Divergence time estimates among European, Maghrebi, and Near Eastern ancestral populations.

<p>We use population-based F_st to estimate divergence time between each of the North African populations and the Qatari (green dots) and Tuscans (purple dots), respectively. We assume point estimates of effective population sizes based on autosomal haplotype heterozygosity estimates from Li et al. <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002397#pgen.1002397-Li1" target="_blank">[17]</a> (<i>Material and Methods</i>). We further estimate ancestral population clusters assigned at <i>k</i> = 5–8 with ADMIXTURE. Assuming that the ancestral clustering procedure has removed, or at least mitigated, the effect of recent migrations into Mediterranean populations, we then use F_st to estimate divergence times between these ancestral clusters. The range of <i>k</i>-based estimates for Maghrebi versus Near Eastern ancestry is indicated with light green polygon. The range of <i>k</i>-based estimates for Magrebi versus European ancestry is indicated with light purple polygon.</p

Genome admixture deconvolution karyogram of an Egyptian.

<p>A single Egyptian individual is presented for ancestry assuming <i>k</i> = 4 source populations: Saharawi [SAH], Nilotic-speaking Maasai [MKK], Spanish Basque [BAS] and Arabic Qatari [QAT]. Maasai segments (which were inferred from <i>k</i> = 3 and were highly diverged from the SAH, QAT, BAS segments) are layered on top of the inferred Maghrebi/Qatari/Basque ancestral karyogram, for <i>k</i> = 4 putative source populations.</p

Correlation between ancestry proportions inferred from ADMIXTURE and PCADMIX.

<p>We compare the proportions of ancestry inferred from assuming 3 ancestral populations in individuals from South Morocco A) using a clustering algorithm set to <i>k</i> = 8 (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002397#pgen-1002397-g001" target="_blank">Figure 1</a>) summing the sub-Saharan ancestry, both Qatar and European ancestry, and Maghrebi ancestry. We compared these estimates (left bar) to our PCA-based local ancestry assignment estimates (right bar). The three ancestral populations were Saharawi [SAH], Bantu-speaking Luhya [LWK], and Spanish Basque [BAS]. B) Genome admixture deconvolution on chromosome 1 of sixteen South Moroccans. Using a principal component-based method of admixture deconvolution, we assign local ancestry to South Moroccan individuals. We implement our PCA-based method for k = 3, and choose the ancestral populations based on the three ancestral populations were Saharawi [SAH], Bantu-speaking Luhya [LWK], and Spanish Basque [BAS]. Chromosome 1 for all sixteen South Moroccans is presented for both the maternal and paternal haplotypes.</p

Multidimensional scaling components discriminating genetic clusters in Africa.

<p>We used multidimensional scaling (MDS) to discriminate clusters of genetic variation within Africa and neighboring regions. MDS was applied to the pairwise, individual identity-by-state (IBS) matrix of 279,500 SNPs using PLINK 1.07 software <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002397#pgen.1002397-Purcell1" target="_blank">[45]</a>. The top three MDS components were plotted together using R 2.11.1. A) In axis one is plotted dimension 1 and in axis two is plotted dimension 2. B) In axis one is plotted dimension 1 and in axis two is plotted dimension 3. Population colors match <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002397#pgen-1002397-g001" target="_blank">Figure 1B</a> (<i>k</i> = 8), with the exception that the Fulani group was highlighted as a distinct population, indicated in light green. North African populations are all indicated in turquoise except for Tunisians that are shown in dark blue.</p

Map of samples and population structure of North Africa and neighboring populations.

<p>A) On the map, star symbols indicate the location of new population samples and circles indicate previously published samples. The decreasing proportions of Maghrebi ancestry is indicated in a west-to-east gradient of blue across North Africa. An unsupervised clustering algorithm, <i>ADMIXTURE </i><a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002397#pgen.1002397-Alexander1" target="_blank">[25]</a> was used to analyze population structure among 13 African, 2 European and 1 Near Eastern populations based on approximately 300 K autosomal SNP loci in common. The two main gradients of ancestry in North Africa (at <i>k</i> = 8), Maghrebi and Near Eastern, are emphasized with arrows. Other population colors in open circles match the colors displayed in the population structure analysis. B) We plot the full dataset assuming <i>k</i> = 2,4,6,8 ancestral populations. <i>k</i> = 10 and log likelihoods are presented in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002397#pgen.1002397.s001" target="_blank">Figure S1</a>.</p

Contribution of <i>FKBP5</i> Genetic Variation to Gemcitabine Treatment and Survival in Pancreatic Adenocarcinoma

<div><p>Purpose</p><p>FKBP51, (<i>FKBP5</i>), is a negative regulator of Akt. Variability in <i>FKBP5</i> expression level is a major factor contributing to variation in response to chemotherapeutic agents including gemcitabine, a first line treatment for pancreatic cancer. Genetic variation in <i>FKBP5</i> could influence its function and, ultimately, treatment response of pancreatic cancer.</p><p>Experimental Design</p><p>We set out to comprehensively study the role of genetic variation in <i>FKBP5</i> identified by Next Generation DNA resequencing on response to gemcitabine treatment of pancreatic cancer by utilizing both tumor and germline DNA samples from 43 pancreatic cancer patients, including 19 paired normal-tumor samples. Next, genotype-phenotype association studies were performed with overall survival as well as with <i>FKBP5</i> gene expression in tumor using the same samples in which resequencing had been performed, followed by functional genomics studies.</p><p>Results</p><p>In-depth resequencing identified 404 <i>FKBP5</i> single nucleotide polymorphisms (SNPs) in normal and tumor DNA. SNPs with the strongest associations with survival or <i>FKBP5</i> expression were subjected to functional genomic study. Electromobility shift assay showed that the rs73748206 “A(T)” SNP altered DNA-protein binding patterns, consistent with significantly increased reporter gene activity, possibly through its increased binding to Glucocorticoid Receptor (GR). The effect of rs73748206 was confirmed on the basis of its association with <i>FKBP5</i> expression by affecting the binding to GR in lymphoblastoid cell lines derived from the same patients for whom DNA was used for resequencing.</p><p>Conclusion</p><p>This comprehensive <i>FKBP5</i> resequencing study provides insights into the role of genetic variation in variation of gemcitabine response.</p></div

Models of migration into North Africa.

1<p>Segments from the migrant population were required to be greater than 3 cM in length. Given this minimum threshold, 815 migrant Egyptian and 1,275 migrant South Moroccan segments were discarded.</p>2<p>Two migration models were tested: a “pulse” model assumes a single episode of migration occurred at T₀ followed by no further migration, a “continuous” model assumes constant migration from T₀ to the present day. Log likelihoods given either model were compared and we present the model with the highest log likelihood.</p>3<p>The maximum likelihood estimate of time since migration initially began “T₀” from the migrant population into the admixed: population. We assume prior migration between the populations was zero. Time since migration began T₀ is indicated in generations.</p>4<p>The 95% confidence interval was estimated by sampling migrant Egyptian segments (n = 1,246) and migrant South Moroccan segments (n = 2,770) 1,000 times with replacement.</p

Distribution of long segments that are identical by descent (IBD) for pairs of individuals.

<p>The cumulative amount of DNA shared IBD between each pair of individuals within a population was summed. We display counts of the number of pairs sharing cumulative IBD in A) Tunisian Berbers, B) Saharawi and B) North Moroccans.</p

Functional characterization of the rs73748206 using pancreatic cancer patients’ lymphoblastoid cell lines (LCLs).

<p>(A) ChIP assay using lymphoblastoid cells with known genotype for the rs73748206 single nucleotide polymorphism (SNP). Left panel indicates a picture of gel electrophoresis. Lanes 1, 6, 9 correspond to DNA ladders. Lanes 2 to 5 are polymerase chain reaction (PCR) products from DNA that was bound to human GR (glucocorticoid receptor) antibody (GR) (lanes 2 and 3) or immunoglobulin G (IgG) control (lanes 4 and 5). The inputs for the variant (V) (lane 7) and wild type (WT) (lane 8) were PCR amplification products of pools of sheared DNA from the entire genome. Right panel is a ChIP qPCR analysis as % of input, shown as fold change relative to IgG. (B) SNP-related differences in relative FKBP5 and GR gene expression after exposure to dexamethasone for 24 h in variant (VAR) and WT cells; (*) p-value <i><</i>0.05. (C) SNP-related differences in <i>FKBP5</i> and <i>GR</i> expression (left and right panels, respectively), and gemcitabine response in variant (VAR), n = 1, and WT, n = 3, cell lines. Data is representative of three independent experiments (mean +/− SEM); (*) p-value <0.05.</p

SNP selection for functional genomic studies.

<p>(A) Association of patients’ survival (left panel) and <i>FKBP5</i> expression (right panel) with groups of <i>FKBP5</i> SNPs using Difference in Minor Allele Frequency test. These SNPs are grouped into 10–50 SNPs/group and are arranged based on their chromosome localization. Color indicates different length of SNP window. (B) Flow chart demonstrates the selection criteria of SNPs for the functional characterization studies. Abbreviations: SNP, single nucleotide polymorphism; ns cSNP, non-synonymous coding SNP; DMAF, Difference in Minor Allele Frequency.</p