Project description:Currently only four genome sequences for Limnothrix spp. are publicly available, and information on the genetic properties of cyanobacteria belonging to this genus is limited. In this study, we report the draft genome of Limnothrix sp. CACIAM 69d, isolated from the reservoir of a hydroelectric dam located in the Amazon ecosystem, from where cyanobacterial genomic data are still scarce. Comparative genomic analysis of Limnothrix revealed the presence of key enzymes in the cyanobacterial central carbon metabolism and how it is well equipped for environmental sulfur and nitrogen acquisition. Additionally, this work covered the analysis of Limnothrix CRISPR-Cas systems, pathways related to biosynthesis of secondary metabolites and assembly of extracellular polymeric substances and their exportation. A trans-AT PKS gene cluster was identified in two strains, possibly related to the novel toxin Limnothrixin biosynthesis. Overall, the draft genome of Limnothrix sp. CACIAM 69d adds new data to the small Limnothrix genome library and contributes to a growing representativeness of cyanobacterial genomes from the Amazon region. The comparative genomic analysis of Limnothrix made it possible to highlight unique genes for each strain and understand the overall features of their metabolism.
Project description:Background: Ependymomas encompass multiple, clinically relevant tumor types based on localization and molecular profiles. Although tumors of the methylation class “spinal ependymoma” (SP-EPN) represent the most common intramedullary neoplasms in children and adults, their developmental origin is ill-defined, molecular data are scarce, and the potential heterogeneity within SP-EPN remains unexplored. The only known recurrent genetic events in SP-EPN are loss of chromosome 22q and NF2 mutations, but neither types and frequency of these alterations nor their clinical meaning have been described in a large, epigenetically defined series. Methods: We mapped SP-EPN transcriptomes (n=76) to developmental atlases of the developing and adult spinal cord to uncover potential developmental origins of these tumors. In addition, transcriptomic, epigenetic (n=234), genetic (n=140), and clinical analyses (n=115) were integrated for a detailed overview on this entity. Results: Integration of transcriptomic ependymoma data with single-cell atlases of the spinal cord identified mature adult ependymal cells to display highest similarities to SP-EPN. Unsupervised hierarchical clustering of tumor data together with integrated analysis of methylation profiles identified two molecular SP-EPN subtypes. Subtype 1 predominantly contained NF2 wild type sequences with regular NF2 expression but revealed more extensive copy number alterations. Subtype 2 harbored previously known germline or sporadic NF2 mutations and was NF2-deficient in most cases, more often showed multilocular disease, and demonstrated a significantly reduced progression-free survival. Conclusion: Based on integrated molecular profiling of a large tumor series we identify two distinct SP-EPN subtypes with important implications for genetic counseling, patient surveillance, and drug development priorities.
Project description:Background: Ependymomas encompass multiple, clinically relevant tumor types based on localization and molecular profiles. Although tumors of the methylation class “spinal ependymoma” (SP-EPN) represent the most common intramedullary neoplasms in children and adults, their developmental origin is ill-defined, molecular data are scarce, and the potential heterogeneity within SP-EPN remains unexplored. The only known recurrent genetic events in SP-EPN are loss of chromosome 22q and NF2 mutations, but neither types and frequency of these alterations nor their clinical meaning have been described in a large, epigenetically defined series. Methods: We mapped SP-EPN transcriptomes (n=76) to developmental atlases of the developing and adult spinal cord to uncover potential developmental origins of these tumors. In addition, transcriptomic, epigenetic (n=234), genetic (n=140), and clinical analyses (n=115) were integrated for a detailed overview on this entity. Results: Integration of transcriptomic ependymoma data with single-cell atlases of the spinal cord identified mature adult ependymal cells to display highest similarities to SP-EPN. Unsupervised hierarchical clustering of tumor data together with integrated analysis of methylation profiles identified two molecular SP-EPN subtypes. Subtype 1 predominantly contained NF2 wild type sequences with regular NF2 expression but revealed more extensive copy number alterations. Subtype 2 harbored previously known germline or sporadic NF2 mutations and was NF2-deficient in most cases, more often showed multilocular disease, and demonstrated a significantly reduced progression-free survival. Conclusion: Based on integrated molecular profiling of a large tumor series we identify two distinct SP-EPN subtypes with important implications for genetic counseling, patient surveillance, and drug development priorities.
Project description:Background: Ependymomas encompass multiple, clinically relevant tumor types based on localization and molecular profiles. Although tumors of the methylation class “spinal ependymoma” (SP-EPN) represent the most common intramedullary neoplasms in children and adults, their developmental origin is ill-defined, molecular data are scarce, and the potential heterogeneity within SP-EPN remains unexplored. The only known recurrent genetic events in SP-EPN are loss of chromosome 22q and NF2 mutations, but neither types and frequency of these alterations nor their clinical meaning have been described in a large, epigenetically defined series. Methods: We mapped SP-EPN transcriptomes (n=76) to developmental atlases of the developing and adult spinal cord to uncover potential developmental origins of these tumors. In addition, transcriptomic, epigenetic (n=234), genetic (n=140), and clinical analyses (n=115) were integrated for a detailed overview on this entity. Results: Integration of transcriptomic ependymoma data with single-cell atlases of the spinal cord identified mature adult ependymal cells to display highest similarities to SP-EPN. Unsupervised hierarchical clustering of tumor data together with integrated analysis of methylation profiles identified two molecular SP-EPN subtypes. Subtype 1 predominantly contained NF2 wild type sequences with regular NF2 expression but revealed more extensive copy number alterations. Subtype 2 harbored previously known germline or sporadic NF2 mutations and was NF2-deficient in most cases, more often showed multilocular disease, and demonstrated a significantly reduced progression-free survival. Conclusion: Based on integrated molecular profiling of a large tumor series we identify two distinct SP-EPN subtypes with important implications for genetic counseling, patient surveillance, and drug development priorities.
Project description:Argonaute (Ago) proteins are key players in RNA interference in eukaryotes, where they function as RNA-guided RNA endonucleases. Prokaryotic Argonautes (pAgos) are much more diverse than their eukaryotic counterparts but their cellular functions and mechanisms of action remain largely unknown. Some pAgos were shown to use small DNA guides for endonucleolytic cleavage of complementary DNA in vitro. However, previously studied pAgos from thermophilic prokaryotes function at elevated temperatures, which limits their potential use as a tool in genomic applications. Here, we describe two pAgos from mesophilic bacteria, Clostridium butyricum (CbAgo) and Limnothrix rosea (LrAgo), that act as DNA-guided DNA nucleases at physiological temperatures. In comparison with previously studied pAgos, CbAgo and LrAgo do not show strong preferences for the 5'-nucleotide in guide DNA and can use not only 5'-phosphorylated but also 5'-hydroxyl DNA guides. Both CbAgo and LrAgo can tolerate guide/target mismatches in the seed region, but are sensitive to mismatches in the 3'-guide region. Both pAgos can perform programmable endonucleolytic cleavage of double-stranded DNA substrates, showing enhanced activity at AT-rich regions and at elevated temperatures. The biochemical characterization of mesophilic pAgo proteins paves the way for their use for DNA manipulations both in vitro and in vivo.
Project description:C-phycocyanin (C-PC) is a blue colored accessory photosynthetic pigment found in cyanobacteria. Some of the medicinal properties of Spirulina have been attributed to this pigment, which includes anticancer, antioxidant, and anti-inflammatory activity. We have screened cyanobacteria isolated from freshwater habitats in Florida for their high content of C-PC. Of 125 strains tested, one filamentous strain identified as Limnothrix sp. was selected for further research. This strain produced 18% C-PC of total dry biomass. Here we describe a simple method for obtaining C-PC of high purity without the use of ion exchange chromatography. The procedure is based on pigment precipitation from the cell lysate with an appropriate concentration of ammonium sulfate, then purification with activated carbon and chitosan, followed by a sample concentration using tangential flow filtration. We have shown that when the lower concentration of ammonium sulfate was used, C-PC with higher purity index was recovered. Characterization of C-PC from Limnothrix showed that it had an absorbance maximum at 620nm and fluorescence at 639nm. The molecular mass of intact C-PC was estimated to be ~50kDa with ? and ? subunits forming dimmers. When C-PC content per unit biomass was compared to that of marketed Spirulina powder, we found that Limnothrix was superior. C-phycocyanin from Limnothrix had an antioxidative activity on DPPH free radicals similar to that found in a natural antioxidant - rutin.