Luísa Azevedo

An X-linked haplotype of Neandertal origin is present among all non-African populations

Recent work on the Neandertal genome has raised the possibility of admixture between Neandertals and the expanding population of Homo sapiens who left Africa between 80 and 50 Kya (thousand years ago) to colonize the rest of the world. Here, we provide evidence of a notable presence (9% overall) of a Neandertal-derived X chromosome segment among all contemporary human populations outside Africa. Our analysis of 6,092 X-chromosomes from all inhabited continents supports earlier contentions that a mosaic of lineages of different time depths and different geographic provenance could have contributed to the genetic constitution of modern humans. It indicates a very early admixture between expanding African migrants and Neandertals prior to or very early on the route of the out-of-Africa expansion that led to the successful colonization of the planet.

Frequency and pattern of heteroplasmy in the complete human mitochondrial genome.

Determining the levels of human mitochondrial heteroplasmy is of utmost importance in several fields. In spite of this, there are currently few published works that have focused on this issue. In order to increase the knowledge of mitochondrial DNA (mtDNA) heteroplasmy, the main goal of this work is to investigate the frequency and the mutational spectrum of heteroplasmy in the human mtDNA genome. To address this, a set of nine primer pairs designed to avoid co-amplification of nuclear DNA (nDNA) sequences of mitochondrial origin (NUMTs) was used to amplify the mitochondrial genome in 101 individuals. The analysed individuals represent a collection with a balanced representation of genders and mtDNA haplogroup distribution, similar to that of a Western European population. The results show that the frequency of heteroplasmic individuals exceeds 61%. The frequency of point heteroplasmy is 28.7%, with a widespread distribution across the entire mtDNA. In addition, an excess of transitions in heteroplasmy were detected, suggesting that genetic drift and/or selection may be acting to reduce its frequency at population level. In fact, heteroplasmy at highly stable positions might have a greater impact on the viability of mitochondria, suggesting that purifying selection must be operating to prevent their fixation within individuals. This study analyses the frequency of heteroplasmy in a healthy population, carrying out an evolutionary analysis of the detected changes and providing a new perspective with important consequences in medical, evolutionary and forensic fields.

Gains, losses and changes of function after gene duplication: study of the metallothionein family.

Metallothioneins (MT) are small proteins involved in heavy metal detoxification and protection against oxidative stress and cancer. The mammalian MT family originated through a series of duplication events which generated four major genes (MT1 to MT4). MT1 and MT2 encode for ubiquitous proteins, while MT3 and MT4 evolved to accomplish specific roles in brain and epithelium, respectively. Herein, phylogenetic, transcriptional and polymorphic analyses are carried out to expose gains, losses and diversification of functions that characterize the evolutionary history of the MT family. The phylogenetic analyses show that all four major genes originated through a single duplication event prior to the radiation of mammals. Further expansion of the MT1 gene has occurred in the primate lineage reaching in humans a total of 13 paralogs, five of which are pseudogenes. In humans, the reading frame of all five MT1 pseudogenes is reconstructed by sequence homology with a functional duplicate revealing that loss of invariant cysteines is the most frequent event accounting for pseudogeneisation. Expression analyses based on EST counts and RT-PCR experiments show that, as for MT1 and MT2, human MT3 is also ubiquitously expressed while MT4 transcripts are present in brain, testes, esophagus and mainly in thymus. Polymorphic variation reveals two deleterious mutations (Cys30Tyr and Arg31Trp) in MT4 with frequencies reaching about 30% in African and Asian populations suggesting the gene is inactive in some individuals and physiological compensation for its loss must arise from a functional equivalent. Altogether our findings provide novel data on the evolution and diversification of MT gene duplicates, a valuable resource for understanding the vast set of biological processes in which these proteins are involved.

Discussion on common data analysis strategies used in MS-based proteomics

Current proteomics technology is limited in resolving the proteome complexity of biological systems. The main issue at stake is to increase throughput and spectra quality so that spatiotemporal dimensions, population parameters and the complexity of protein modifications on a quantitative scale can be considered. MS‐based proteomics and protein arrays are the main players in large‐scale proteome analysis and an integration of these two methodologies is powerful but presently not sufficient for detailed quantitative and spatiotemporal proteome characterization. Improvements of instrumentation for MS‐based proteomics have been achieved recently resulting in data sets of approximately one million spectra which is a large step in the right direction. The corresponding raw data range from 50 to 100 Gb and are frequently made available. Multidimensional LC‐MS data sets have been demonstrated to identify and quantitate 2000–8000 proteins from whole cell extracts. The analysis of the resulting data sets requires several steps from raw data processing, to database‐dependent search, statistical evaluation of the search result, quantitative algorithms and statistical analysis of quantitative data. A large number of software tools have been proposed for the above‐mentioned tasks. However, it is not the aim of this review to cover all software tools, but rather discuss common data analysis strategies used by various algorithms for each of the above‐mentioned steps in a non‐redundant approach and to argue that there are still some areas which need improvements.

Ancestral origin of the ATTCT repeat expansion in spinocerebellar ataxia type 10 (SCA10)

Spinocerebellar ataxia type 10 (SCA10) is an autosomal dominant neurodegenerative disease characterized by cerebellar ataxia and seizures. The disease is caused by a large ATTCT repeat expansion in the ATXN10 gene. The first families reported with SCA10 were of Mexican origin, but the disease was soon after described in Brazilian families of mixed Portuguese and Amerindian ancestry. The origin of the SCA10 expansion and a possible founder effect that would account for its geographical distribution have been the source of speculation over the last years. To unravel the mutational origin and spread of the SCA10 expansion, we performed an extensive haplotype study, using closely linked STR markers and intragenic SNPs, in families from Brazil and Mexico. Our results showed (1) a shared disease haplotype for all Brazilian and one of the Mexican families, and (2) closely-related haplotypes for the additional SCA10 Mexican families; (3) little or null genetic distance in small normal alleles of different repeat sizes, from the same SNP lineage, indicating that they are being originated by a single step mechanism; and (4) a shared haplotype for pure and interrupted expanded alleles, pointing to a gene conversion model for its generation. In conclusion, we show evidence for an ancestral common origin for SCA10 in Latin America, which might have arisen in an ancestral Amerindian population and later have been spread into the mixed populations of Mexico and Brazil.

Epistatic interactions modulate the evolution of mammalian mitochondrial respiratory complex components.

BackgroundThe deleterious effect of a mutation can be reverted by a second-site interacting residue. This is an epistatic compensatory process explaining why mutations that are deleterious in some species are tolerated in phylogenetically related lineages, rendering evident that those mutations are, by all means, only deleterious in the species-specific context. Although an extensive and refined theoretical framework on compensatory evolution does exist, the supporting evidence remains limited, especially for protein models. In this current study, we focused on the molecular mechanism underlying the epistatic compensatory process in mammalian mitochondrial OXPHOS proteins using a combination of in-depth structural and sequence analyses.ResultsModeled human structures were used in this study to predict the structural impairment and recovery of deleterious mutations alone and combined with an interacting compensatory partner, respectively. In two cases, COI and COIII, intramolecular interactions between spatially linked residues restore the folding pattern impaired by the deleterious mutation. In a third case, intermolecular contact between mitochondrial CYB and nuclear CYT1 encoded components of the cytochrome bc1 complex are likely to restore protein binding. Moreover, we observed different modes of compensatory evolution that have resulted in either a quasi-simultaneous occurrence of a mutation and corresponding compensatory partner, or in independent occurrences of mutations in distinct lineages that were always preceded by the compensatory site.ConclusionEpistatic interactions between individual replacements involving deleterious mutations seems to follow a parsimonious model of evolution in which genomes hold pre-compensating states that subsequently tolerate deleterious mutations. This phenomenon is likely to have been constraining the variability at coevolving sites and shaping the interaction between the mitochondrial and the nuclear genome.

Novel TTC19 mutation in a family with severe psychiatric manifestations and complex III deficiency

Complex III of the mitochondrial respiratory chain (CIII) catalyzes transfer of electrons from reduced coenzyme Q to cytochrome c. Low biochemical activity of CIII is not a frequent etiology in disorders of oxidative metabolism and is genetically heterogeneous. Recently, mutations in the human tetratricopeptide 19 gene (TTC19) have been involved in the etiology of CIII deficiency through impaired assembly of the holocomplex. We investigated a consanguineous Portuguese family where four siblings had reduced enzymatic activity of CIII in muscle and harbored a novel homozygous mutation in TTC19. The clinical phenotype in the four sibs was consistent with severe olivo–ponto–cerebellar atrophy, although their age at onset differed slightly. Interestingly, three patients also presented progressive psychosis. The mutation resulted in almost complete absence of TTC19 protein, defective assembly of CIII in muscle, and enhanced production of reactive oxygen species in cultured skin fibroblasts. Our findings add to the array of mutations in TTC19, corroborate the notion of genotype/phenotype variability in mitochondrial encephalomyopathies even within a single family, and indicate that psychiatric manifestations are a further presentation of low CIII.

Molecular mechanisms underlying large genomic deletions in ornithine transcarbamylase (OTC) gene.

Ornithine transcarbamylase deficiency (OTCD) is an X‐linked urea cycle error causing hyperammonemia and orotic aciduria. Clinical diagnosis is generally confirmed by mutation detection. However, in ∼20% of the patients, no mutation is found by conventional mutation‐searching strategies, which fail to detect deletions spanning at least a whole exon, large rearrangements, or mutations at non‐coding regions. To detect large deletions or duplications, we have applied the multiplex ligation‐dependent probe amplification (MLPA) methodology to three OTCD patients (two females and one male). MLPA revealed copy number alterations of OTC exons in all of them. The two females were found to be heterozygous for deletions of either exon 2 or exons 6–9, and the male was confirmed to lack all OTC exons. Females’ characterization of the deletion breakpoints by long polymerase chain reaction and sequencing revealed the mutations c.78‐3544_217‐129del5921 and c.541‐600_1005 + 1880del10862 corresponding to exon 2 and exon 6–9 deletions, respectively. Examination of the deletion‐flanking regions suggests that exon 2 deletion probably resulted from replication slippage facilitated by a secondary structure formed by two inverted Alu repeats, whereas an Alu–Alu homologous recombination was probably responsible for the exon 6–9 deletion. This work contributes to the identification of novel disease‐causing mutations in OTCD and increases the knowledge on possible mutational mechanisms generating deletions in OTC.

Trans-species polymorphism in humans and the great apes is generally maintained by balancing selection that modulates the host immune response

SummaryKnown examples of ancient identical-by-descent genetic variants being shared between evolutionarily related species, known as trans-species polymorphisms (TSPs), result from counterbalancing selective forces acting on target genes to confer resistance against infectious agents. To date, putative TSPs between humans and other primate species have been identified for the highly polymorphic major histocompatibility complex (MHC), the histo-blood ABO group, two antiviral genes (ZC3HAV1 and TRIM5), an autoimmunity-related gene LAD1 and several non-coding genomic segments with a putative regulatory role. Although the number of well-characterized TSPs under long-term balancing selection is still very small, these examples are connected by a common thread, namely that they involve genes with key roles in the immune system and, in heterozygosity, appear to confer genetic resistance to pathogens. Here, we review known cases of shared polymorphism that appear to be under long-term balancing selection in humans and the great apes. Although the specific selective agent(s) responsible are still unknown, these TSPs may nevertheless be seen as constituting important adaptive events that have occurred during the evolution of the primate immune system.

NAMPT and NAPRT1 : novel polymorphisms and distribution of variants between normal tissues and tumor samples

Nicotinamide phosphoribosyltransferase (NAMPT) and nicotinate phosphoribosyltransferase domain containing 1 (NAPRT1) are the main human NAD salvage enzymes. NAD regulates energy metabolism and cell signaling, and the enzymes that control NAD availability are linked to pathologies such as cancer and neurodegeneration. Here, we have screened normal and tumor samples from different tissues and populations of origin for mutations in human NAMPT and NAPRT1, and evaluated their potential pathogenicity. We have identified several novel polymorphisms and showed that NAPRT1 has a greater genetic diversity than NAMPT, where any alteration can have a greater functional impact. Some variants presented different frequencies between normal and tumor samples that were most likely related to their population of origin. The novel mutations described that affect protein structure or expression levels can be functionally relevant and should be considered in a disease context. Particularly, mutations that decrease NAPRT1 expression can predict the usefulness of Nicotinic Acid in tumor treatments with NAMPT inhibitors.