Candida species are the most common cause of opportunistic fungal infection worldwide. We report the genome sequences of six Candida species and compare these and related pathogens and nonpathogens. There are significant expansions of cell wall, secreted, and transporter gene families in pathogenic species, suggesting adaptations associated with virulence. Large genomic tracts are homozygous in three diploid species, possibly resulting from recent recombination events. Surprisingly, key components of the mating and meiosis pathways are missing from several species. These include major differences at the Mating-type loci (MTL); Lodderomyces elongisporus lacks MTL, and components of the a1/alpha2 cell identity determinant were lost in other species, raising questions about how mating and cell types are controlled. Analysis of the CUG leucine to serine genetic code change reveals that 99% of ancestral CUG codons were erased and new ones arose elsewhere. Lastly, we revise the C. albicans gene catalog, identifying many new genes.

Human colorectal cancers (CRCs) display a large number of genetic and epigenetic alterations, some of which are causally involved in tumorigenesis (drivers) and others that have little functional impact (passengers). To help distinguish between these two classes of alterations, we used a transposonbased genetic screen in mice to identify candidate genes for CRC. Mice harboring mutagenic Sleeping Beauty (SB) transposons were crossed to mice expressing SB transposase in gastrointestinal tract epithelium. Most of the offspring developed intestinal lesions including intraepithelial neoplasia, adenomas, and adenocarcinomas. Analysis of over 16,000 transposon insertions identified 77 candidate CRC genes, 60 of which are mutated and/or dysregulated in human CRC and thus are most likely to drive tumorigenesis. These genes include APC, PTEN and SMAD4. The screen also * Publisher's Disclaimer: This manuscript has been accepted for publication in Science. This version has not undergone final editing.Please refer to the complete version of record at http://www.sciencemag.org/. Their manuscript may not be reproduced or used in any manner that does not fall within the fair use provisions of the

The pathways by which oncogenes, such as MLL-AF9, initiate transformation and leukemia in humans and mice are incompletely defined. In a study of target cells and oncogene dosage, we found that Mll-AF9, when under endogenous regulatory control, efficiently transformed LSK (Lin(-)Sca1(+)c-kit(+)) stem cells, while committed granulocyte-monocyte progenitors (GMPs) were transformation resistant and did not cause leukemia. Mll-AF9 was expressed at higher levels in hematopoietic stem (HSC) than GMP cells. Mll-AF9 gene dosage effects were directly shown in experiments where GMPs were efficiently transformed by the high dosage of Mll-AF9 resulting from retroviral transduction. Mll-AF9 upregulated expression of 192 genes in both LSK and progenitor cells, but to higher levels in LSKs than in committed myeloid progenitors.

Background: Osteoblast differentiation requires the coordinated stepwise expression of multiple genes. Histone deacetylase inhibitors (HDIs) accelerate the osteoblast differentiation process by blocking the activity of histone deacetylases (HDACs), which alter gene expression by modifying chromatin structure. We previously demonstrated that HDIs and HDAC3 shRNAs accelerate matrix mineralization and the expression of osteoblast maturation genes (e.g. alkaline phosphatase, osteocalcin). Identifying other genes that are differentially regulated by HDIs might identify new pathways that contribute to osteoblast differentiation.

Vascular smooth muscle cells (VSMCs) play important roles in cardiovascular disorders and biology. Outlined in this paper is a step-by-step procedure for isolating aortic VSMCs from adult C57BL6J male mice by enzymatic digestion of the aorta using collagenase. The plating, culturing, and subculturing of the isolated cells are discussed in detail along with techniques to characterize VSMC phenotype by gene expression and immunofluorescence. Traction force microscopy was used to characterize contractility of single subcultured VSMCs at baseline.

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IntroductionContinuous-flow left ventricular assist devices (LVADs) are an established therapy for patients with end-stage heart failure. The short- and long-term impact of these devices on peripheral blood gene expression has not been characterized, and may provide insight into the molecular pathways mediated in response to left ventricular remodeling and an improvement in overall systemic circulation. We performed RNA sequencing to identify genes and pathways influenced by these devices.MethodsRNA was extracted from blood of 9 heart failure patients (8 male) prior to LVAD implantation, and at 7 and 180 days postoperatively. Libraries were sequenced on an Illumina HiSeq2000 and sequences mapped to the human Ensembl GRCh37.67 genome assembly.ResultsA specific set of genes involved in regulating cellular immune response, antigen presentation, and T cell activation and survival were down-regulated 7 days after LVAD placement. 6 months following LVAD placement, the expression levels of these genes were significantly increased; yet importantly, remained significantly lower than age and sex-matched samples from healthy controls.ConclusionsIn summary, this genomic analysis identified a significant decrease in the expression of genes that promote a healthy immune response in patients with heart failure that was partially restored 6 months following LVAD implant.

In order to understand the pathophysiology of leukemia, we need to study the effects of leukemic oncogenes on the rare hematopoietic stem and progenitor cells. We investigated the self-renewal capabilities of the various hematopoietic cell types derived from Mll-AF9 knock-in mice. We used the murine knock-in model since it offers the advantage of a single copy of the Mll-fusion gene under the control of the endogenous promoter present in every hematopoietic stem/progenitor cell. In methylcellulose cultures, we compared myeloid colony formation of Mll-AF9 cells to wild type progenitor populations over three generations of plating. In the first generation of plating, the Mll-AF9 common myeloid progenitors (CMPs) and granulocyte-macrophage progenitors (GMPs) formed more colonies than the hematopoietic stem cells (HSCs) and common lymphoid progenitors (CLPs). However, at the third generation of plating, colony numbers formed by Mll-AF9 HSCs and CLPs were significantly greater than those formed by CMPs and GMPs. By the third generation only occasional colonies were found in the wild type groups. These results demonstrate that while Mll-AF9 led to an increase in self-renewal of all 4 cell types studied, these effects were more pronounced in HSCs and CLPs. To identify the downstream genes that mediate the growth deregulatory effects of Mll-AF9, we compared gene expression profiles of Mll-AF9 derived cells to their wild type counterparts. To assess gene expression levels, we extracted RNA from wild type and Mll-AF9 HSCs, CLPs, CMPs and GMPs. We then amplified and labeled the RNA for analysis by Affymetrix murine 430 2.0 genome arrays. In an unsupervised analysis, the various Mll-AF9 cells clustered with their corresponding wild type counterparts, indicating that the expression of most genes was not significantly altered by Mll-AF9. To identify the genes that are differentially expressed in the Mll-AF9 derived cells, we performed a two-way ANOVA (with the genotype and cell type as the two variables) allowing for a false discovery rate of 10%. In this analysis, we found that 76 genes were up-regulated in all Mll-AF9 progenitor cells compared to their wild-type counterparts. This list included known targets of Mll-fusion proteins Hoxa5, Hoxa7, Hoxa9 and Hoxa10. Also included were Evi1 and Mef2c, two genes that have been implicated in promoting enhanced self-renewal of murine hematopoietic cells. Importantly, in wild type mice, these 6 genes were expressed at higher levels in HSCs and CLPs compared to CMPs and GMPs (average 3–25 fold). While we observed an average 2–10 fold increase in expression of these genes in all Mll-AF9 cell types compared to their respective wild type controls, the expression level was 3–8 fold higher in Mll-AF9 HSCs and CLPs compared to CMPs and GMPs. Thus, the expression of genes known to be intrinsically related to self-renewal is further enhanced as a result of the Mll-AF9 fusion gene. In conclusion, while activation of the Mll-AF9 genetic program and the resulting enhanced self-renewal occurs in all 4 cell types studied, these effects are greatest in HSCs and CLPs. Thus, HSCs and CLPs are likely to be more efficient than CMPs and GMPs in producing cellular expansion and targets for cooperating mutations resulting in leukemia.