The human mitochondrial molecular clock is the rate at which mutations have been accumulating in the mitochondrial genome of hominids during the course of human evolution. The archeological record of human activity from early periods in human prehistory is relatively limited and its interpretation has been controversial. Because of the uncertainties from the archeological record, scientists have turned to molecular dating techniques in order to refine the timeline of human evolution. A major goal of scientists in the field is to develop an accurate hominid mitochondrial molecular clock which could then be used to confidently date events that occurred during the course of human evolution.
Estimates of the mutation rate of human mitochondrial DNA (mtDNA) vary greatly depending on the available data and the method used for estimation. The two main methods of estimation, phylogeny-based methods and pedigree-based methods, have produced mutation rates that differ by almost an order of magnitude. Current research has been focused on resolving the high variability obtained from different rate estimates.
Rate variability A major assumption of the molecular clock theory is that mutations within a particular genetic system occur at a statistically uniform rate and this uniform rate can be used for dating genetic events. In practice the assumption of a single uniform rate is an oversimplification. Though a single mutation rate is often applied, it is often a composite or an average of several different mutation rates. Many factors influence observed mutation rates and these factors include the type of samples, the region of the genome studied and the time period covered.
Actual vs. observed rates The rate at which mutations occur during reproduction, the germline mutation rate, is thought to be higher than all observed mutation rates, because not all mutations are successfully passed down to subsequent generations. mtDNA is only passed down along the matrilineal line, and therefore mutations passed down to sons are lost. Random genetic drift may also cause the loss of mutations. For these reasons, the actual mutation rate will not be equivalent to the mutation rate observed from a population sample.
Population size Population dynamics are believed to influence observed mutation rates. When a population is expanding, more germline mutations are preserved in the population. As a result, observed mutation rates tend to increase in an expanding population. When populations contract, as in a population bottleneck, more germline mutations are lost. Population bottlenecks thus tend to slow down observed mutation rates. Since the emergence of the species homo sapiens about 200,000 years ago, the human population has expanded from a few thousand individuals living in Africa to over 8 billion worldwide. However, the expansion has not been uniform, so the history of human populations may consist of both bottlenecks and expansions.
Structural variability The mutation rate across the mitochondrial genome is not uniformly distributed. Certain regions of the genome are known to mutate more rapidly than others. The Hypervariable regions are known to be highly polymorphic relative to other parts of the genome.
The rate at which mutations accumulate in coding and non-coding regions of the genome also differs as mutations in the coding region are subject to purifying selection. For this reason, some studies avoid coding region or nonsynonymous mutations when calibrating the molecular clock. only consider synonymous mutations, they have recalibrated the molecular clock of human mtDNA as 7990 years per synonymous mutation over the mitochondrial genome. consider both coding and non-coding region mutations to arrive at a single mutation rate, but apply a correction factor to account for selection in the coding region.
Temporal variability The mutation rate has been observed to vary with time. Mutation rates within the human species are faster than those observed along the human-ape lineage. The mutation rate is also thought to be faster in recent times, since the beginning of the Holocene 11,000 years ago.
Parallel mutations and saturation Parallel mutation (sometimes referred to as Homoplasy) or convergent evolution occurs when separate lineages have the same mutation independently occur at the same site in the genome. Saturation occurs when a single site experiences multiple mutations. Parallel mutations and saturation result in the underestimation of the mutation rate because they are likely to be overlooked.
Heteroplasmy Individuals affected by heteroplasmy have a mixture of mtDNA types, some with new mutations and some without. The new mutations may or may not be passed down to subsequent generations. Thus the presence of heteroplasmic individuals in a sample may complicate the calculation of mutation rates.
Methods
Pedigree based Pedigree methods estimate the mutation rate by comparing the mtDNA sequences of a sample of parent/offspring pairs or analyzing mtDNA sequences of individuals from a deep-rooted genealogy. The number of new mutations in the sample is counted and divided by the total number of parent-to-child DNA transmission events to arrive at a mutation rate.
Phylogeny based Phylogeny based methods are estimated by first reconstructing the haplotype of the most recent common ancestor (MRCA) of a sample of two or more genetic lineages. A requirement is that the time to the most recent common ancestor (TMRCA) of the sample of lineages must already be known from other independent sources, usually the archeological record. The average number of mutations that have accumulated since the MRCA is then computed and divided by the TMRCA to arrive at the mutation rate. The human mutation rate is usually estimated by comparing the sequences of modern humans and chimpanzees and then reconstructing the ancestral haplotype of the chimpanzee-human common ancestor. According to the paleontological record the last common ancestor of humans may have lived around 6 million years ago.
Pedigree vs. phylogeny comparison Rates obtained by pedigree methods are about 10 times faster than those obtained by phylogenetic methods. Several factors acting together may be responsible for this difference. As pedigree methods record mutations in living subjects, the mutation rates from pedigree studies are closer to the germline mutation rate. Pedigree studies use genealogies that are only a few generations deep whereas phylogeny based methods use timescales that are thousands or millions of years deep. According to Henn et al. 2009, phylogeny based methods take into account events that occur over long time scales and are thus less affected by stochastic fluctuations. Howell et al. 2003 suggests that selection, saturation, parallel mutations and genetic drift are responsible for the differences observed between pedigree based methods and phylogeny based methods.
Estimating based on AMH archaeology Methods/parameters for archaeologically estimated dates of mitochondrial Eve Study Sequencetype TAnchor(location) Referencing method(correction method) Restriction fragments 40, 30, and 12 Ka(Australia, New Guinea New World) archaeologically defined migrations matched with estimated sequence divergence rates Genomic 40 to 55 Ka(Papua New Guinea)14.5 to 21.5 Ka(Haps H1 and H3) PNG following Haplogroup P Anatomically modern humans (AMH) spread out of Africa and over a large area of Eurasia and left artifacts along the northern coast of the Southwest, South, Southeast and East Asia. did not rely on a predicted TCHLCA to estimate single-nucleotide polymorphism (SNP) rates. Instead, they used evidence of colonization in Southeast Asia and Oceania to estimate mutation rates. In addition they used RFLP technology (Restriction fragment length polymorphism) to examine differences between DNA. Using these techniques this group came up with a TMRCA of 140,000 to 290,000 years. Cann et al. (1987) estimated the TMRCA of humans to be approximately 210 ky and the most recent estimates Soares et al. 2009 (using 7 million year chimpanzee human mtDNA MRCA) differ by only 9%, which is relatively close considering the wide confidence range for both estimates and calls for more ancient TCHLCA.
have reevaluated the predicted migrations globally and compared those to the actual evidence. This group used the coding regions of sequences. They postulate that the molecular clock based on chimp-human comparisons is not reliable, particularly in predicting recent migrations, such as founding migrations into Europe, Australia, and the Americans. With this technique this group came up with a TMRCA of 82,000 to 134,000 years.
Estimating based on CHLCA Because chimps and humans share a matrilineal ancestor, establishing the geological age of that last ancestor allows the estimation of the mutation rate. The chimp-human last common ancestor (CHLCA) is frequently applied as an anchor for mt-TMRCA studies with ranges between 4 and 13 million years cited in the literature. This is one source of variation in the time estimates. The other weakness is the non-clocklike accumulation of SNPs, would tend to make more recent branches look older than they actually are.see:
+ SNP rates as described by Soares et al. (2009) Regions(s) Subregions(or site within codon) SNP rate(per site * year) Controlregion HVR I 1.6 × 10−7 HVR II 2.3 × 10−7 remaining 1.5 × 10−8 Protein-coding (1st and 2nd) 8.8 × 10−9 (3rd) 1.9 × 10−8 DNA encoding rRNA (rDNA) 8.2 × 10−9 DNA encoding tRNA (tDNA) 6.9 × 10−9 other 2.4 × 10−8 TCHLCA assumed 6.5 Ma, relative rate to 1st & 2nd codons These two sources may balance each other or amplify each other depending on the direction of the TCHLCA error. There are two major reasons why this method is widely employed. First the pedigree based rates are inappropriate for estimates for very long periods of time. Second, while the archaeology anchored rates represent the intermediate range, archaeological evidence for human colonization often occurs well after colonization. For example, colonization of Eurasia from west to east is believed to have occurred along the Indian Ocean. However, the oldest archaeological sites that also demonstrate anatomically modern humans (AMH) are in China and Australia, greater than 42,000 years in age. However the oldest Indian site with AMH remains is from 34,000 years, and another site with AMH compatible archaeology is in excess of 76,000 years in age. Therefore, application of the anchor is a subjective interpretation of when humans were first present.
A simple measure the sequence divergence between humans and chimps can be bound by observing the SNPs. Given that the mitogenome is about 16553 base pairs in length (each base-pair which can be aligned with known references is called a site), the formula is:
The '2' in the denominator is derived from the 2 lineages, human and chimpanzee, that split from the CHLCA. Ideally it represents the accumulation of mutations on both lineages but in different positions (SNPs). As long as the number of SNP observed approximates the number of mutations this formula works well. However, at rapidly evolving sites mutations are obscured by saturation affects. Sorting positions within the mitogenome by rate and compensating for saturation are alternative approaches.See: ,
Because the TCHLCA is subject to change with more paleontological information, the equation described above allows the comparison of TMRCA from different studies.