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New insights into the apoptotic process in mollusks: characterization of caspase genes in Mytilus galloprovincialis.

Romero A, Estévez-Calvar N, Dios S, Figueras A, Novoa B - PLoS ONE (2011)

Bottom Line: Caspase proteins constitute the core of the apoptotic machinery and can be categorized as either initiators or effectors of apoptosis.Evaluation of the genes' tissue expression patterns revealed extremely high expression levels within the gland and gills, where the apoptotic process is highly active due to the clearance of damaged cells.Hemocytes also showed high expression values, probably due to of the role of apoptosis in the defense against pathogens.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Investigaciones Marinas, Consejo Superior de Investigaciones Científicas, Vigo, Spain.

ABSTRACT
Apoptosis is an essential biological process in the development and maintenance of immune system homeostasis. Caspase proteins constitute the core of the apoptotic machinery and can be categorized as either initiators or effectors of apoptosis. Although the genes encoding caspase proteins have been described in vertebrates and in almost all invertebrate phyla, there are few reports describing the initiator and executioner caspases or the modulation of their expression by different stimuli in different apoptotic pathways in bivalves. In the present work, we characterized two initiator and four executioner caspases in the mussel Mytilus galloprovincialis. Both initiators and executioners showed structural features that make them different from other caspase proteins already described. Evaluation of the genes' tissue expression patterns revealed extremely high expression levels within the gland and gills, where the apoptotic process is highly active due to the clearance of damaged cells. Hemocytes also showed high expression values, probably due to of the role of apoptosis in the defense against pathogens. To understand the mechanisms of caspase gene regulation, hemocytes were treated with UV-light, environmental pollutants and pathogen-associated molecular patterns (PAMPs) and apoptosis was evaluated by microscopy, flow cytometry and qPCR techniques. Our results suggest that the apoptotic process could be tightly regulated in bivalve mollusks by overexpression/suppression of caspase genes; additionally, there is evidence of caspase-specific responses to pathogens and pollutants. The apoptotic process in mollusks has a similar complexity to that of vertebrates, but presents unique features that may be related to recurrent exposure to environmental changes, pollutants and pathogens imposed by their sedentary nature.

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Related in: MedlinePlus

Induction of caspase genes in hemocytes treated with different PAMPs at 1, 3 and 6 h pt.Results represent the mean ± SD of 4 experimental hemocyte pools. Data were analyzed using the Student’s t-test. a, b, c, d, e and f indicate significant differences (p<0.05) of caspase-2, -8, 3/7-1, -3/7-2, -3/7-3 and -3/7-4, respectively.
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pone-0017003-g009: Induction of caspase genes in hemocytes treated with different PAMPs at 1, 3 and 6 h pt.Results represent the mean ± SD of 4 experimental hemocyte pools. Data were analyzed using the Student’s t-test. a, b, c, d, e and f indicate significant differences (p<0.05) of caspase-2, -8, 3/7-1, -3/7-2, -3/7-3 and -3/7-4, respectively.

Mentions: In order to analyze the implication of caspases in the pathogen-induced immune response, hemocytes were treated with various PAMPs (Figure 9). The PAMPs that mimicked bacterial infections (LPS, CPGs and LTA) were able to modulate caspase gene expression. LPS and CPGs induced a less than 3.5-fold change in gene expression. Initiator caspase-2 and -8 were significantly down-regulated after 3 h pt for all treatments. By the end of the experiment (6 h pt) LPS and LTA treatments had induced a significant up-regulation of the caspase-2 gene, whereas the CPGs treatment significantly up-regulated caspase-8 expression. Executioner caspase-3/7-1 and -3/7-2 followed similar kinetics for all treatments. LPS and LTA induced an increase of both caspases during the time course, caspase-3/7-2 expression reaching levels significantly different than controls as early as 3 h pt; caspase-3/7-1 was not significantly up-regulated in LTA treated samples at the 3 h pt time point. During CPGs treatment, the expression levels of caspase-3/7-1 and -3/7-2 peaked after 3 h pt followed by a significant decrease of the expression after 6 h pt. Caspase-3/7-3 and -3/7-4 were less affected by the treatments during the time course. Caspase-3/7-3 was consistently significantly down-regulated after 3 h pt in all treatments, recovering initial expression values at 6 h pt in LPS and LTA treatments or reaching maximum values in CPGs treatment. No significant changes were recorded for caspase-3/7-4 expression.


New insights into the apoptotic process in mollusks: characterization of caspase genes in Mytilus galloprovincialis.

Romero A, Estévez-Calvar N, Dios S, Figueras A, Novoa B - PLoS ONE (2011)

Induction of caspase genes in hemocytes treated with different PAMPs at 1, 3 and 6 h pt.Results represent the mean ± SD of 4 experimental hemocyte pools. Data were analyzed using the Student’s t-test. a, b, c, d, e and f indicate significant differences (p<0.05) of caspase-2, -8, 3/7-1, -3/7-2, -3/7-3 and -3/7-4, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3037946&req=5

pone-0017003-g009: Induction of caspase genes in hemocytes treated with different PAMPs at 1, 3 and 6 h pt.Results represent the mean ± SD of 4 experimental hemocyte pools. Data were analyzed using the Student’s t-test. a, b, c, d, e and f indicate significant differences (p<0.05) of caspase-2, -8, 3/7-1, -3/7-2, -3/7-3 and -3/7-4, respectively.
Mentions: In order to analyze the implication of caspases in the pathogen-induced immune response, hemocytes were treated with various PAMPs (Figure 9). The PAMPs that mimicked bacterial infections (LPS, CPGs and LTA) were able to modulate caspase gene expression. LPS and CPGs induced a less than 3.5-fold change in gene expression. Initiator caspase-2 and -8 were significantly down-regulated after 3 h pt for all treatments. By the end of the experiment (6 h pt) LPS and LTA treatments had induced a significant up-regulation of the caspase-2 gene, whereas the CPGs treatment significantly up-regulated caspase-8 expression. Executioner caspase-3/7-1 and -3/7-2 followed similar kinetics for all treatments. LPS and LTA induced an increase of both caspases during the time course, caspase-3/7-2 expression reaching levels significantly different than controls as early as 3 h pt; caspase-3/7-1 was not significantly up-regulated in LTA treated samples at the 3 h pt time point. During CPGs treatment, the expression levels of caspase-3/7-1 and -3/7-2 peaked after 3 h pt followed by a significant decrease of the expression after 6 h pt. Caspase-3/7-3 and -3/7-4 were less affected by the treatments during the time course. Caspase-3/7-3 was consistently significantly down-regulated after 3 h pt in all treatments, recovering initial expression values at 6 h pt in LPS and LTA treatments or reaching maximum values in CPGs treatment. No significant changes were recorded for caspase-3/7-4 expression.

Bottom Line: Caspase proteins constitute the core of the apoptotic machinery and can be categorized as either initiators or effectors of apoptosis.Evaluation of the genes' tissue expression patterns revealed extremely high expression levels within the gland and gills, where the apoptotic process is highly active due to the clearance of damaged cells.Hemocytes also showed high expression values, probably due to of the role of apoptosis in the defense against pathogens.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Investigaciones Marinas, Consejo Superior de Investigaciones Científicas, Vigo, Spain.

ABSTRACT
Apoptosis is an essential biological process in the development and maintenance of immune system homeostasis. Caspase proteins constitute the core of the apoptotic machinery and can be categorized as either initiators or effectors of apoptosis. Although the genes encoding caspase proteins have been described in vertebrates and in almost all invertebrate phyla, there are few reports describing the initiator and executioner caspases or the modulation of their expression by different stimuli in different apoptotic pathways in bivalves. In the present work, we characterized two initiator and four executioner caspases in the mussel Mytilus galloprovincialis. Both initiators and executioners showed structural features that make them different from other caspase proteins already described. Evaluation of the genes' tissue expression patterns revealed extremely high expression levels within the gland and gills, where the apoptotic process is highly active due to the clearance of damaged cells. Hemocytes also showed high expression values, probably due to of the role of apoptosis in the defense against pathogens. To understand the mechanisms of caspase gene regulation, hemocytes were treated with UV-light, environmental pollutants and pathogen-associated molecular patterns (PAMPs) and apoptosis was evaluated by microscopy, flow cytometry and qPCR techniques. Our results suggest that the apoptotic process could be tightly regulated in bivalve mollusks by overexpression/suppression of caspase genes; additionally, there is evidence of caspase-specific responses to pathogens and pollutants. The apoptotic process in mollusks has a similar complexity to that of vertebrates, but presents unique features that may be related to recurrent exposure to environmental changes, pollutants and pathogens imposed by their sedentary nature.

Show MeSH
Related in: MedlinePlus