1,518 research outputs found
Genomic insights into fine-scale recombination variation in adaptively diverging threespine stickleback fish (Gasterosteus aculeatus)
Meiotic recombination is one of the major molecular mechanisms generating
genetic diversity and influencing genome evolution. By shuffling allelic
combinations, it can directly influence the patterns and efficacy of natural
selection. Studies in various organisms have shown that the rate and placement of
recombination varies substantially within the genome, among individuals,
between sexes and among different species. It is hypothesized that this variation
plays an important role in genome evolution. In this PhD thesis, I investigated the
extent and molecular basis of recombination variation in adaptively diverging
threespine stickleback fish (Gasterosteus aculeatus) to further understand its
evolutionary implications. I used both ChIP-sequencing and whole genome
sequencing of pedigrees to empirically identify and quantify double strand breaks
(DSBs) and meiotic crossovers (COs). Whole genome sequencing of large nuclear
families was performed to identify meiotic crossovers in 36 individuals of
diverging marine and freshwater ecotypes and their hybrids. This produced the
first genome-wide high-resolution sex-specific and ecotype-specific map of
contemporary recombination events in sticklebacks. The results show striking
differences in crossover number and placement between sexes. Females recombine
nearly 1.76 times more than males and their COs are distributed all over the
chromosome while male COs predominantly occur near the chromosomal
periphery. When compared among ecotypes a significant reduction in overall
recombination rate was observed in hybrid females compared to pure forms. Even
though the known loci underlying marine-freshwater adaptive divergence tend to
fall in regions of low recombination, considerable female recombination is
observed in the regions between adaptive loci. This suggests that the sexual
dimorphism in recombination phenotype may have important evolutionary
implications.
At the fine-scale, COs and male DSBs are nonrandomly distributed
involving ‘semi-hot’ hotspots and coldspots of recombination. I report a significant
association of male DSBs and COs with functionally active open chromatin regions
like gene promoters, whereas female COs did not show an association more than
expected by chance. However, a considerable number of COs and DSBs away from
any of the tested open chromatin marks suggests possibility of additional novel
mechanisms of recombination regulation in sticklebacks.
In addition, we developed a novel method for constructing individualized
recombination maps from pooled gamete DNA using linked read sequencing
technology by 10X Genomics®. We tested the method by contrasting recombination
profiles of gametic and somatic tissue from a hybrid mouse and stickleback fish.
Our pipeline faithfully detects previously described recombination hotspots in
mice at high resolution and identify many novel hotspots across the genome in
both species and thereby demonstrate the efficiency of the novel method. This
method could be employed for large scale QTL mapping studies to further
understand the genetic basis of recombination variation reported in this thesis.
By bridging the gap between natural populations and lab organisms with
large clutch sizes and tractable genetic tools, this work shows the utility of the
stickleback system and provides important groundwork for further studies of
heterochiasmy and divergence in recombination during adaptation to differing
environments
Integrative Analysis of Low- and High-Resolution eQTL
The study of expression quantitative trait loci (eQTL) is a powerful way of detecting transcriptional regulators at a genomic scale and for elucidating how natural genetic variation impacts gene expression. Power and genetic resolution are heavily affected by the study population: whereas recombinant inbred (RI) strains yield greater statistical power with low genetic resolution, using diverse inbred or outbred strains improves genetic resolution at the cost of lower power. In order to overcome the limitations of both individual approaches, we combine data from RI strains with genetically more diverse strains and analyze hippocampus eQTL data obtained from mouse RI strains (BXD) and from a panel of diverse inbred strains (Mouse Diversity Panel, MDP). We perform a systematic analysis of the consistency of eQTL independently obtained from these two populations and demonstrate that a significant fraction of eQTL can be replicated. Based on existing knowledge from pathway databases we assess different approaches for using the high-resolution MDP data for fine mapping BXD eQTL. Finally, we apply this framework to an eQTL hotspot on chromosome 1 (Qrr1), which has been implicated in a range of neurological traits. Here we present the first systematic examination of the consistency between eQTL obtained independently from the BXD and MDP populations. Our analysis of fine-mapping approaches is based on ‘real life’ data as opposed to simulated data and it allows us to propose a strategy for using MDP data to fine map BXD eQTL. Application of this framework to Qrr1 reveals that this eQTL hotspot is not caused by just one (or few) ‘master regulators’, but actually by a set of polymorphic genes specific to the central nervous system
Genome-Wide Control of the Distribution of Meiotic Recombination
Meiotic recombination events are not randomly distributed in the genome but occur in specific regions called recombination hotspots. Hotspots are predicted to be preferred sites for the initiation of meiotic recombination and their positions and activities are regulated by yet-unknown controls. The activity of the Psmb9 hotspot on mouse Chromosome 17 (Chr 17) varies according to genetic background. It is active in strains carrying a recombinant Chr 17 where the proximal third is derived from Mus musculus molossinus. We have identified the genetic locus required for Psmb9 activity, named Dsbc1 for Double-strand break control 1, and mapped this locus within a 6.7-Mb region on Chr 17. Based on cytological analysis of meiotic DNA double-strand breaks (DSB) and crossovers (COs), we show that Dsbc1 influences DSB and CO, not only at Psmb9, but in several other regions of Chr 17. We further show that CO distribution is also influenced by Dsbc1 on Chrs 15 and 18. Finally, we provide direct molecular evidence for the regulation in trans mediated by Dsbc1, by showing that it controls the CO activity at the Hlx1 hotspot on Chr 1. We thus propose that Dsbc1 encodes for a trans-acting factor involved in the specification of initiation sites of meiotic recombination genome wide in mice
Regulatory Hotspots in the Malaria Parasite Genome Dictate Transcriptional Variation
The determinants of transcriptional regulation in malaria parasites remain elusive. The presence of a well-characterized gene expression cascade shared by different Plasmodium falciparum strains could imply that transcriptional regulation and its natural variation do not contribute significantly to the evolution of parasite drug resistance. To clarify the role of transcriptional variation as a source of stain-specific diversity in the most deadly malaria species and to find genetic loci that dictate variations in gene expression, we examined genome-wide expression level polymorphisms (ELPs) in a genetic cross between phenotypically distinct parasite clones. Significant variation in gene expression is observed through direct co-hybridizations of RNA from different P. falciparum clones. Nearly 18% of genes were regulated by a significant expression quantitative trait locus. The genetic determinants of most of these ELPs resided in hotspots that are physically distant from their targets. The most prominent regulatory locus, influencing 269 transcripts, coincided with a Chromosome 5 amplification event carrying the drug resistance gene, pfmdr1, and 13 other genes. Drug selection pressure in the Dd2 parental clone lineage led not only to a copy number change in the pfmdr1 gene but also to an increased copy number of putative neighboring regulatory factors that, in turn, broadly influence the transcriptional network. Previously unrecognized transcriptional variation, controlled by polymorphic regulatory genes and possibly master regulators within large copy number variants, contributes to sweeping phenotypic evolution in drug-resistant malaria parasites
The Genomic Distribution of L1 Elements: The Role of Insertion Bias and Natural Selection
LINE-1 (L1) retrotransposons constitute the most successful family of retroelements in mammals and account for as much as 20% of mammalian DNA. L1 elements can be found in all genomic regions but they are far more abundant in AT-rich, gene-poor, and low-recombining regions of the genome. In addition, the sex chromosomes and some genes seem disproportionately enriched in L1 elements. Insertion bias and selective processes can both account for this biased distribution of L1 elements. L1 elements do not appear to insert randomly in the genome and this insertion bias can at least partially explain the genomic distribution of L1. The contrasted distribution of L1 and Alu elements suggests that postinsertional processes play a major role in shaping L1 distribution. The most likely mechanism is the loss of recently integrated L1 elements that are deleterious (negative selection) either because of disruption of gene function or their ability to mediate ectopic recombination. By comparison, the retention of L1 elements because of some positive effect is limited to a small fraction of the genome. Understanding the respective importance of insertion bias and selection will require a better knowledge of insertion mechanisms and the dynamics of L1 inserts in populations
Genesis of ancestral haplotypes: RNA modifications and reverse transcription–mediated polymorphisms
Understanding the genesis of the block haplotype structure of the genome is a major challenge. With the completion of the sequencing of the Human Genome and the initiation of the HapMap project the concept that the chromosomes of the mammalian genome are a mosaic, or patchwork, of conserved extended block haplotype sequences is now accepted by the mainstream genomics research community. Ancestral Haplotypes (AHs) can be viewed as a recombined string of smaller Polymorphic Frozen Blocks (PFBs). How have such variant extended DNA sequence tracts emerged in evolution? Here the relevant literature on the problem is reviewed from various fields of molecular and cell biology particularly molecular immunology and comparative and functional genomics. Based on our synthesis we then advance a testable molecular and cellular model. A critical part of the analysis concerns the origin of the strand biased mutation signatures in the transcribed regions of the human and higher primate genome, A-to-G versus T-to-C (ratio ~1.5 fold) and C-to-T versus G-to-A (≥1.5 fold). A comparison and evaluation of the current state of the fields of immunoglobulin Somatic Hypermutation (SHM) and Transcription-Coupled DNA Repair focused on how mutations in newly synthesized RNA might be copied back to DNA thus accounting for some of the genome-wide strand biases (e.g., the A-to-G vs T-to-C component of the strand biased spectrum). We hypothesize that the genesis of PFBs and extended AHs occurs during mutagenic episodes in evolution (e.g., retroviral infections) and that many of the critical DNA sequence diversifying events occur first at the RNA level, e.g., recombination between RNA strings resulting in tandem and dispersed RNA duplications (retroduplications), RNA mutations via adenosine-to-inosine pre-mRNA editing events as well as error prone RNA synthesis. These are then copied back into DNA by a cellular reverse transcription process (also likely to be error-prone) that we have called "reverse transcription-mediated long DNA conversion." Finally we suggest that all these activities and others can be envisaged as being brought physically under the umbrella of special sites in the nucleus involved in transcription known as "transcription factories."
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