Functional conservation of the apoptotic machinery from coral to man: the diverse and complex Bcl-2 and caspase repertoires of Acropora millepora.

BACKGROUND: Apoptotic cell death is a defining and ubiquitous characteristic of metazoans, but its evolutionary origins are unclear. Although Caenorhabditis and Drosophila played key roles in establishing the molecular bases of apoptosis, it is now clear that cell death pathways of these animals do not reflect ancestral characteristics. Conversely, recent work suggests that the apoptotic networks of cnidarians may be complex and vertebrate-like, hence characterization of the apoptotic complement of representatives of the basal cnidarian class Anthozoa will help us to understand the evolution of the vertebrate apoptotic network. RESULTS: We describe the Bcl-2 and caspase protein repertoires of the coral Acropora millepora, making use of the comprehensive transcriptomic data available for this species. Molecular phylogenetics indicates that some Acropora proteins are orthologs of specific mammalian pro-apoptotic Bcl-2 family members, but the relationships of other Bcl-2 and caspases are unclear. The pro- or anti-apoptotic activities of coral Bcl-2 proteins were investigated by expression in mammalian cells, and the results imply functional conservation of the effector/anti-apoptotic machinery despite limited sequence conservation in the anti-apoptotic Bcl-2 proteins. A novel caspase type ("Caspase-X"), containing both inactive and active caspase domains, was identified in Acropora and appears to be restricted to corals. When expressed in mammalian cells, full-length caspase-X caused loss of viability, and a truncated version containing only the active domain was more effective in inducing cell death, suggesting that the inactive domain might modulate activity in the full-length protein. Structure prediction suggests that the active and inactive caspase domains in caspase-X are likely to interact, resulting in a structure resembling that of the active domain in procaspase-8 and the inactive caspase domain in the mammalian c-FLIP anti-apoptotic factor. CONCLUSIONS: The data presented here confirm that many of the basic mechanisms involved in both the intrinsic and extrinsic apoptotic pathways were in place in the common ancestor of cnidarians and bilaterians. With the identification of most or all of the repertoires of coral Bcl-2 and caspases, our results not only provide new perspectives on the evolution of apoptotic pathways, but also a framework for future experimental studies towards a complete understanding of coral bleaching mechanisms, in which apoptotic cell death might be involved.

The acute transcriptional response of the coral Acropora millepora to immune challenge: expression of GiMAP/IAN genes links the innate immune responses of corals with those of mammals and plants.

BACKGROUND: As a step towards understanding coral immunity we present the first whole transcriptome analysis of the acute responses of Acropora millepora to challenge with the bacterial cell wall derivative MDP and the viral mimic poly I:C, defined immunogens provoking distinct but well characterised responses in higher animals. RESULTS: These experiments reveal similarities with the responses both of arthropods and mammals, as well as coral-specific effects. The most surprising finding was that MDP specifically induced three members of the GiMAP gene family, which has been implicated in immunity in mammals but is absent from Drosophila and Caenorhabditis. Like their mammalian homologs, GiMAP genes are arranged in a tandem cluster in the coral genome. CONCLUSIONS: A phylogenomic survey of this gene family implies ancient origins, multiple independent losses and lineage-specific expansions during animal evolution. Whilst functional convergence cannot be ruled out, GiMAP expression in corals may reflect an ancestral role in immunity, perhaps in phagolysosomal processing.

Defining the origins of the NOD-like receptor system at the base of animal evolution.

Distinguishing self from nonself and the onset of defense effector mechanisms upon recognition of pathogens are essential for the survival of all life forms in the animal kingdom. The family of nucleotide -binding and oligomeriszation domain-like receptors (NLRs) was first identified in vertebrates and comprises a group of pivotal sensor protein of the innate immune system for microbial cell wall components or danger signals. Here, we provide first evidence that early diverging metazoans have large and complex NLR repertoires. The cnidarian NACHT/NB-ARC genes include novel combinations of domains, and the number of one specific type (NB-ARC and tetratricopeptide repeat containing) in Hydra is particularly large. We characterize the transcript structure and expression patterns of a selected HyNLR, HyNLR type 1 and describe putative interaction partners. In a heterologous expression system, we show induced proximity recruitment of an effector caspase (HyDD-Caspase) to the HyNLR type 1 protein upon oligomerization indicating a potential role of caspase activation downstream of NLR activation in Hydra. These results add substantially to our understanding of the ancestral innate immune repertoire as well as providing the first insights into putative cytoplasmic defense mechanisms at the base of animal evolution.

Skeletal abnormalities and extra-skeletal ossification in mice with restricted Gsalpha deletion caused by a renin promoter-Cre transgene.

We have recently generated a transgenic mouse line (termed hRen-Cre) that expresses Cre-recombinase under the control of a 12.2-kb fragment of the human renin promoter. In the present study, we have crossed hRen-Cre mice with a mouse strain in which exon 1 of the Gnas gene is flanked by loxP sites. Gnas encodes the alpha-subunit of the stimulatory G protein (Gs alpha). Our aim has been to generate a mouse model with locally restricted inactivation of Gs alpha to extend studies of the role of Gs alpha function in vivo. Mice with local Cre-mediated inactivation of Gs alpha (rCre-Gs alpha) are viable and fertile. Their most obvious phenotype consists of marked skeletal malformations of the forelimbs in which computer-tomography scans reveal shortened and fused extremity bones. Extraskeletal ossifications occur in the subcutis and in skeletal muscles associated with the affected long bones. Plasma calcium, phosphate and parathyroid hormone are normal. Skin histology has demonstrated diffuse mineralization and ossification associated with the basal cells of hair follicles. This phenotype in part resembles syndromes in humans associated with loss-of-function of Gs alpha, such as Albright hereditary osteodystrophy and progressive osseous heteroplasia. The renal phenotype of rCre-Gs alpha mice is inconspicuous. Plasma renin concentration, ambient urine osmolarity, and the glomerular filtration rate of rCre-Gs alpha mice do not differ from controls. The absence of measurable functional changes in the renin-angiotensin system indicates insufficient Cre expression in juxtaglomerular granular cells in this strain of mice. Nevertheless, the present report reaffirms the importance of Gs alpha signaling for bone development and the suppression of ectopic ossification.