Project description:Chlorella sp. HS2 is a halotolerant microalga exhibiting relatively high biomass productivity and substantially high lipid accumulation in marine growth media, which suggests this alga as an important crop for industrial algal cultivation systems. To determine pathways leading to HS2's acclimation responses to salt stress, we performed RNA-seq analysis with triplicated cultures grown in freshwater and marine media at both exponential and stationary growth phases. We then run de novo assembly to obtain HS2 transcriptome, which in turn was annotated and processed to extract dysregulated pathways. Results showed a large proportion of down-regulated genes, for instance photosynthesis and TCA pathways. Photosynthesis appeared, however, to recover at the stationary phase, while the general down-regulation pattern was maintained.
Project description:Highly invasive integrin and protease-independent amoeboid migration is often employed by cancer cells at the invasive front, though little is known about the transcriptomic changes underlying the switch to an amoeboid migratory mode. Using a metastatic melanoma cell line expressing photoconvertible Dendra2 protein (WM983c-D2), tumour spheroids were cultured in 3D collagen matrices and imaged over time. Spheroids grown from WM983c-D2 cells generate cells of three distinct phenotypes: 1) compact, non-invading cells organised at the spheroid surface with no visual cell protrusions, hereafter termed ‘epithelial’; 2) cells still attached to the spheroid periphery that have commenced tumour escape, exhibiting cellular protrusions and an elongated phenotype, hereafter termed ‘escaping’; 3) singly migrating cells exhibiting a rounded phenotype and no lamellipodia consistent with amoeboid cell migration, hereafter termed 'amoeboid'. Individual amoeboid and escaping cells, as well as groups of epithelial cells at the spheroid edge were photoconverted and single cell sorted via FACS following enzymatic digestion of the collagen matrix. 10 Epithelial, 12 escaping and 13 amoeboid cells were subjected to scRNA-seq and differential expression analyses in order to identify changes in gene expression as melanoma cells convert from a non-invasive epithelial state to invasive amoeboid cells.
Project description:Dunaliella tertiolecta is an extremophilic, green alga from the Chlorophyte lineage. It is found in coastal marine environments around the world. D. tertiolecta can tolerate extremes of heat, light, pH, and salinity. D. tertiolecta is under development for the production biofuels and other bioproducts because it can produce large quantities of neutral lipids, and it can be grown in open raceway ponds using only the inputs of seawater and sunlight. This isolate of D. tertiolecta (UTEX LB 999) was found in Oslofjord, Norway in 1938. This accession includes an RNA-Seq analysis of D. tertiolecta cultures grown in iron-replete (1.5 µM) or iron-deficient (0 µM) media.
Project description:Amoeboid and mesenchymal migration of cancer cells both contribute to metastatic spreading of tumors. To characterize gene expression profiles underlying the different migratory modes, we performed RNA sequencing of HT1080 fibrosarcoma cells undergoing mesenchymal-amoeboid transition induced by either doxycycline-inducible constitutively active RhoA or dasatinib treatment. Cells were kept in three-dimensional collagen gels with or without induction for 48 hours. RNA was isolated with a modified Chomczynski protocol. RNA-seq libraries were constructed from DNase I treated, rRNA depleted total RNA and sequenced with Illumina 2000/2500 sequencers.
Project description:Microglia, the resident immune cells of the central nervous system (CNS), have two distinct phenotypes in the developing brain: amoeboid form, known to be amoeboid microglial cells (AMC) and ramified form, known to be ramified microglial cells (RMC) alongside several intermediate forms. The AMC are characterized by being proliferative, phagocytic and migratory whereas the RMC are quiescent and exhibit a slow turnover rate. The AMC transform into RMC with advancing age, and this transformation is indicative of the gradual shift in the microglial functions. Both AMC and RMC respond to CNS inflammation, and they become hypertrophic when they are activated by trauma, infection or neurodegenerative stimuli. The molecular mechanisms and functional significance of morphological transformation of microglia during normal development and in disease conditions is not clear. It is hypothesized that AMC and RMC are functionally regulated by a specific set of genes encoding various signaling molecules and transcription factors. To address this, we carried out cDNA microarray analysis using lectin-labeled AMC and RMC isolated from frozen tissue sections of the corpus callosum of 5-day and 4-week old rat brain respectively, by laser capture microdissection (LCM). The global gene expression profiles of both microglial phenotypes were compared and the differentially expressed genes in AMC and RMC were clustered based on their functional annotations. This genome wide comparative analysis helps in identifying genes that are specific to AMC and RMC. The novel and specific molecules identified in both microglial phenotypes can be targeted for therapeutic purposes in developing and adult brain diseases. We used microarrays to identify the genes specific to amoeboid and ramified microglia. RNA was isolated from the laser-captured amoeboid and ramified microglia from the corpus callosum of 5-day and 4-week old rat brain. The RNA was hybridised onto Affymetrix Rat 230 2.0 array.
Project description:Recently we demonstrated that amoeboid microglia in white matter regions are essential for proper oligodendrocyte homeostasis and myelinogenesis in the first postnatal week of mice. Amoeboid microglia in the corpus callosum change their activation profile already within few days after postnatal day (P)7 and microglia of the cerebellum show similar features to callosal microglia. Here we expanded our previous transcriptional analysis and performed bulk RNA sequencing of microglia during development in a detailed way in P7, P10 and P42 microglia from corpus callosum, cortex and cerebellum. The goal of this study was to identify a specific gene profile for both, white matter and grey matter microglia during development.
Project description:The green alga Volvox carteri is a model organism for the development of multicellularity. It has a spherical shape with a complete division of labor between around 2000 somatic cells and 16 reproductive cells. When comparing Volvox with its unicellular relative Chlamydomonas rheinhardtii, one striking observation is the similarity in the protein coding genes [1]. Additionally, Baulcombe and colleagues showed that Chlamydomonas contains functional RNAi and miRNA machineries [2]. We deep sequenced small RNAs associated with one Argonaute protein of the female Volvox strain Vol6 during its vegetative growth phase. Using these data, we established a miRNA identification pipeline that takes into account plant miRNA feature in general and also uses parameters employed in finding miRNAs in Chlamydomonas. Other small RNAs that are functionally incorporated into Ago are characterized. 1. Prochnik, S.E., et al., Genomic analysis of organismal complexity in the multicellular green alga Volvox carteri. Science, 2010. 329(5988): p. 223-6. 2. Molnar, A., et al., miRNAs control gene expression in the single-cell alga Chlamydomonas reinhardtii. Nature, 2007. 447(7148): p. 1126-9. Examination of small RNAs bound to an Argonaute protein of Volvox carteri