Project description:In the social amoebae (Dictyostelia) quorum sensing system mediates aggregation of single cells into multicellular aggregates by chemotactic movement towards gradients of diffusible molecules known as acrasins. The acrasin of P. violaceum is the unusual dipeptide N-propionyl-gamma-L-glutamyl-L-ornithine-delta-lactam-ethylester, known as glorin. Phylogenetic analysis has indicated that P. violaceum is more related to the most derived group 4 dictyostelids than to the ancient group 2 polysphondylids such as P. pallidum. Nevertheless it has been reported that P. pallidum cells respond to glorin in chemotaxis assays. This has led to the assumption that glorin-based communication may be the most ancient form of intercellular communication that Dictyostelia invented to organize early steps of multicellular development. In this study we show that glorin mediates rapid changes in gene expression at the transition from vegetative growth to aggregation, apparently without pronounced cross-talk with the cyclic AMP-based communication system that coordinates post-aggregation events in this species. We describe glorin-mediated changes in gene expression in the social amoeba Polysphondylium pallidum at the transition from unicellular growth to multicellular development. Comparison of gene expression in growing cells versus cells starving for 2 or 3 hours in the presence or absence of glorin.
Project description:In the social amoebae (Dictyostelia) quorum sensing system mediates aggregation of single cells into multicellular aggregates by chemotactic movement towards gradients of diffusible molecules known as acrasins. The acrasin of P. violaceum is the unusual dipeptide N-propionyl-gamma-L-glutamyl-L-ornithine-delta-lactam-ethylester, known as glorin. Phylogenetic analysis has indicated that P. violaceum is more related to the most derived group 4 dictyostelids than to the ancient group 2 polysphondylids such as P. pallidum. Nevertheless it has been reported that P. pallidum cells respond to glorin in chemotaxis assays. This has led to the assumption that glorin-based communication may be the most ancient form of intercellular communication that Dictyostelia invented to organize early steps of multicellular development. In this study we show that glorin mediates rapid changes in gene expression at the transition from vegetative growth to aggregation, apparently without pronounced cross-talk with the cyclic AMP-based communication system that coordinates post-aggregation events in this species. We describe glorin-mediated changes in gene expression in the social amoeba Polysphondylium pallidum at the transition from unicellular growth to multicellular development.
Project description:Myxococcus xanthus is a model organism for studying social behaviors and cell differentiation in bacteria. Upon nutrient depletion, M. xanthus cells initiate a developmental program that culminates in formation of spore-filled fruiting bodies and peripheral rods outside of fruiting bodies. Completion of this developmental program depends on fine-tuned spatial and temporal regulation of gene expression, intercellular communication, signaling by nucleotide-based second messengers, and motility. In order to understand stage-specific gene expression during growth and development, we extracted total RNA from vegetative cells (referred as 0 h of development) and from cells developed for 6, 12, 18 and 24 h under submerged conditions in two replicates.
Project description:Myxococcus xanthus is a model organism for studying social behaviors and cell differentiation in bacteria. Upon nutrient depletion, M. xanthus cells initiate a developmental program that culminates in formation of spore-filled fruiting bodies and peripheral rods outside of fruiting bodies. Completion of this developmental program depends on fine-tuned spatial and temporal regulation of gene expression, intercellular communication, signaling by nucleotide-based second messengers, and motility. In order to understand regulation of gene expression during growth and development, transcription start sites were identified using Cappable-seq. To this end, we extracted total RNA from vegetative cells (referred as 0 h of development) and from cells developed for 6, 12, 18 and 24 h under submerged conditions in two replicates.
Project description:In many developmental systems, morphogenesis is coupled with dramatic changes in spatiotemporal gene expression, often orchestrated by the coordinated action of transcription factors. Development of the social soil amoebae Dictyostelium discoideum proceeds through a sequence of morphological and transcriptional changes, but the role of transcription factors in development is not well understood. GtaC, a GATA-type zinc-finger transcription factor, is essential for Dictyostelium development. It decodes pulsatile extracellular cAMP signals during early development and mediates cell-type differentiation at later stages. Here, we studied the developmental regulatory roles of GtaC through the concerted analysis of temporal ChIP- and RNA-sequencing data from strains that carry different alleles of gtaC. We show that GtaC exhibits temporally distinctive DNA-binding patterns throughout early development, accompanied by largely cotemporaneous expression of its target genes. We also show that GtaC binds DNA in two modes. One of these modes exhibits binding preferences for canonical GATA-like sequences, the regulatory consequences accompanying which is predominantly up-regulation of target gene expression. The other binding mode is mostly associated with down-regulation. Among its targets we find transcription factors that are essential for development as well as genes involved in cAMP signaling and cell-type specification. Our results suggest that GtaC is a master regulator that regulates multiple physiological processes during early development, when Dictyostelium transitions from a group of unicellular amoebae to an integrated multicellular organism. Cotemporaneous transcriptional profiling and ChIP sequencing during early Dictyostelium development
Project description:Metastasis is a major cause of mortality, and remains a final frontier in the search for a cure for cancer. While there has been much research on the ‘seed’ (metastatic tumor cells) and the ‘soil’ (colonized host tissue), interactions between metastatic cancer cells and stromal endothelial cells, which occur at multiple stages during metastasis, are less well understood. Here we report a dynamic regulation of the endothelium by cancer cells through the formation of nanoscale intercellular membrane bridges, which act as physical conduits for intercellular communication in vitro and in vivo, including horizontal transfer of microRNAs (miRNA). The communication between the tumor cell and the endothelium upregulates markers associated with pathological endothelium, which is reversed by pharmacological inhibition of these nanoscale conduits. These results lead us to define the notion of “metastatic hijack”: cancer cell-induced transformation of healthy endothelium into pathological endothelium via horizontal communication through the nanoscale conduits. Pharmacological perturbation of these nanoscale membrane bridges decreases metastatic foci in syngeneic- and human xenograft-breast cancer models. Targeting the formation of these nanoscale membrane bridges may potentially emerge as a new therapeutic opportunity in the management of metastatic cancer. A miRNA microarray was used to evaluate the transport of endogenous microRNAs. The intercellular transfer-ve and intercellular transfer+ve samples were sorted from the same endothelial cell population with the only difference being the occurrence of intercellular transport. The heat map shows potential miRNA candidates for exogenous transfer on two independent biological replicates. These miRNA candidates were significantly up-regulated in the cells receiving transfer of intercellular contents. HUVECs that were not exposed to cancer cells were used as a baseline control.
Project description:In many developmental systems, morphogenesis is coupled with dramatic changes in spatiotemporal gene expression, often orchestrated by the coordinated action of transcription factors. Development of the social soil amoebae Dictyostelium discoideum proceeds through a sequence of morphological and transcriptional changes, but the role of transcription factors in development is not well understood. GtaC, a GATA-type zinc-finger transcription factor, is essential for Dictyostelium development. It decodes pulsatile extracellular cAMP signals during early development and mediates cell-type differentiation at later stages. Here, we studied the developmental regulatory roles of GtaC through the concerted analysis of temporal ChIP- and RNA-sequencing data from strains that carry different alleles of gtaC. We show that GtaC exhibits temporally distinctive DNA-binding patterns throughout early development, accompanied by largely cotemporaneous expression of its target genes. We also show that GtaC binds DNA in two modes. One of these modes exhibits binding preferences for canonical GATA-like sequences, the regulatory consequences accompanying which is predominantly up-regulation of target gene expression. The other binding mode is mostly associated with down-regulation. Among its targets we find transcription factors that are essential for development as well as genes involved in cAMP signaling and cell-type specification. Our results suggest that GtaC is a master regulator that regulates multiple physiological processes during early development, when Dictyostelium transitions from a group of unicellular amoebae to an integrated multicellular organism.