Phylogenomic analysis reveals extensive phylogenetic mosaicism in the Human GPCR Superfamily

A novel high throughput phylogenomic analysis (HTP) was applied to the rhodopsin G-protein coupled receptor (GPCR) family. Instances of phylogenetic mosaicism between receptors were found to be frequent, often as instances of correlated mosaicism and repeated mosaicism. A null data set was constructed with the same phylogenetic topology as the rhodopsin GPCRs. Comparison of the two data sets revealed that mosaicism was found in GPCRs in a higher frequency than would be expected by homoplasy or the effects of topology alone. Various evolutionary models of differential conservation, recombination and homoplasy are explored which could result in the patterns observed in this analysis. We find that the results are most consistent with frequent recombination events. A complex evolutionary history is illustrated in which it is likely frequent recombination has endowed GPCRs with new functions. The pattern of mosaicism is shown to be informative for functional prediction for orphan receptors. HTP analysis is complementary to conventional phylogenomic analyses revealing mosaicism that would not otherwise have been detectable through conventional phylogenetics

EM-mosaic detects mosaic point mutations that contribute to congenital heart disease.

BackgroundThe contribution of somatic mosaicism, or genetic mutations arising after oocyte fertilization, to congenital heart disease (CHD) is not well understood. Further, the relationship between mosaicism in blood and cardiovascular tissue has not been determined.MethodsWe developed a new computational method, EM-mosaic (Expectation-Maximization-based detection of mosaicism), to analyze mosaicism in exome sequences derived primarily from blood DNA of 2530 CHD proband-parent trios. To optimize this method, we measured mosaic detection power as a function of sequencing depth. In parallel, we analyzed our cohort using MosaicHunter, a Bayesian genotyping algorithm-based mosaic detection tool, and compared the two methods. The accuracy of these mosaic variant detection algorithms was assessed using an independent resequencing method. We then applied both methods to detect mosaicism in cardiac tissue-derived exome sequences of 66 participants for which matched blood and heart tissue was available.ResultsEM-mosaic detected 326 mosaic mutations in blood and/or cardiac tissue DNA. Of the 309 detected in blood DNA, 85/97 (88%) tested were independently confirmed, while 7/17 (41%) candidates of 17 detected in cardiac tissue were confirmed. MosaicHunter detected an additional 64 mosaics, of which 23/46 (50%) among 58 candidates from blood and 4/6 (67%) of 6 candidates from cardiac tissue confirmed. Twenty-five mosaic variants altered CHD-risk genes, affecting 1% of our cohort. Of these 25, 22/22 candidates tested were confirmed. Variants predicted as damaging had higher variant allele fraction than benign variants, suggesting a role in CHD. The estimated true frequency of mosaic variants above 10% mosaicism was 0.14/person in blood and 0.21/person in cardiac tissue. Analysis of 66 individuals with matched cardiac tissue available revealed both tissue-specific and shared mosaicism, with shared mosaics generally having higher allele fraction.ConclusionsWe estimate that ~ 1% of CHD probands have a mosaic variant detectable in blood that could contribute to cardiac malformations, particularly those damaging variants with relatively higher allele fraction. Although blood is a readily available DNA source, cardiac tissues analyzed contributed ~ 5% of somatic mosaic variants identified, indicating the value of tissue mosaicism analyses

Genetic Normalization of Differentiating Aneuploid Human Embryos

Early embryogenesis involves a series of dynamic processes, many of which are currently not well described or understood. Aneuploidy and aneuploid mosaicism, a mixture of aneuploid and euploid cells within one embryo, in early embryonic development are principal causes of developmental failure.^1,2^ Here we show that human embryos demonstrate a significant rate of genetic correction of aneuploidy, or "genetic normalization" when cultured from the cleavage stage on day 3 (Cleavage) to the blastocyst stage on day 5 (Blastocyst) using routine in vitro fertilization (IVF) laboratory conditions. One hundred and twenty-six human Cleavage stage embryos were evaluated for clinically indicated preimplantation genetic screening (PGS). Sixty-four of these embryos were found to be aneuploid following Cleavage stage embryo biopsy and single nucleotide polymorphism (SNP) 23 chromosome molecular karyotype (microarray). Of these, 25 survived to the Blastocyst stage of development and repeat microarray evaluation was performed. The inner cell mass (ICM), containing cells destined to form the fetus, and the trophectoderm (TE), containing cells destined to form the placenta were evaluated. Sixteen of 25 embryos (64%) [95% CI: 44-80%] possessed diploid karyotypes in both the ICM and TE cell populations. An additional three Blastocyst stage embryos showed genetic correction of the TE but not the ICM and one Blastocyst stage embryo showed the reverse. Mosaicism (exceeding 5%), was not detected in any of the ICM and TE samples analyzed. Recognizing that genetic normalization may occur in developing human embryos has important implications for stem cell biology, preimplantation and developmental genetics, embryology, and reproductive medicine. 

1)Hassold, T. et al. A cytogenetic study of 1000 spontaneous abortions. Ann Hum Genet. 44, 151-78 (1980).
2)Menasha, J., Levy, B., Hirschhorn, K., & Kardon, N.B. Incidence and spectrum of chromosome abnormalities in spontaneous abortions: new insights from a 12-year study. Genet Med. 7, 251-63 (2005)

X-chromosome inactivation mosaicism in the three germ layers and the germ line of the mouse embryo

Electrophoretic variant forms of the X-linked enzyme phosphoglycerate kinase (PGK-1, E.C.2, 7, 2, 3) have been used to examine X-chromosome mosaicism in tissues from 121/2-day post coitum heterozygous female mouse embryos. Samples of yolk-sac endoderm, neural ectoderm, heart (mesoderm), liver (endoderm) and germ cells were analysed from each embryo. In all tissues except yolk-sac endoderm, both PGK-1 isozymes were expressed. The extent of covariance among tissues with respect to the PGK-1 isozyme contribution is consistent with all tissues being derived from the same pool of cells after X-inactivation. The covariance among tissues gives an estimate of the size of this pool (47 cells) and places the earliest time of X-inactivation in epiblast cells between 41/2 and 51/2 days post coitum. From the independent variance among tissues within an individual, the average primordial precursor pool size for the three germ layers and the germ line itself was estimated as 193 cells

On the origin of trisomy 21 Down syndrome

Background: Down syndrome, characterized by an extra chromosome 21 is the most common genetic cause for congenital malformations and learning disability. It is well known that the extra chromosome 21 most often originates from the mother, the incidence increases with maternal age, there may be aberrant maternal chromosome 21 recombination and there is a higher recurrence in young women. In spite of intensive efforts to understand the underlying reason(s) for these characteristics, the origin still remains unknown. We hypothesize that maternal trisomy 21 ovarian mosaicism might provide the major causative factor. Results: We used fluorescence in situ hybridization (FISH) with two chromosome 21-specific probes to determine the copy number of chromosome 21 in ovarian cells from eight female foetuses at gestational age 14–22 weeks. All eight phenotypically normal female foetuses were found to be mosaics, containing ovarian cells with an extra chromosome 21. Trisomy 21 occurred with about the same frequency in cells that had entered meiosis as in pre-meiotic and ovarian mesenchymal stroma cells. Conclusion: We suggest that most normal female foetuses are trisomy 21 ovarian mosaics and the maternal age effect is caused by differential selection of these cells during foetal and postnatal development until ovulation. The exceptional occurrence of high-grade ovarian mosaicism may explain why some women have a child with Down syndrome already at young age as well as the associated increased incidence at subsequent conceptions. We also propose that our findings may explain the aberrant maternal recombination patterns previously found by family linkage analysis

Complex patterns of male germline instability and somatic mosaicism in myotonic dystrophy type 1

The genetic basis of myotonic dystrophy type 1 (DM1) is the expansion of a CTG repeat in the 3' untranslated region of DM1PK . Once into the disease range, the repeat becomes highly unstable and is biased toward expansion in both somatic and germline tissues. Intergenerational differences usually reveal an increase in allele length, concordant with the clinical anticipation characteristic of DM1, but there have also been cases with intergenerational contractions of the repeat length, accompanied by apparent anticipation. In order to gain a better understanding of this intergenerational behaviour, we have obtained semen samples from six DM males and used single molecule analyses to compare the allele distributions present in their sperm and blood with those of their offspring. We have confirmed that the male germline mutational pathway is distinct from that of the soma, but the extent of variation is highly variable from one individual to another and not obviously correlated with progenitor allele length. Nonetheless, in all cases the alleles present in the father's sperm overlap with those observed in their offspring. These data also provide further indications that the interpretation of intergenerational transmissions by standard analyses is frequently compromised by the masking of germline differences by age-dependent somatic expansion in the parent

Considering Intra-individual Genetic Heterogeneity to Understand Biodiversity

In this chapter, I am concerned with the concept of Intra-individual Genetic Hetereogeneity (IGH) and its potential influence on biodiversity estimates. Definitions of biological individuality are often indirectly dependent on genetic sampling -and vice versa. Genetic sampling typically focuses on a particular locus or set of loci, found in the the mitochondrial, chloroplast or nuclear genome. If ecological function or evolutionary individuality can be defined on the level of multiple divergent genomes, as I shall argue is the case in IGH, our current genetic sampling strategies and analytic approaches may miss out on relevant biodiversity. Now that more and more examples of IGH are available, it is becoming possible to investigate the positive and negative effects of IGH on the functioning and evolution of multicellular individuals more systematically. I consider some examples and argue that studying diversity through the lens of IGH facilitates thinking not in terms of units, but in terms of interactions between biological entities. This, in turn, enables a fresh take on the ecological and evolutionary significance of biological diversity