Biology subjects

Aboobaker, A.

Publications and source records attributed to Aboobaker, A..

7 recordsLinked to original sources

bioRxiv2017

A platform for efficient transgenesis in Macrostomum lignano, a flatworm model organism for stem cell research

Regeneration-capable flatworms are informative research models to study the mechanisms of stem cell regulation, regeneration and tissue patterning. However, the lack of transgenesis methods significantly hampers their wider use. Here we report development of a transgenesis method for Macrostomum lignano, a basal flatworm with excellent regeneration capacity. We demonstrate that microinjection of DNA constructs into fertilized one-cell stage eggs, followed by a low dose of irradiation, frequently results in random integration of the transgene in the genome and its stable transmission through the germline. To facilitate selection of promoter regions for transgenic reporters, we assembled and annotated the M. lignano genome, including genome-wide mapping of transcription start regions, and showed its utility by generating multiple stable transgenic lines expressing fluorescent proteins under several tissue-specific promoters. The reported transgenesis method and annotated genome sequence will permit sophisticated genetic studies on stem cells and regeneration using M. lignano as a model organism.

bioengineering↗

bioRxiv2017

The Abrogation Of Condensin Function Provides Independent Evidence For Defining The Self-Renewing Population Of Pluripotent Stem Cells

Heterogeneity of planarian neoblast stem cells has been categorised on the basis of single cell expression analyses and subsequent experiments to demonstrate lineage relationships. Some data suggest that despite gene expression heterogeneity amongst cells in the cell cycle, in fact only one sub-population, known as sigma neoblasts, can self-renew. Without the tools to perform live in vivo lineage analysis, we instead took an alternative approach to provide independent evidence for defining the self-renewing stem cell population. We exploited the role of highly conserved condensin proteins to functionally assay neoblast self-renewal properties. Condensins are involved in forming properly condensed chromosomes to allow cell division to proceed during mitosis, and their abrogation inhibits mitosis and can lead to repeated endoreplication of the genome in cells that make repeated attempts to divide. We find that planarians possess only the condensin I complex, and that this is required for normal stem cell function. Abrogation of condensin function led to rapid stem cell depletion accompanied by the appearance of giant cells with increased DNA content. Using previously discovered markers of heterogeneity we show that enlarged cells are always from the sigma-class of the neoblast population and we never observe evidence for endoreplication for the other neoblast subclasses. Overall, our data establish that condensins are essential for stem cell maintenance and provide independent evidence that only sigma-neoblasts are capable of multiple rounds of cell division and hence self-renewal.

developmental biology↗

bioRxiv2017

MLL3/4 Prevents Stem Cell Hyperplasia And Controls Differentiation Programs In A Planarian Cancer Stem Cell Model

Currently, little is known about the evolution of epigenetic regulation in animal stem cells. Using the planarian stem cell system to investigate the role of the COMPASS family of MLL3/4 histone methyltransferases, we demonstrate that their role as tumour suppressors in stem cells is conserved over a large evolutionary distance in animals. This also suggested the potential conservation of a genome wide epigenetic regulation program in animal stem cells, so we assessed the regulatory effects of Mll3/4 loss of function by performing RNA-seq and ChIP-seq on the G2/M planarian stem cell population, part of which contributes to the formation of outgrowths. We find many oncogenes and tumour suppressors among the affected genes that are therefore likely candidates for mediating MLL3/4 tumour suppression function in mammals, where little is known about in vivo regulatory targets. Our work demonstrates conservation of an important epigenetic regulatory program in animals and highlights the utility of the planarian model system for studying epigenetic regulation.

developmental biology↗

bioRxiv2017

Epigenetic analyses of the planarian genome reveals conservation of bivalent promoters in animal stem cells.

Planarian flatworms have an indefinite capacity to regenerate missing or damaged body parts owing to a population of pluripotent adult stems cells called neoblasts (NBs). Currently, little is known about the importance of the epigenetic status of NBs and how histone modifications regulate homeostasis and cellular differentiation. We have developed an improved and optimized ChIP-seq protocol for NBs in Schmidtea mediterranea and have generated genome-wide profiles for the active marks H3K4me3 and H3K36me3, and suppressive marks H3K4me1 and H3K27me3. The genome-wide profiles of these marks were found to correlate well with NB gene expression profiles. We found that genes with little transcriptional activity in the NB compartment but which switch on in post-mitotic progeny during differentiation are bivalent, being marked by both H3K4me3 and H3K27me3 at promoter regions. In further support of this hypothesis bivalent genes also have a high level of paused RNA Polymerase II at the promoter-proximal region. Overall, this study confirms that epigenetic control is important for the maintenance of a NB transcriptional program and makes a case for bivalent promoters as a conserved feature of animal stem cells and not a vertebrate specific innovation. By establishing a robust ChIP-seq protocol and analysis methodology, we further promote planarians as a promising model system to investigate histone modification mediated regulation of stem cell function and differentiation.

developmental biology↗

bioRxiv2016

The innate immune systems of malacostracan crustaceans exhibit both conserved and evolutionarily distinct components

Growing demands for aquatic sources of animal proteins have attracted significant investments in aquaculture research in recent years. The crustacean aquaculture industry has undergone substantial growth to accommodate a rising global demand, however such large-scale production is susceptible to pathogen-mediated destruction. It is clear that a thorough understanding of the crustacean innate immune system is imperative for future research into combating current and future pathogens of the main food crop species. Through a comparative genomics approach utilising extant data from 55 species, we describe the innate immune system of crustaceans from the Malacostraca class. We identify 7407 malacostracan genes from 39 gene families implicated in different aspects of host defence and demonstrate dynamic evolution of innate immunity components within this group. Malacostracans have achieved flexibility in recognising infectious agents through divergent evolution and expansion of pathogen recognition receptors genes. Antiviral RNAi, Toll and JAK-STAT signal transduction pathways have remained conserved within Malacostraca, although the Imd pathway appears to lack several key components. Immune effectors such as the antimicrobial peptides (AMPs) have unique evolutionary profiles, with many malacostracan AMPs not found in other arthropod groups. Lastly, we describe four putative novel immune gene families, characterised by distinct protein domains, potentially representing important evolutionary novelties of the malacostracan immune system.

evolutionary biology↗

bioRxiv2016

Analyses of the core eukaryotic protein subunit of telomerase support extensive adaptation to different evolutionary and life histories in the Metazoa

Most animals employ telomerase, which consists of a catalytic subunit known as the telomerase reverse transcriptase (TERT) and an RNA template, to maintain telomere ends. Given the importance of TERT and the apparent importance of telomere biology in core metazoan life history traits like ageing and the control of somatic cell proliferation, we hypothesised that TERT would have patterns of sequence and regulatory evolution reflecting adaptations to diverse evolutionary and life histories across the Animal Kingdom. To test this, we performed a complete investigation of the evolutionary history of TERT across animals. We show that although TERT is almost ubiquitous across Metazoa, it has undergone substantial sequence evolution in canonical motifs. Beyond the known canonical motifs, we also identify and compare regions that are highly variable between lineages, but for which conservation exists within phyla. Recent data have highlighted the importance of alternate splice forms of TERT in non-canonical functions in some animals. Although animals may share some conserved introns, we find that the selection of exons for alternative splicing appears to be highly variable, and regulation by alternative splicing appears to be a very dynamic feature of TERT evolution. We show that even within a closely related group of triclad flatworms, where alternative splicing of TERT was previously correlated with reproductive strategy, we observe highly diverse alternative splicing patterns. Our work establishes that the evolutionary history and structural evolution of TERT involves previously unappreciated levels of change, supporting the view that this core eukaryotic protein has adapted to the requirements of diverse animal life histories.

evolutionary biology↗

bioRxiv2016

A shielded irradiation assay to investigate mechanisms of in vivo stem cell migration in planarians

Migration of stem cells underpins the physiology of metazoan animals. For tissues to be maintained, stem cells and their progeny must migrate and differentiate in the correct positions. This need is even more acute after tissue damage by wounding or pathogenic infections. Inappropriate migration also underpins the formation of metastasis. Despite this, few mechanistic studies address stem cell migration during repair or homeostasis in adult tissues. Here, we present a shielded X-ray irradiation assay that allows us to follow stem cell migration in the planarians. We demonstrate that we can use this system to study the molecular control of stem cell migration and show that snail and zeb-1 EMT transcription factors homologs are necessary for cell migration to wound sites and for the establishment of migratory cell morphology. Our work establishes planarians as a suitable model for further in depth study of the processes controlling stem cell migration in vivo.

developmental biology↗