylogenetic tree for human mtDNA is now very
extensive as it is found that only persons who have a maternal relationship
within a genealogical time frame have identical mitochondrial sequences. And, as one compares the mtDNA sequences from
persons who are more and more distantly related there is an increasing number
of mutational differences between the sequences.
There are several papers that describe the
phylogenetic tree; in particular, the paper from Kivisild,
et al. (2006) has some very useful diagrams.
This paper also gives estimates as to when the different branches of the
phylogenetic tree were formed. As an
example, the diagrams show that Haplogroup H and V have a common source from
about 16,000 +/- 6,000 years ago.
The author’s own mtDNA sequence shows his maternal
origin is in Haplogroup V and has this set of mutational differences from the
T72C
A263G 309.1C 315.1C A750G
A1438G A2706G A2880G
G4580A A4769G
C7028T A8860G A15326G
C15904T A16162G
T16298C
The list contains 16 mutations of which 14 are base
substitutions and 2 are insertions.
As Haplogroup V and Haplogroup H arose from a common
source it is possible to split this list of mutations into two parts.
The first part contains the 6 mutations that have
occurred after the split from the common source on the line leading to
Haplogroup V:
T72C A2880G G4580A C15904T
A16162G T16298C
whilst
the second part contains the other 10 mutations that occur on the line to the
A263G 309.1C 315.1C A750G
A1438G
A2706G A4769G C7028T
A8860G A15326G
However, if the list of mutations as given above is
viewed from the
G263A
C309d C315d G750A
G1438A
G2706A G4769A T7028C G8860A
G15326A
Figure 1
illustrates the list of mutations divided into these two parts.
Figure 1. The mutations that
have occurred along the lines leading to Haplogroups H and V from HV
So, over the last 16,000 years these 16 mutations have
appeared in the two lines to Haplogroups H and V; but how might this have
happened?
Personification
A point to be made at this stage is that in many places
in this paper, and in the accompanying computer model, it has been found useful
to personify a mutation as it passes through the different stages of its
propagation; but it is important to appreciate that all the steps occur at
random and a mutation does not have purpose, ambition, nor any
form of intelligence.
Modelling
the Propagation of Mitochondrial Mutations
A computer
model that simulates the propagation of mitochondrial mutations has been
written to accompany this paper. This
model shows how a new mutation might go through the phases of propagation of a
mutation from an initial mutational event to the states of heteroplasmy
and homoplasmy.
The
computer model is available in text form as supplementary file at:
http://www.jogg.info/51/nucloid_model.txt
And
as a web page, accompanied by instructions for use and a discussion of the modeling process at:
http://jogg/info/51/nucleoid_model.html
The
computer model is set up as a web page and can be viewed with any web
browser. However, the model runs
appreciably faster under Google Chrome, as compared to other browsers,
as this browser uses compiled Javascript.
Discussion:
Mitochondria
are small organelles found in all the cells of the body. They are mainly involved in the production of
“energy” within the cells and have been considered to act as small “power
stations.” But, whereas most processes
within a cell are controlled by the nucleus, mitochondria act independently and
are thought to have arisen from bacteria that have formed a symbiotic
relationship with the cell. For this
reason mitochondria appear to behave more like bacteria than nucleated cells.
Most cells
contain relatively few mitochondria, perhaps up to several hundred. But in a fully developed egg cell, an oocyte, there may as many as 200,000. Mitochondria appear to replicate by division
and this process is independent of cell division. So, in a cell that does not divide as it
matures, such as an oocyte, the number of
mitochondria may become very high.
However, in the cells of a tissue where there is a high rate of cell
division the number of mitochondria per cell falls to a very low number.
The recent
papers by Shoubridge and Wai
(Shoubridge, 2007, Wai,
2008) deal particularly well with the subject of the behaviour of mitochondria
in mammalian cells and discuss some of the problems concerning the emergence of
mitochondrial mutations.
The
Propagation of a Mitochondrial Mutation
A new
mitochondrial mutation cannot just appear suddenly in every one of an individual’s
billions of mitochondrial
But
because mitochondria act like bacteria it is possible to infer that a
mitochondrial mutation propagates in the same manner as a mutated bacterial
strain is known to arise. In bacteria
for the emergence of a new bacterial strain there needs in the first instance
to be a mutation occurring in the genetic code of just one organism, this
mutated bacterium then goes through many generations until bacteria with this
mutation form a new strain. In bacteria
a mutation can develop because it offers an advantage to the bacteria, or at
random with no advantage being present.
As
described with bacteria, the spread of a mitochondrial mutation has to begin
with an initial mutational event occurring in just one mtDNA molecule
and this is followed by the mutation gradually coming to be present in more and
more mtDNA molecules, until finally the state of homoplasmy
is reached when it is considered that every mtDNA molecule in the body has the
mutation.
In this
paper the gradual increase in the prevalence of a new mutation is described as
the propagation of a mutation; and this process which can be split into two
phases.
The first
phase includes the steps that may be involved in going from an initial
mutational event through to when many, or all, of the mtDNA molecules,
firstly in a single mitochondrial nucleoid, and later
a single mitochondrion, have the new mutation.
This first phase can occur in any cell in the body, and not necessarily
in a cell line that is going to be passed to a descendant.
However,
the second phase deals with the special case where the new mutation has present
in a germ cell line that goes on to produce the egg cells, the oocytes. In this
phase a new mutation propagates and leads to heteroplasmy and homoplasmy in descendants.
Figure 2
illustrates the physical relationship between mtDNA molecules, a mitochondrial nucleoid, a mitochondrion and a cell.
Figure 2.
(a) mtDNA
molecules in a nucleoid, (b) Nucleoids
in a mitochondrion, (c) Mitochondria in a cell.
Initial
Mutational Events
Any mtDNA
mutation must in the first instance come from an initial mutational event
affecting a strand of mtDNA; and will usually involve either:
a
substitution of one base for another,
a
deletion of one or more neighbouring bases, or
the
insertion of one of more extra bases,
It is also
possible that an initial mutational event may, on occasions, be more
complex, and lead to several different changes in a
mtDNA molecule occurring at the same time.
For example, a deletion of bases at one point and their insertion at
another may occur.
Presumably, these initial mutational events
occur as errors when a mtDNA strand is being
replicated and are very common. But what
is unusual is for an initial event to lead to the development of a new homoplasmic mutation.
In this paper it is assumed that initial events
occur at random and do not exert any advantage, or disadvantage, to the
mitochondrion, cell, or person, in which they are found. In this respect, the discussion of any
possible positive or negative selection of individuals with certain
mtDNA mutations is beyond the scope of this paper.
However, a very interesting paper on selection
has recently been published by Doublet, et al.,(Doublet,
2008) and whilst the authors are discussing the mitochondria in a crustacean,
some of their conclusions on the general behaviour of mitochondria are similar
to those expressed in this paper.
Modelling initial mutational events
In the computer model the user is asked to select the
number of initial events from the range 1 - 10,000,000; and this number
then becomes the number of ‘runs’ that the model will use. In most instances an initial mutational event will not propagate, but each
mutational event is considered to have the same chance of successful
propagation as any other.
Propagation to a Mitochondrial Nucleoid
After its creation as an initial event, the
next stage in the successful propagation of a mutation is when, by random,
many, or all, of the mtDNA molecules in a particular mitochondrial nucleoid have the mutation.
In this paper this successful propagation is described as the capture
of a nucleoid by a mutation; and whereas initial
events are likely to be very common, a capture of a nucleoid is presumably very uncommon.
Unfortunately, the biology of mitochondrial nucleoids is poorly understood - but a nucleoid
appears to be a functionally separate area containing just a fraction of the
mtDNA molecules found within a single mitochondrion. The recent paper by Gilkerson,
et al. (2008) contains a detailed account of what is known about mitochondrial nucleoids, together with a very useful set of references.
An inference that can be drawn from the concept of the
capture of a mitochondrial nucleoid is that
different nucleoids in the same mitochondrion may
become captured by different mutations.
That is, the mtDNA molecules in one nucleoid
may contain one particular mutation whilst other nucleoids
may have other mutations
Phase A: Modelling the capture of a mitochondrial nucleoid
In the computer model the user is asked to determine
the Nucleoid Size from the range 5 -
30, and this value represents the average number of mtDNA molecules in
each nucleoid within a mitochondrion.
With the selections of initial events and Nucleoid Size made, the user can now click A:START to run the model.
The model will then for each initial event in
turn show if a new mutation can capture a nucleoid.
In simple terms the model replicates a mtDNA molecule chosen at random and then deletes a mtDNA
molecule, again chosen at random, over and over, until either the new mutation
is lost or it captures the nucleoid.
Propagation to a Mitochondrion
Once a new mutation has become found in most, or all, of
the mtDNA molecules in a nucleoid, the mutation may
then become dominant within a mitochondrion by replication occurring at random,
and without suggesting that there might be any selective advantage to the
‘mutated’ nucleoid.
It is not known how nucleoids
are replicated - but they do act as functionally separate organelles. As such it is likely that a nucleoid increases in size as its mtDNA molecules are
duplicated; and then the nucleoid divides into two
forming daughter nucleoids which each contain roughly
half the original number of mtDNA molecules.
This replication of nucleoids
appears to happen at random, and this may lead over a number of replication
cycles to the situation where all the nucleoids in a
mitochondrion are the descendants of just one nucleoid. So, if this nucleoid
does contain a new mutation in its
Phase A: Modelling the capture of a mitochondrion
In the computer model the user is asked to determine
the Nucleoid Number from
the range 5 - 20, and this value represents the average number of mtDNA
molecules in a mitochondrion. The user
also needs to specify ‘mitochondrion’ as opposed to ‘nucleoid’.
With the selections of initial events, Nucleoid Size, Nucleoid Number
and ‘mitochondrion’ made, the user can now click A:START
to run the model.
The model will then for each initial event in
turn show if a new mutation can capture a mitochondrion.
In simple terms the model will as before first try to capture
a nucleoid, and when successful, proceeds with this nucleoid to see if, at random, it captures the
mitochondrion.
These results suggest that with a Nucleoid
Number of 10 mtDNA molecules per
mitochondrion capture of a mitochondrion may occur, but as the Nucleoid Number is increased then mitochondrial capture
becomes very much less likely.
The Persistence of a Mutated Mitochondrion
Once a mitochondrion has most, or all, of its mtDNA
molecules containing a new mutation, it is possible that a fairly stable
situation has been reached. Mitochondria
are organelles that appear to have a long life within cells so it is likely
that a mutated mitochondrion once it has arisen may persist in tissues.
It is interesting to speculate that the phenomenon
whereby many new mutations appear in mtDNA taken from cancerous tissue that the
mutations have arisen because of the proliferation of persistent mutated
mitochondria in the tissue; and not as a result of the actual cancer causing
mutations in the mtDNA of its cells.
The Importance of the Germ Cell Line
The previous discussion has concerned itself with how
a new mutation might appear in any line of cells in an organism, but in
genealogy we are concerned with how mutations are transferred from a person in
one generation to their offspring in the next generation. And, for this to happen it is necessary to
consider that mutated mitochondria are present in the germ cell line—meaning
that for a new mutation to appear, mutated mitochondria need to be
present in the egg cells, the oocytes.
After fertilisation, a human oocyte
enlarges and divides repeatedly to form the first stage of embryonic growth,
the blastula. This has the form
of a hollow ball of cells; and each of the different cells of the blastula
goes on to divide and make a different cell line; and these cell lines in turn
form the different tissues of the body.
One of these lines in a female embryo is the cell line that form the primordial germ cells which over
time form the next set of oocytes; and
mitochondria containing mtDNA molecules with a new mutation must be present in
this cell line if the new mutation is to be taken forward to the next
generation.
Another important cell line to be considered is the
one which goes on to produce buccal cells (cells forming the inner
surface of the cheeks); as sequencing of the mitochondrial
But, as before whenever there is replication of cells
and their organelles, there is a random allocation of the organelles into the cells. So when different cell lines are produced
from the blastula it is possible that some cell lines may have a below
average number of mutated mitochondria in their cells and other cell
lines an above average number. If the
new primordial germ cell line has a higher proportion of mutated
mitochondria in its cells than were present in the germ cell line of
the parent then the new mutation has successfully
achieved a higher level of propagation.
Figure 3 illustrates
these stages in embryonic growth and shows how two cell lines may vary in the
number of mutated mitochondria that they contain in their cells.
Figure 3. New cell lines
containing different proportions of mutated mitochondria
Heteroplasmy and Homoplasmy
The above discussion has described how it is possible for
the cells in a particular cell line of the body to acquire a new mutation; and
how if the cell line is the Germ Cell Line that produces the egg cells,
the oocytes, for the next generation it is
possible for the proportion of mtDNA molecules with the new mutation to be
higher in the next, and subsequent generations.
At present it is not feasible to test cells from the Germ
Cell Line for Genealogical purposes, so it is usual to test buccal
cells which are very easy to collect using a mouth swab. A buccal cell sample is generally
assumed to represent the cells of the Germ Cell Line - but as the above
discussion has pointed out, there may be differences in the proportion of
mutated mtDNA molecules in different cell lines.
The mtDNA molecules obtained in a cell sample can
theoretically be considered as having heteroplasmy at very low levels,
but it is really only possible when sequencing for Genealogical purposes for heteroplasmy
to be detected when a new mutation is present in maybe a tenth of the mtDNA molecules,
i.e. at a level of, maybe, 10%. This
also means that homoplasmy is assumed on
testing when the new mutation might actually only be present in about 90% of
mtDNA molecules.
Just how a new mutation present at a low level of heteroplasmy
propagates to become a homoplasmic mutation is
not properly understood. But it is
likely that in many cases where a new mutation is being propagated successfully
that heteroplasmy levels rise slowly over a number of generations. This, however, does not mean that the level
of heteroplasmy rises with every generation, and indeed the level can be
expected to fall in some generations.
But whilst the gradual rise in levels of heteroplasmy
over a number of generations is possibly the usual way in which a new mutation
is propagated, this does not explain the observation that on occasions new
mutations can be seen to appear with a high level of heteroplasmy, or
even homoplasmy, in an individual, but
the mutation is not found in the individual’s mother.
However, one explanation of this phenomenon may be to
consider that at a particular stage in the development of Germ Cells in an embryo
there is such rapid growth that each
cell contains very few mitochondria.
And, if one of these mitochondria should contain a new mutation it is
possible that the mutation may appear to propagate suddenly over a single
generation. But, as discussed above,
this does not mean the initial mutational event is recent, as the first
phase of propagation leading to the presence of at least a
single mutated mitochondria will have needed to have occurred beforehand.
The recent papers by Shoubridge
and Wai (Shoubridge, 2007, Wai, 2008) discuss this phenomenon in some detail and
describe it as being a genetic bottleneck.
The Computer Model accompanying this paper attempts to
illustrate how a new mutation might propagate successfully from a very low
level of heteroplasmy to homoplasmy -
or, as happens in most cases, fails in its attempt to propagate. The model allows for the number of
mitochondria per cell to be reduced to very low levels, thereby simulating the
effect of a genetic bottleneck.
Phase B:
Modelling the
steps from Mitochondrial Capture to the appearance of Heteroplasmy and Homoplasmy
This part of the model shows how the level of heteroplasmy
might vary from one generation to the next; and whether homoplasmy
is achieved.
The starting condition can be considered as
representing the presence of a single mitochondrion with a new mutation in all
of its mtDNA molecules; and running the model shows over a series of Runs whether
the states of heteroplasmy and homoplasmy are
achieved.
In the computer model the user is asked to determine:
- the number of Runs from the range 10 - 1000,
- the Mitochondrial Number from the range 10 - 800.
In effect this number fixes the initial level of
heteroplasmy. Choosing ‘200’
mitochondria will give an initial level of 1/200, or 0.5%.
- the number of Replications from the range 5 - 20.
This number can be considered to represent the number of
cell divisions at the ‘blastula’ stage.
The higher the number selected the more randomisation will occur between
generations.
- the number of Generations form the range 10 - 300.
This number alters the number of generations that are
to be followed.
- there is also the option of suppressing the display of the Full
Results, which can be useful when the output from the model is large.
The model is run by choosing B:START. Results for three sets of example inputs are
shown below:
Conclusions
The above discussion and accompanying Computer Model
offer some ideas as to how mitochondrial mutations might arise and be
propagated.
First, a mutation appears as an initial mutational
event, before becoming the dominant form in the mitochondrial
But if a particular mutation cannot be detected on
sequencing as being heteroplasmic, does this mean the mutation is totally
absent, or is it present in some of the mtDNA molecules of an individual? The ideas discussed above do suggest that the
mitochondria of an individual may contain many new mutations, each of which has
a very small chance of propagating over future generations to be a heteroplasmic
or homoplasmic mutation.
However, other problems still remain to be solved:
For example, there are many examples in the
phylogenetic tree where a new mutation appears in branches near to each other,
but is absent in many other branches.
The usual explanation of this phenomenon has in the past involved the
concepts of parallel mutations and back mutations.
A parallel mutation being said to occur when
the same mutation has appeared in different places in the tree, i.e. the
mutation is said to have emerged by coincidence in different places. This phenomenon is generally considered
simply to be the result of the randomness of nature as there is only a limited number of possible mutations that can
occur and so it is likely the same mutation will occur by chance in different
places in the phylogenetic tree.
And, a back mutation is said to occur when a
new mutation is found higher up the tree, but is absent in lower branches where
it would be expected. This phenomenon is
difficult to explain as it would appear to depend on a mutation having a
memory of its previous state--something that is unlikely to occur.
However, this paper describes how a new mutation must
in the first instance appear as an initial mutational event and then may
propagate to be a heteroplasmic or homoplasmic
mutation. And, whereas the ideas
described here do not explain fully the finding of back mutations in the
phylogenetic tree, they do suggest a mechanism by which a new mutation may not
appear in all the descendants of an individual.
There is clearly still a great deal to be learnt about
the behaviour of mitochondrial mutations and better models will be produced,
but the present paper together with its Computer Model appears able to explain
many of the observed phenomena.
Supplementary Information
A listing of the code for the computer model is
available both in text form and as a web page at:
http://www.jogg.info/51/files/nucloid_model.txt
http://jogg/info/51/files/nucleoid_model.html
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