Unknown,Transcriptomics,Genomics,Proteomics

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Expression data analysis of murine pulmonary cryptococcosis induced by C. gattii

ABSTRACT: Our previous investigation indicated that high-virulence C. gattii (C. gattii TIMM 4097) tend to reside in the alveoli, whereas low-virulence C. gattii (C. gattii TIMM 4903) tend to be washed out from the alveoli and move into the central side of the respiratory system. To test this hypothesis, we performed microarray assay. C. gattii TIMM 4097 and C. gattii TIMM 4903 were prepared for the study. The mice were anesthetized and the spore suspension was intratracheally injected into each mouse. Mice were sacrificed 15 days after the infection and then examined. Lung of the mice infected with C. gattii TIMM 4097 and C. gattii TIMM 4903 and normal controls were obtained (n=3, respectively. RNA targets preparation was performed according to the manufacturer's protocol using GeneChip(R) 3' IVT Express Kit (Affymetrix). One hundred nanograms of total RNA were converted into double-stranded cDNA template for transcription. In vitro transcription synthesized amplified RNA (aRNA) and incorporated a biotin-conjugated nucleotide. After purification and fragmentation of aRNA, 12.5 ug of them was hybridized to GeneChip(R) Mouse Genome 430 2.0 Array (Affymetrix).The Probe Array was scanned using a GeneChip(R) Scanner 3000 7G

ORGANISM(S): Mus musculus

SUBMITTER: Yoichiro Okubo

PROVIDER: E-GEOD-48595 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Sox9 acts together with Sox5 or Sox6 as a master regulator for chondrocyte differentiation; however, how these transcription factors functionally interact and collaborate to regulate chondrogenesis remains unclear. Here we show that the protein kinase MLTK plays an essential role in the onset of chondrogenesis through triggering the induction of Sox6 by Sox9. Knockdown of MLTK in Xenopus embryos results in drastic loss of craniofacial cartilages without defects in neural crest formation. We also find that Sox6 is specifically induced during craniofacial chondrogenesis and this induction is inhibited by MLTK knockdown. Remarkably, Sox6-knockdown embryos display essentially the same phenotype as the MLTK-knockdown embryos; the drastic loss of cartilages and the marked down-regulation of genes involved in chondrogenesis. Microarray analysis reveals a remarkable similarity between Sox6-depleted and MLTK-depleted embryos in their gene expression pattern. Moreover, we find that ectopic expression of MLTK can induce Sox6 expression in a Sox9-dependent manner. These results identify a novel, key regulator for chondrogenesis. We used microarrays to describe the genome-wide gene expression profiles of xMLTK-depleted and xSox6-depleted embryos. CoMO, xMLTK-MO, or xSox6-MO was injected into the animal pole of all blastomeres (10 ng/cell) at the four-cell stage. The heads of embryos were cut out and harvested at 72 hours after fertilization (corresponding to about stage 41). Total RNA was extracted using TRIsol reagent, treated with DNase (TURBO DNase, Ambion), and then purified using RNeasy Mini Kit (QIAGEN) according to the manufacturerM-bM-^@M-^Ys instruction. The quality of total RNA was assessed using the Agilent 2100 BioAnalyzer. Reverse transcription to synthesize first-strand cDNA, second-strand cDNA synthesis, in vitro transcription to synthesize Biotin-modified aRNA, aRNA purification and fragmentation of the labeled aRNA were performed using the GeneChip 3M-bM-^@M-^Y IVT Express Kit and hybridization to the Affymetrix GeneChip Xenopus laevis Genome 2.0 Array was performed using the GeneChip Hybridization Wash and Stain Kit according to the manufacturerM-bM-^@M-^Ys instruction. Hybridized arrays were scanned using an Affymetrix GeneChip Scanner. Scanned Chip images were analyzed with GeneChip operating Software v.1.4 (GCOS) and GeneSpring GX 11.0.2. (Agilent technologies).

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Expression data analysis of murine pulmonary cryptococcosis induced by C. gattii

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