Inference of Ancestral Recombination Graphs through Topological Data Analysis

The recent explosion of genomic data has underscored the need for interpretable and comprehensive analyses that can capture complex phylogenetic relationships within and across species. Recombination, reassortment and horizontal gene transfer constitute examples of pervasive biological phenomena that cannot be captured by tree-like representations. Starting from hundreds of genomes, we are interested in the reconstruction of potential evolutionary histories leading to the observed data. Ancestral recombination graphs represent potential histories that explicitly accommodate recombination and mutation events across orthologous genomes. However, they are computationally costly to reconstruct, usually being infeasible for more than few tens of genomes. Recently, Topological Data Analysis (TDA) methods have been proposed as robust and scalable methods that can capture the genetic scale and frequency of recombination. We build upon previous TDA developments for detecting and quantifying recombination, and present a novel framework that can be applied to hundreds of genomes and can be interpreted in terms of minimal histories of mutation and recombination events, quantifying the scales and identifying the genomic locations of recombinations. We implement this framework in a software package, called TARGet, and apply it to several examples, including small migration between different populations, human recombination, and horizontal evolution in finches inhabiting the Gal\'apagos Islands.Comment: 33 pages, 12 figures. The accompanying software, instructions and example files used in the manuscript can be obtained from https://github.com/RabadanLab/TARGe

Isolation of a Chloramphenicol-Resistant Protein From λ-Infected Cells

During infection with bacteriophage ϕX174, a protein, made in the presence of 30 γ chloramphenicol/ml but not made in the presence of concentrations greater than 100 γ/ml, has been shown to be essential for viral DNA replication. A protein with similar chromatographic properties and “chloramphenicol resistance” has now been demonstrated in cells infected with λ bacteriophage

Germline Mutations and Polymorphisms in the Origins of Cancers in Women

Several female malignancies including breast, ovarian, and endometrial cancers can be characterized based on known somatic and germline mutations. Initiation and propagation of tumors reflect underlying genomic alterations such as mutations, polymorphisms, and copy number variations found in genes of multiple cellular pathways. The contributions of any single genetic variation or mutation in a population depend on its frequency and penetrance as well as tissue-specific functionality. Genome wide association studies, fluorescence in situ hybridization, comparative genomic hybridization, and candidate gene studies have enumerated genetic contributors to cancers in women. These include p53, BRCA1, BRCA2, STK11, PTEN, CHEK2, ATM, BRIP1, PALB2, FGFR2, TGFB1, MDM2, MDM4 as well as several other chromosomal loci. Based on the heterogeneity within a specific tumor type, a combination of genomic alterations defines the cancer subtype, biologic behavior, and in some cases, response to therapeutics. Consideration of tumor heterogeneity is therefore important in the critical analysis of gene associations in cancer

Identifying mRNA targets of microRNA dysregulated in cancer: with application to clear cell Renal Cell Carcinoma

BACKGROUND. MicroRNA regulate mRNA levels in a tissue specific way, either by inducing degradation of the transcript or by inhibiting translation or transcription. Putative mRNA targets of microRNA identified from seed sequence matches are available in many databases. However, such matches have a high false positive rate and cannot identify tissue specificity of regulation. RESULTS. We describe a simple method to identify direct mRNA targets of microRNA dysregulated in cancers from expression level measurements in patient matched tumor/normal samples. The word "direct" is used here in a strict sense to: a) represent mRNA which have an exact seed sequence match to the microRNA in their 3'UTR, b) the seed sequence match is strictly conserved across mouse, human, rat and dog genomes, c) the mRNA and microRNA expression levels can distinguish tumor from normal with high significance and d) the microRNA/mRNA expression levels are strongly and significantly anti-correlated in tumor and/or normal samples. We apply and validate the method using clear cell Renal Cell Carcinoma (ccRCC) and matched normal kidney samples, limiting our analysis to mRNA targets which undergo degradation of the mRNA transcript because of a perfect seed sequence match. Dysregulated microRNA and mRNA are first identified by comparing their expression levels in tumor vs normal samples. Putative dysregulated microRNA/mRNA pairs are identified from these using seed sequence matches, requiring that the seed sequence be conserved in human/dog/rat/mouse genomes. These are further pruned by requiring a strong anti-correlation signature in tumor and/or normal samples. The method revealed many new regulations in ccRCC. For instance, loss of miR-149, miR-200c and mir-141 causes gain of function of oncogenes (KCNMA1, LOX), VEGFA and SEMA6A respectively and increased levels of miR-142-3p, miR-185, mir-34a, miR-224, miR-21 cause loss of function of tumor suppressors LRRC2, PTPN13, SFRP1, ERBB4, and (SLC12A1, TCF21) respectively. We also found strong anti-correlation between VEGFA and the miR-200 family of microRNA: miR-200a*, 200b, 200c and miR-141. Several identified microRNA/mRNA pairs were validated on an independent set of matched ccRCC/normal samples. The regulation of SEMA6A by miR-141 was verified by a transfection assay. CONCLUSIONS. We describe a simple and reliable method to identify direct gene targets of microRNA in any cancer. The constraints we impose (strong dysregulation signature for microRNA and mRNA levels between tumor/normal samples, evolutionary conservation of seed sequence and strong anti-correlation of expression levels) remove spurious matches and identify a subset of robust, tissue specific, functional mRNA targets of dysregulated microRNA.Cancer Institute of New Jersy; New Jersey Commission for Cacner Research; Lineberger Comprehensive Cancer Center Tissue Procurement and Genomics Core Facility; Crawford Fun

Physical and Functional Interaction between p53 Mutants and Different Isoforms of p73

p53 is the most frequently inactivated tumor suppressor gene in human cancer, whereas its homologue, p73, is rarely mutated. Similarly to p53, p73 can promote growth arrest or apoptosis when overexpressed in certain p53-null tumor cells. It has previously been shown that some human tumor-derived p53 mutants can exert gain of function activity. The molecular mechanism underlying this activity remains to be elucidated. We show here that human tumor-derived p53 mutants (p53His175 and p53Gly281) associate in vitro and in vivo with p73 alpha, beta, gamma, and delta. This association occurs under physiological conditions, as verified in T47D and SKBR3 breast cancer cell lines. The core domain of mutant p53 is sufficient for the association with p73, whereas both the specific DNA binding and the oligomerization domains of p73 are required for the association with mutant p53. Furthermore, p53His175 and p53Gly281 mutants markedly reduce the transcriptional activity of the various isoforms of p73. Thus, human tumor-derived p53 mutants can associate with p73 not only physically but also functionally. These findings define a network involving mutant p53 and the various spliced isoforms of p73 that may confer upon tumor cells a selective survival advantage

A Double-Blind, Placebo-Controlled Withdrawal Trial of Dexmethylphenidate Hydrochloride in Children with Attention Deficit Hyperactivity Disorder

Objectives: d,l-threo-methylphenidate HCl (d,l-MPH) is the most common treatment of attention deficit hyperactivity disorder (ADHD). A previous report showed placebo-controlled efficacy for the purified disomer (dexmethylphenidate hydrochloride, d-MPH, Focalin™) with a 2:1 potency compared to dl, and suggested a 6-hour duration of action. This study complements that report by studying the effect of placebo-controlled discontinuation and retesting the duration of action. Methods: A 6-week, open-label titration of d-MPH (2.5–10 mg twice-a-day) was followed by a doubleblind, placebo-controlled, 2-week withdrawal study of responders. Results: In the open titration, 82% of the 89 enrolled patients achieved a Clinical Global Impression— Improvement (CGI-I) rating of much or very much improved. Only 5 patients discontinued for adverse events. Seventy-five patients continued into the placebo-controlled discontinuation. For the randomly assigned d-MPH (n = 35) and placebo (n = 40) groups, mean ages, respectively, were 10.1 ± 2.9 and 9.9 ± 2.7 years, 86% and 78% were male, and 70.6% and 80.0% took the ceiling dose of 10 mg twice-daily, respectively. Each group had 80% combined-type ADHD and 20% inattentive type. By the end of the 2- week, placebo-masked withdrawal, significantly more placebo patients (24 of 39) than d-MPH continuers (6 of 35) relapsed (61.5% versus 17.1%, p = 0.001). Compared to d-MPH continuers, placebo patients deteriorated significantly more in the 2-week period on teacher ratings of the 18 ADHD symptoms rated 0– 3 (p = 0.028), the 3 p.m. and 6 p.m. parent ADHD symptom ratings (p = 0.0026 and p = 0.0381, respectively), and clinic (2–3 p.m.) and home (6 p.m.) Math Tests (p = 0.024 and p < 0.0001, respectively). The 6 p.m. scores replicated the significant effect at 6 hours reported in the previous study. Conclusions: d-MPH is safe, tolerable, and effective, with a 6-hour duration of effect suggested by the significant difference from placebo at 6 hours on a double-blind discontinuation