Macrophage Ontogeny Underlies Differences in Tumor-Specific Education in Brain Malignancies.

Extensive transcriptional and ontogenetic diversity exists among normal tissue-resident macrophages, with unique transcriptional profiles endowing the cells with tissue-specific functions. However, it is unknown whether the origins of different macrophage populations affect their roles in malignancy. Given potential artifacts associated with irradiation-based lineage tracing, it remains unclear if bone-marrow-derived macrophages (BMDMs) are present in tumors of the brain, a tissue with no homeostatic involvement of BMDMs. Here, we employed multiple models of murine brain malignancy and genetic lineage tracing to demonstrate that BMDMs are abundant in primary and metastatic brain tumors. Our data indicate that distinct transcriptional networks in brain-resident microglia and recruited BMDMs are associated with tumor-mediated education yet are also influenced by chromatin landscapes established before tumor initiation. Furthermore, we demonstrate that microglia specifically repress Itga4 (CD49D), enabling its utility as a discriminatory marker between microglia and BMDMs in primary and metastatic disease in mouse and human

Integrating sequence and array data to create an improved 1000 Genomes Project haplotype reference panel

A major use of the 1000 Genomes Project (1000GP) data is genotype imputation in genome-wide association studies (GWAS). Here we develop a method to estimate haplotypes from low-coverage sequencing data that can take advantage of single-nucleotide polymorphism (SNP) microarray genotypes on the same samples. First the SNP array data are phased to build a backbone (or 'scaffold') of haplotypes across each chromosome. We then phase the sequence data 'onto' this haplotype scaffold. This approach can take advantage of relatedness between sequenced and non-sequenced samples to improve accuracy. We use this method to create a new 1000GP haplotype reference set for use by the human genetic community. Using a set of validation genotypes at SNP and bi-allelic indels we show that these haplotypes have lower genotype discordance and improved imputation performance into downstream GWAS samples, especially at low-frequency variants. © 2014 Macmillan Publishers Limited. All rights reserved

Phytochrome from green plants: assay, purification, and characterization

Phytochrome from the chlorophyllous cells of light-grown higher plants and green algae has been isolated and characterized. We have developed a simple procedure that separates chlorophyll from phytochrome in crude extracts from green tissue thus permitting spectral measurement of the phytochrome in such extracts for the first time. Spectral and immunochemical analysis of phytochrome from green oat tissue indicates the presence of two distinct species of the molecule: a minority species (approx. 20%) that is recognized by antibodies directed against phytochrome from etiolated tissue and that has an apparent molecular mass of 124 kilodaltons (kD), the same as that of the native molecule from etiolated tissue; and a majority species (approx. 80%) that is not recognized by anti-etiolated tissue phytochrome Ig and has a Pr absorbance maximum some 14 nm shorter than its etiolated tissue counterpart. Mixing experiments have established that these different molecular species preexist in the green cell and are not the results of posthomogenization modifications. Attempts to purify the phytochrome from green tissue by immunoaffinity chromatography have been thwarted by the lack of immunological cross-reactivity referred to. We have begun to identify monoclonal antibodies specific for antigenic sites distributed throughout the length of the etiolated-tissue phytochrome polypeptide. Axenic cultures of the alga Mesotaenium have been established and preliminary spectral analysis of phytochrome isolated from these cells has been carried out

Phytochrome-imposed oscillations in PIF3 protein abundance regulate hypocotyl growth under diurnal light/dark conditions in Arabidopsis

Arabidopsis seedlings display rhythmic growth when grown under diurnal conditions, with maximal elongation rates occurring at the end of the night under short-day photoperiods. Current evidence indicates that this behavior involves the action of the growth-promoting bHLH factors PHYTOCHROME-INTERACTING FACTOR 4 (PIF4) and PHYTOCHROME-INTERACTING FACTOR 5 (PIF5) at the end of the night, through a coincidence mechanism that combines their transcriptional regulation by the circadian clock with control of protein accumulation by light. To assess the possible role of PIF3 in this process, we have analyzed hypocotyl responses and marker gene expression in pif single- and higher-order mutants. The data show that PIF3 plays a prominent role as a promoter of seedling growth under diurnal light/dark conditions, in conjunction with PIF4 and PIF5. In addition, we provide evidence that PIF3 functions in this process through its intrinsic transcriptional regulatory activity, at least in part by directly targeting growth-related genes, and independently of its ability to regulate phytochrome B (phyB) levels. Furthermore, in sharp contrast to PIF4 and PIF5, our data show that the PIF3 gene is not subject to transcriptional regulation by the clock, but that PIF3 protein abundance oscillates under diurnal conditions as a result of a progressive decline in PIF3 protein degradation mediated by photoactivated phyB, and consequent accumulation of the bHLH factor during the dark period. Collectively, the data suggest that phyB-mediated, post-translational regulation allows PIF3 accumulation to peak just before dawn, at which time it accelerates hypocotyl growth, together with PIF4 and PIF5, by directly regulating the induction of growth-related genes