Characterization of the small RNAs of Volvox carteri
ABSTRACT: 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 . Additionally, Baulcombe and colleagues showed that Chlamydomonas contains functional RNAi and miRNA machineries . We deep sequenced small RNAs of the female Volvox strain HK10 in different life stages (asexual reproduction to sexual reproduction), each time dividing the samples into somatic cells and reproductive cells. This allowed for the observation not only of differences in individual life stages, but also for monitoring sRNA content in the two cell types. We show that Volvox expresses miRNAs and that they are 2’-O-methylated at the 3’ end. The expression profiles of several miRNAs were validated by Northern blotting showing a differential expression both between cell types and between life stages. Intriguingly, most miRNAs do not seem to be conserved between Volvox and Chlamydomonas, raising the interesting question if this changed miRNome leads to differently targeted mRNAs thus resulting in cell differentiation. Since only little is known about the transcriptome of Volvox, we performed RNASeq in order to analyze potential miRNA targets. In conclusion, most miRNA in Volvox are not conserved in Chlamydomonas although the two species are evolutionary close together. This suggests that dramatic changes in the miRNA expression might be one of the driving forces for the development of multicellularity. 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 of Volvox carteri during different stages of its life cycle
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 . Additionally, Baulcombe and colleagues showed that Chlamydomonas contains functional RNAi and miRNA machineries . 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
Project description:Motivation: MicroRNAs (miRNAs) are short regulatory RNAs derived from a longer precursor RNA. miRNA biogenesis has been studied in animals and plants, recently elucidating more diverse and complex aspects, such as non-conserved, speciesspecific, and heterogeneous miRNA precursor populations. Small RNA sequencing data can be used to computationally determine genomic loci of miRNA precursors. The challenge is to predict a valid miRNA precursor from inhomogeneous read coverage: while the mature miRNA typically produces hundreds of sequence reads, the remaining part of the precursor is covered very sparsely. Results: We introduce a new conservation-independent method for the identification of miRNA precursors, that allows for speciesspecific heterogeneous precursor populations. The algorithm requires small RNA sequencing data and evaluates precursor secondary structures, with key parameters that can be adjusted based on the specific organism under investigation (within animals, plants, algae). We illustrate the validity of results from our algorithm using sequencing data for the two Volvocine algae Chlamydomonas reinhardtii (Chlamydomonas) and Volvox carteri (Volvox). Both organisms show little cross-species miRNA sequence conservation, and a heterogeneous miRNA precursor population. We validate our list of Chlamydomonas miRNAs with annotated miRNAs, and demonstrate excellent agreement. Furthermore, we are able to identify additional novel miRNA precursors, with structures ranging from simple mammalian-like hairpins to precursor structures indicating the creation of multiple mature/star miRNA duplexes. Novel miRNAs identified in Volvox show no similarity to mature miRNAs in Chlamydomonas. These results confirm the need for conservationindependent miRNA identification methods. Examination of small RNAs of Volvox carteri during different stages of its life cycle
Project description:We identified 174 miRNAs expressed in Volvox carteri. Some of Volvox miRNAs are highly enriched in gonidia or somatic cells. Subsequently, we predicted the targets of Volvox miRNAs and found many of target genes were regulated through mRNA degradation. Conservation analysis suggests the common origin of miRNA between Volvox and Chlamydomonas and high frequency of birth and death of Volvox miRNAs. Identification of miRNAs in multicellular green Volvox
Project description:Germ-soma differentiation is a hallmark of complex multicellular organisms, yet its origins are not well understood. Volvox carteri is a simple multicellular green alga that has recently evolved a simple germ-soma dichotomy with only two cell types: large germ cells called gonidia and small terminally differentiated somatic cells. Here, we provide a comprehensive characterization of the gonidial and somatic transcriptomes of Volvox to uncover fundamental differences between the molecular and metabolic programming of these cell types. We found extensive transcriptome differentiation between cell types, with somatic cells expressing a more specialized program overrepresented in younger, lineage-specific genes and gonidial cells expressing a more generalist program overrepresented in more ancient genes that shared striking overlap with metazoan stem-cell-specific genes. Directed analyses of metabolic pathways revealed a strong dichotomy between cell types with gonidial cells expressing growth-related genes and somatic cells expressing an altruistic metabolic program geared towards the assembly of flagella, which support organismal motility, and the conversion of storage carbon to sugars, which act as donors of extracellular matrix glycoproteins whose secretion enables massive organismal expansion. Volvox orthologs of Chlamydomonas diurnally regulated genes were analyzed for cell-type distribution and found to be strongly partitioned, with expression of dark-phase genes overrepresented in somatic cells and light-phase genes overrepresented in gonidial cells, a result that is consistent with cell type programs in Volvox arising by cooption of temporal regulons in a unicellular ancestor. Together our findings reveal fundamental molecular, metabolic, and evolutionary mechanisms that underlie the origins of germ-soma differentiation in Volvox and provide a template for understanding the acquisition of germ-soma differentiation in other multicellular lineages. Overall design: Gonidial cells and somatic cells of Volvox were separated and RNA was extracted from each cell type pool. RNA-seq libraries were generated from the extracted RNA and sequenced using high-throughput Illumina sequencing. A total of 4 RNA-seq samples were generated, 2 gonidial cell biological replicates and 2 somatic cell biological replicates.
Project description:RNA populations in Chlamydomonas reinhardtii Keywords: Highly parallel pyrosequencing Small RNAs were prepared from Chlamydomonas reinhardtii total extracts,ligated to a 3' adaptor and a 5' acceptor sequentially, and then RT-PCR amplified. PCR products were reamplified using a pair of 454 cloning primers and provided to 454 Life Sciences (Branford, CT) for sequencing. For technical details, see Tao Zhao, Guanglin Li, Shijun Mi, Shan Li, Gregory J. Hannon, Xiu-Jie Wang, and Yijun Qi. 2007. A Complex System of Small RNAs in the Unicellular Green Alga Chlamydomonas reinhardtii. Genes & Development
Project description:The recent identification of catalytically active peptidylglycine -amidating monooxygenase (PAM) in Chlamydomonas reinhardtii, a unicellular green alga, suggested the presence of a PAM-like gene and peptidergic signaling in the last eukaryotic common ancestor (LECA). Homologs of prototypical neuropeptide precursors and essential peptide processing enzymes (subtilisin-like prohormone convertases and carboxypeptidase B-like enzymes) were identified in the C. reinhardtii genome. Reasoning that sexual reproduction by C. reinhardtii requires extensive communication between cells, we used mass spectroscopy to identify proteins recovered from the soluble secretome of mating gametes and searched for evidence that the putative peptidergic processing enzymes were functional.After fractionation by SDS-PAGE, signal peptide-containing proteins that remained intact or were subjected to cleavage were identified. The C. reinhardtii mating secretome contained multiple matrix metalloproteinases, cysteine endopeptidases and serine carboxypeptidases, along with one subtilisin-like proteinase. Published transcriptomic studies support a role for these proteases in sexual reproduction. Multiple extracellular matrix proteins (ECM) were identified in the soluble mating secretome. Several pherophorins, ECM glycoproteins homologous to the Volvox sex-inducing pheromone, were present; most contained typical peptide processing sites and had been cleaved, generating stable N- or C-terminal fragments. Our data suggest that subtilisin endoproteases and matrix metalloproteinases similar to those important in vertebrate peptidergic and growth factor signaling play an important role in stage transitions during the life cycle of C. reinhardtii.
Project description:Small RNAs (21-24 nt) are pivotal regulators of gene expression that guide both transcriptional and post-transcriptional silencing mechanisms in diverse eukaryotes, including most if not all plants. MicroRNAs (miRNAs) and short interfering RNAs (siRNAs) are the two major types, both of which have a demonstrated and important role in plant development, stress responses and pathogen resistance. In this work, we used a deep sequencing approach (Sequencing-By-Synthesis, or SBS) to develop sequence resources of small RNAs from cultures of Volvox carteri (in control, phosphate starvation and sulphate starvation conditions). The high depth of the resulting datasets enabled us to examine in detail critical small RNA features as size distribution, tissue-specific regulation and sequence conservation between different organs in this species. We also developed database resources and a dedicated website (http://smallrna.udel.edu/) with computational tools for allowing other users to identify new miRNAs or siRNAs involved in specific regulatory pathways, verify the degree of conservation of these sequences in other plant species and map small RNAs on genes or larger regions of the genome under study. Small RNA libraries were derived from cultures of Volvox carteri in control, phosphate starvation and sulphate starvation conditions. Total RNA was isolated using the TriReagent (Molecular Research Center), and submitted to Illumina (Hayward, CA, http://www.illumina.com) for small RNA library construction using approaches described in (Lu et al., 2007) with minor modifications. The small RNA libraries were sequenced with the Sequencing-By-Synthesis (SBS) technology by Illumina. PERL scripts were designed to remove the adapter sequences and determine the abundance of each distinct small RNA. We thank Kan Nobuta and Gayathri Mahalingam for assistance with the computational methods.
Project description:Whole Genome Metabolism of "Volvox carteri f. nagariensis"
This is a whole genome metabolism model of Volvox carteri f. nagariensis.
This model has been automatically generated by the SuBliMinaL Toolbox
and libAnnotationSBML using information coming from from KEGG (release 66, April 2013, accessed via the resource's web services interface) and, where relevant, augmented with metabolic pathway information extracted from MetaCyc (version 17.0, March 2013).
This model has been produced by the path2models
project and is currently hosted on BioModels Database
and identified by: BMID000000140930
Other models with the same genus include BMID000000002039 BMID000000002040 BMID000000002041 BMID000000002042 BMID000000002043 BMID000000002044 BMID000000002045 BMID000000002046 BMID000000002047 BMID000000002048 BMID000000002049 BMID000000002050 BMID000000002051 BMID000000002052 BMID000000002053 BMID000000002054 BMID000000002055 BMID000000002056 BMID000000002057 BMID000000002058 BMID000000002059 BMID000000002060 BMID000000002061 BMID000000002062 BMID000000002063 BMID000000002064 BMID000000002065 BMID000000002066 BMID000000002067 BMID000000002068 BMID000000036631 BMID000000036632 BMID000000036633 BMID000000036634 BMID000000036635 BMID000000036636 BMID000000036637 BMID000000036638 BMID000000036639 BMID000000036640 BMID000000036641 BMID000000036642 BMID000000036643 BMID000000036644 BMID000000036645 BMID000000036646 BMID000000036647 BMID000000036648 BMID000000036649 BMID000000036650 BMID000000036651 BMID000000036652 BMID000000036653 BMID000000036654 BMID000000036655 BMID000000036656 BMID000000036657 BMID000000036658 BMID000000036659 BMID000000036660 BMID000000036661 BMID000000036662 BMID000000036663 BMID000000036664 BMID000000036665 BMID000000036666 BMID000000036667 BMID000000036668 BMID000000036669 BMID000000036670 BMID000000036671 BMID000000036672 BMID000000036673 BMID000000036674 BMID000000036675 BMID000000036676 BMID000000036677 BMID000000036678 BMID000000036679 BMID000000036680 BMID000000036681 BMID000000036682 BMID000000036683 BMID000000036684 BMID000000036685 BMID000000036686 BMID000000036687 BMID000000036688 BMID000000036689 BMID000000036690 BMID000000036691 BMID000000036692 BMID000000036693 BMID000000036694 BMID000000036695 BMID000000036696 BMID000000036697 BMID000000036698 BMID000000036699 BMID000000036700 BMID000000036701 BMID000000036702 BMID000000036703 BMID000000036704 BMID000000036705 BMID000000036706 BMID000000036707 .
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