Project description:The experiment was performed with the intention of collecting transcriptomic data from isolated cell types (spore, stalk and vegetative cells) in Dictyostelium lacteum as well as from cysts in Polyshpondlyium pallidum. By combining these data with similar cell-type specific RNA-Seq data from other organisms, and by examining the expression patterns of transcription factor genes, we tried to characterize how gene regulation for cell differentiation evolved in Dictyostelia. Specifically ,we dissociated and collected spore and stalk from the fruiting bodies of D. lacteum at 24 hours of development. We also collected exponentially growing vegetative cells of D. lacteum. For collecting P. pallidum cyst samples, cells were induced to encyst with sorbitol, and samples were collected at 0, 8, 16, and 24 h of incubation. RNA was extracted using the RNeasy kit (QIAGEN), and cDNA libraries were made using the Illumina TruSeq kit. The Illumina sequencing platforms (NextSeq500 for D. lacteum samples, and Hi-seq 2000 for P. pallidum samples) were used for sequencing.
Project description:Activating mutations in tyrosine kinase (TK) genes (e.g. FLT3 and KIT) are found in more than 30% of patients with de novo acute myeloid leukemia (AML); many groups have speculated that mutations in other TK genes may be present in the remaining 70%. We performed high-throughput re-sequencing of the kinase domains of 26 TK genes (11 receptor TK and 15 cytoplasmic TK) that are expressed in most AML patients, using genomic DNA from the bone marrow (tumor) and matched skin biopsy samples (germline) from 94 patients with de novo AML; sequence variants were validated in an additional 94 AML tumor samples (14.3 million base pairs of sequence were obtained and analyzed). We identified known somatic mutations in FLT3, KIT, and JAK2 TK genes at the expected frequencies, and found four novel somatic mutations, JAK1V623A, JAK1T478S, DDR1A803V and NTRK1S677N, once each in four respective patients out of 188 tested. We also identified novel germline sequence changes encoding amino acid substitutions (i.e. non-synonymous changes) in 14 TK genes, including TYK2, which had the largest number of non-synonymous sequence variants (11 total detected). Additional studies will be required to define the roles that these somatic and germline TK gene variants play in AML pathogenesis. Experiment Overall Design: 188 patient samples analysed
Project description:Tachykinins (TKs) and their receptors have been shown to be expressed in the mammalian ovary. However, the biological roles for ovarian TKs have yet to be verified. Ci-TK and Ci-TK-R, characterized from the protochordate (ascidian), Ciona intestinalis, are prototypes of vertebrate TKs and their receptors. In the present study, we show a novel biological function of TKs as an inducible factor for oocyte growth using C. intestinalis as a model animal. Immunostaining demonstrated the specific expression of Ci-TK-R in test cells residing in oocytes at the vitellogenic stage. DNA microarray and real-time PCR substantiated that Ci-TK induced gene expression of several proteases including cathepsin D, chymotrypsin, and carboxypeptidase B2 and functionally unidentified lectins or glycosidases in the ovary. The enzymatic activities of the above proteases in the ovary were also shown to be enhanced by Ci-TK. Of particular significance is that treatment of Ciona oocytes with Ci-TK resulted in progression of growth from the vitellogenic stage to the post-vitellogenic stage, which was completely blocked by a TK antagonist or protease inhibitors. These results led to the conclusion that Ci-TK enhances growth of the vitellogenic oocytes via up-regulation of gene expression and enzymatic activities of the proteases. Keywords: tachykinins, ovary
