Proper Sterol Distribution Is Required for Candida albicans Hyphal Formation and Virulence

Candida albicans is an opportunistic fungus responsible for the majority of systemic fungal infections. Multiple factors contribute to C. albicans pathogenicity. C. albicans strains lacking CaArv1 are avirulent. Arv1 has a conserved Arv1 homology domain (AHD) that has a zinc-binding domain containing two cysteine clusters. Here, we explored the role of the CaAHD and zinc-binding motif in CaArv1-dependent virulence. Overall, we found that the CaAHD was necessary but not sufficient for cells to be virulent, whereas the zinc-binding domain was essential, as Caarv1/Caarv1 cells expressing the full-length zinc-binding domain mutants, Caarv1C3S and Caarv1C28S, were avirulent. Phenotypically, we found a direct correlation between the avirulence of Caarv1/Caarv1, Caarrv1AHD, Caarv1C3S, and Caarv1C28S cells and defects in bud site selection, septa formation and localization, and hyphal formation and elongation. Importantly, all avirulent mutant strains lacked the ability to maintain proper sterol distribution. Overall, our results have established the importance of the AHD and zinc-binding domain in fungal invasion, and have correlated an avirulent phenotype with the inability to maintain proper sterol distribution

Properties of the Youngest Protostars in Perseus, Serpens, and Ophiuchus

We present an unbiased census of deeply embedded protostars in Perseus, Serpens, and Ophiuchus, assembled by combining large-scale 1.1 mm Bolocam continuum and Spitzer Legacy surveys. We identify protostellar candidates based on their mid-infrared properties, correlate their positions with 1.1 mm core positions, and construct well-sampled SEDs using our extensive wavelength coverage (lam=1.25-1100 micron). Source classification based on the bolometric temperature yields a total of 39 Class 0 and 89 Class I sources in the three cloud sample. We compare to protostellar evolutionary models using the bolometric temperature-luminosity diagram, finding a population of low luminosity Class I sources that are inconsistent with constant or monotonically decreasing mass accretion rates. This result argues strongly for episodic accretion during the Class I phase, with more than 50% of sources in a ``sub-Shu'' (dM/dt < 1e-6 Msun/yr) accretion state. Average spectra are compared to protostellar radiative transfer models, which match the observed spectra fairly well in Stage 0, but predict too much near-IR and too little mid-IR flux in Stage I. Finally, the relative number of Class 0 and Class I sources are used to estimate the lifetime of the Class 0 phase; the three cloud average yields a Class 0 lifetime of 1.7e5 yr, ruling out an extremely rapid early accretion phase. Correcting photometry for extinction results in a somewhat shorter lifetime (1.1e5 yr). In Ophiuchus, however, we find very few Class 0 sources (N(Class0)/N(ClassI)=0.1-0.2), similar to previous studies of that cloud. The observations suggest a consistent picture of nearly constant average accretion rate through the entire embedded phase, with accretion becoming episodic by at least the Class I stage, and possibly earlier.Comment: 31 pages, 19 figures, 8 tables; accepted for publication in Ap

The Mass Distribution and Lifetime of Prestellar Cores in Perseus, Serpens, and Ophiuchus

We present an unbiased census of starless cores in Perseus, Serpens, and Ophiuchus, assembled by comparing large-scale Bolocam 1.1 mm continuum emission maps with Spitzer c2d surveys. We use the c2d catalogs to separate 108 starless from 92 protostellar cores in the 1.1 mm core samples from Enoch et al. (2006), Young et al. (2006), and Enoch et al. (2007). A comparison of these populations reveals the initial conditions of the starless cores. Starless cores in Perseus have similar masses but larger sizes and lower densities on average than protostellar cores, with sizes that suggest density profiles substantially flatter than r^-2. By contrast, starless cores in Serpens are compact and have lower masses than protostellar cores; future star formation will likely result in lower mass objects than the currently forming protostars. Comparison to dynamical masses estimated from the NH_3 survey of Perseus cores by Rosolowsky et al. (2007) suggests that most of the starless cores are likely to be gravitationally bound, and thus prestellar. The combined prestellar core mass distribution includes 108 cores and has a slope of -2.3+/-0.4 for M>0.8 Msun. This slope is consistent with recent measurements of the stellar initial mass function, providing further evidence that stellar masses are directly linked to the core formation process. We place a lower limit on the core-to-star efficiency of 25%. There are approximately equal numbers of prestellar and protostellar cores in each cloud, thus the dense prestellar core lifetime must be similar to the lifetime of embedded protostars, or 4.5x10^5 years, with a total uncertainty of a factor of two. Such a short lifetime suggests a dynamic, rather than quasi-static, core evolution scenario, at least at the relatively high mean densities (n>2x10^4 cm^-3) to which we are sensitive.Comment: 27 pages, 15 figures, 5 tables, accepted for publication in ApJ. Version with full resolution figures available at http://www.astro.caltech.edu/~menoch/corespaper

Analysis of the astray/robo2 Zebrafish Mutant Reveals that Degenerating Tracts Do Not Provide Strong Guidance Cues for Regenerating Optic Axons

During formation of the optic projection in astray/robo2 mutant zebrafish, optic axons exhibit rostro-caudal pathfinding errors, ectopic midline crossing and increased terminal arbor size. Here we show that these errors persist into adulthood, even when robo2 function is conditionally reduced only during initial formation of the optic projection. Adult errors include massive ectopic optic tracts in the telencephalon. During optic nerve regeneration in astray/robo2 animals, these tracts are not re-populated and ectopic midline crossing is reduced compared to unlesioned mutants. This is despite a comparable macrophage/microglial response and upregulation of contactin1a in oligodendrocytes of entopic and ectopic tracts. However, other errors, such as expanded termination areas and ectopic growth into the tectum, were frequently re-committed by regenerating optic axons. Retinal ganglion cells with regenerating axons re-express robo2 and expression of slit ligands is maintained in some areas of the adult optic pathway. However, slit expression is reduced rostral and caudal to the chiasm, compared to development and ubiquitous overexpression of Slit2 did not elicit major pathfinding phenotypes. This shows that (1) there is not an efficient correction mechanism for large-scale pathfinding errors of optic axons during development; (2) degenerating tracts do not provide a strong guidance cue for regenerating optic axons in the adult CNS, unlike the PNS; and (3) robo2 is less important for pathfinding of optic axons during regeneration than during development

Drug discovery for Diamond-Blackfan anemia using reprogrammed hematopoietic progenitors

Diamond-Blackfan anemia (DBA) is a congenital disorder characterized by the failure of erythroid progenitor differentiation, severely curtailing red blood cell production. Because many DBA patients fail to respond to corticosteroid therapy, there is considerable need for therapeutics for this disorder. Identifying therapeutics for DBA requires circumventing the paucity of primary patient blood stem and progenitor cells. To this end, we adopted a reprogramming strategy to generate expandable hematopoietic progenitor cells from induced pluripotent stem cells (iPSCs) from DBA patients. Reprogrammed DBA progenitors recapitulate defects in erythroid differentiation, which were rescued by gene complementation. Unbiased chemical screens identified SMER28, a small-molecule inducer of autophagy, which enhanced erythropoiesis in a range of in vitro and in vivo models of DBA. SMER28 acted through autophagy factor ATG5 to stimulate erythropoiesis and up-regulate expression of globin genes. These findings present an unbiased drug screen for hematological disease using iPSCs and identify autophagy as a therapeutic pathway in DBA.National Institute of Diabetes and Digestive and Kidney Diseases (U.S.) (Grant R24-DK092760)National Institute of Diabetes and Digestive and Kidney Diseases (U.S.) (Grant R24-DK49216)National Institute of Diabetes and Digestive and Kidney Diseases (U.S.) (Grant U54DK110805)National Heart, Lung, and Blood Institute (Grant UO1-HL100001)National Heart, Lung, and Blood Institute (Grant U01HL134812)National Heart, Lung, and Blood Institute (Grant R01HL04880)National Institutes of Health (U.S.) (Grant R24OD017870-01

Discovery and Validation of Molecular Biomarkers for Colorectal Adenomas and Cancer with Application to Blood Testing

BACKGROUND & AIMS: Colorectal cancer incidence and deaths are reduced by the detection and removal of early-stage, treatable neoplasia but we lack proven biomarkers sensitive for both cancer and pre-invasive adenomas. The aims of this study were to determine if adenomas and cancers exhibit characteristic patterns of biomarker expression and to explore whether a tissue-discovered (and validated) biomarker is differentially expressed in the plasma of patients with colorectal adenomas or cancer. METHODS: Candidate RNA biomarkers were identified by oligonucleotide microarray analysis of colorectal specimens (222 normal, 29 adenoma, 161 adenocarcinoma and 50 colitis) and validated in a previously untested cohort of 68 colorectal specimens using a custom-designed oligonucleotide microarray. One validated biomarker, KIAA1199, was assayed using qRT-PCR on plasma extracted RNA from 20 colonoscopy-confirmed healthy controls, 20 patients with adenoma, and 20 with cancer. RESULTS: Genome-wide analysis uncovered reproducible gene expression signatures for both adenomas and cancers compared to controls. 386/489 (79%) of the adenoma and 439/529 (83%) of the adenocarcinoma biomarkers were validated in independent tissues. We also identified genes differentially expressed in adenomas compared to cancer. KIAA1199 was selected for further analysis based on consistent up-regulation in neoplasia, previous studies and its interest as an uncharacterized gene. Plasma KIAA1199 RNA levels were significantly higher in patients with either cancer or adenoma (31/40) compared to neoplasia-free controls (6/20). CONCLUSIONS: Colorectal neoplasia exhibits characteristic patterns of gene expression. KIAA1199 is differentially expressed in neoplastic tissues and KIAA1199 transcripts are more abundant in the plasma of patients with either cancer or adenoma compared to controls

Latency Associated Peptide Has In Vitro and In Vivo Immune Effects Independent of TGF-β1

Latency Associated Peptide (LAP) binds TGF-β1, forming a latent complex. Currently, LAP is presumed to function only as a sequestering agent for active TGF-β1. Previous work shows that LAP can induce epithelial cell migration, but effects on leukocytes have not been reported. Because of the multiplicity of immunologic processes in which TGF-β1 plays a role, we hypothesized that LAP could function independently to modulate immune responses. In separate experiments we found that LAP promoted chemotaxis of human monocytes and blocked inflammation in vivo in a murine model of the delayed-type hypersensitivity response (DTHR). These effects did not involve TGF-β1 activity. Further studies revealed that disruption of specific LAP-thrombospondin-1 (TSP-1) interactions prevented LAP-induced responses. The effect of LAP on DTH inhibition depended on IL-10. These data support a novel role for LAP in regulating monocyte trafficking and immune modulation

Self-Mating in the Definitive Host Potentiates Clonal Outbreaks of the Apicomplexan Parasites Sarcocystis neurona and Toxoplasma gondii

Tissue-encysting coccidia, including Toxoplasma gondii and Sarcocystis neurona, are heterogamous parasites with sexual and asexual life stages in definitive and intermediate hosts, respectively. During its sexual life stage, T. gondii reproduces either by genetic out-crossing or via clonal amplification of a single strain through self-mating. Out-crossing has been experimentally verified as a potent mechanism capable of producing offspring possessing a range of adaptive and virulence potentials. In contrast, selfing and other life history traits, such as asexual expansion of tissue-cysts by oral transmission among intermediate hosts, have been proposed to explain the genetic basis for the clonal population structure of T. gondii. In this study, we investigated the contributing roles self-mating and sexual recombination play in nature to maintain clonal population structures and produce or expand parasite clones capable of causing disease epidemics for two tissue encysting parasites. We applied high-resolution genotyping against strains isolated from a T. gondii waterborne outbreak that caused symptomatic disease in 155 immune-competent people in Brazil and a S. neurona outbreak that resulted in a mass mortality event in Southern sea otters. In both cases, a single, genetically distinct clone was found infecting outbreak-exposed individuals. Furthermore, the T. gondii outbreak clone was one of several apparently recombinant progeny recovered from the local environment. Since oocysts or sporocysts were the infectious form implicated in each outbreak, the expansion of the epidemic clone can be explained by self-mating. The results also show that out-crossing preceded selfing to produce the virulent T. gondii clone. For the tissue encysting coccidia, self-mating exists as a key adaptation potentiating the epidemic expansion and transmission of newly emerged parasite clones that can profoundly shape parasite population genetic structures or cause devastating disease outbreaks