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Identification of mRNAs that are differentially expressed between serum-stimulated and non-stimulated A. caninum L3

ABSTRACT: Third-stage larvae (L3s) of the canine hookworm, Ancylostoma caninum, endure a period of arrested development preceding transmission to a host. Many of the mRNAs up-regulated at this stage are likely to encode proteins that act at the host-parasite interface and facilitate the transition from a free-living to a parasitic larva. The initial phase of the infection of a mammalian host by A. caninum L3s (herein termed “activation”) can be mimicked in vitro by culturing L3s in serum-containing medium. The mRNAs differentially transcribed between activated and non-activated L3s were identified by suppression subtractive hybridisation (SSH). The analysis of these mRNAs on a custom oligonucleotide microarray printed with the SSH ESTs and publicly available A. caninum ESTs (non-subtracted) yielded a total of 602 differentially expressed mRNAs, of which the most highly represented sequences (27) encoded products belonging to the pathogenesis-related protein (PRP) superfamily and different mechanistic classes of proteases. Comparison of these A. caninum mRNAs with those of Caenorhabditis elegans larvae exiting from developmental (dauer) arrest demonstrated unexpectedly large differences with respect to gene ontology profiles. C. elegans L3 exiting developmental arrest up-regulated the expression of collagens and other (mostly intracellular) molecules involved in growth and development. Such mRNAs are virtually absent from activated hookworm larvae, and instead are represented by an inordinately large number of mRNAs encoding extracellular proteins, suggesting that many of the activation-associated hookworm mRNAs are involved in host-parasite interactions. The near absence of mRNAs associated with reproduction, growth and development among activated hookworm L3s probably reflects their ability to further arrest (i.e. undergo hypobiosis) in tissues of non-permissive hosts or in the external environment when conditions for transmission are unfavourable. Although this should not necessarily invalidate C. elegans dauer exit as a model for hookworm activation, it provides substantial information on the limitations of this free-living nematode as a model organism for the transition of nematode larvae from a free-living to a parasitic state. Keywords: Comparative transcriptomic hybridisation

ORGANISM(S): Ancylostoma caninum

PROVIDER: GSE8155 | GEO | 2007/08/15

SECONDARY ACCESSION(S): PRJNA101039

REPOSITORIES: GEO

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Project description:Many tissue-specific stem cells maintain the ability to produce multiple cell types during long periods of non-division, or quiescence. FOXO transcription factors promote quiescence and stem cell maintenance, but the mechanisms by which FOXO proteins promote multipotency during quiescence are still emerging. The single FOXO ortholog in C. elegans, daf-16, promotes entry into a quiescent and stress-resistant larval stage called dauer in response to adverse environmental cues. During dauer, stem and progenitor cells maintain or re-establish multipotency to allow normal development to resume after dauer. We find that during dauer, daf-16/FOXO prevents epidermal stem cells (seam cells) from prematurely adopting differentiated, adult characteristics. In particular, dauer larvae that lack daf-16 misexpress collagens that are normally adult-enriched. Using col-19p::gfp as an adult cell fate marker, we find that all major daf-16 isoforms contribute to opposing col-19p::gfp expression during dauer. By contrast, daf-16(0) larvae that undergo non-dauer development do not misexpress col-19p::gfp. Adult cell fate and the timing of col-19p::gfp expression are regulated by the heterochronic gene network, including lin-41 and lin-29. lin-41 encodes an RNA-binding protein orthologous to LIN41/TRIM71 in mammals, and lin-29 encodes a conserved zinc finger transcription factor. In non-dauer development lin-41 opposes adult cell fate by inhibiting the translation of lin-29, which directly activates col-19 transcription and promotes adult cell fate. We find that during dauer, lin-41 blocks col-19p::gfp expression, but surprisingly, lin-29 is not required in this context. Additionally, daf-16 promotes the expression of lin-41 in dauer larvae. The col-19p::gfp misexpression phenotype observed in dauer larvae with reduced daf-16 requires the downregulation of lin-41, but does not require lin-29. Taken together, this work demonstrates a novel role for daf-16/FOXO as a heterochronic gene that promotes expression of lin-41/TRIM71 to contribute to multipotent cell fate in a quiescent stem cell model.

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Identification of mRNAs that are differentially expressed between serum-stimulated and non-stimulated A. caninum L3

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