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Expression of Ciona intestinalis variable region-containing chitin-binding proteins during development of the gastrointestinal tract and their role in host-microbe interactions.

Liberti A, Melillo D, Zucchetti I, Natale L, Dishaw LJ, Litman GW, De Santis R, Pinto MR - PLoS ONE (2014)

Bottom Line: The expression of VCBP-A and -C is detected first in discrete areas of larva endoderm and becomes progressively localized during differentiation in the stomach and intestine, marking the development of gut tracts.In "small adults" (1-2 cm juveniles) expression of VCBP-C persists and VCBP-A gradually diminishes, ultimately replaced by expression of VCBP-B.A potential role for VCBPs in gut-microbiota interactions and homeostasis is indicated.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Physiology and Evolution, Stazione Zoologica Anton Dohrn, Villa Comunale, Napoli, Italy.

ABSTRACT
Variable region-containing chitin-binding proteins (VCBPs) are secreted, immune-type molecules that have been described in both amphioxus, a cephalochordate, and sea squirt, Ciona intestinalis, a urochordate. In adult Ciona, VCBP-A, -B and -C are expressed in hemocytes and the cells of the gastrointestinal tract. VCBP-C binds bacteria in the stomach lumen and functions as an opsonin in vitro. In the present paper the expression of VCBPs has been characterized during development using in situ hybridization, immunohistochemical staining and quantitative polymerase chain reaction (qPCR) technologies. The expression of VCBP-A and -C is detected first in discrete areas of larva endoderm and becomes progressively localized during differentiation in the stomach and intestine, marking the development of gut tracts. In "small adults" (1-2 cm juveniles) expression of VCBP-C persists and VCBP-A gradually diminishes, ultimately replaced by expression of VCBP-B. The expression of VCBP-A and -C in stage 7-8 juveniles, at which point animals have already started feeding, is influenced significantly by challenge with either Gram-positive or -negative bacteria. A potential role for VCBPs in gut-microbiota interactions and homeostasis is indicated.

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Immunogold localization of VCBP-A by anti-VCBP-A antibody in adult stomach sections.In stomach epithelium VCBP-A is detected in both the cytoplasm and within large vacuoles of the absorptive cells (A and B). In the lamina propria, gold particles are localized in the granular amoebocytes (C and D). In some instance, amoebocytes expressing VCBP-A are found associated with the basement membrane of the epithelium (C) and apposed to the plasma membrane of the epithelial cells (D); gold particles also are present in the basal rim of the plasma membrane of the stomach epithelium (D). (B) Higher magnification of the area marked in (A). bm, basement membrane; cy, cytoplasm; ga, granular amoebocyte; v, vacuole; arrowheads, gold particles; asterisk, lamina propria. (Scale bars: A, C and D, 1 µm; B, 0,5 µm).
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pone-0094984-g009: Immunogold localization of VCBP-A by anti-VCBP-A antibody in adult stomach sections.In stomach epithelium VCBP-A is detected in both the cytoplasm and within large vacuoles of the absorptive cells (A and B). In the lamina propria, gold particles are localized in the granular amoebocytes (C and D). In some instance, amoebocytes expressing VCBP-A are found associated with the basement membrane of the epithelium (C) and apposed to the plasma membrane of the epithelial cells (D); gold particles also are present in the basal rim of the plasma membrane of the stomach epithelium (D). (B) Higher magnification of the area marked in (A). bm, basement membrane; cy, cytoplasm; ga, granular amoebocyte; v, vacuole; arrowheads, gold particles; asterisk, lamina propria. (Scale bars: A, C and D, 1 µm; B, 0,5 µm).

Mentions: In order to clarify why VCBP-A protein could be detected in an as yet unidentified stomach cell, in which VCBP-A transcripts were not detected by ISH, IHC analysis was carried out at the ultrastructural level. VCBP-A protein can be localized to the cytoplasm and within the large vacuoles of the absorptive cells (Fig. 9A and B). VCBP-A also could be detected in the lamina propria, both as a secreted molecule and in the granular amoebocytes populating this compartment (Fig. 9C). Quite often this type of hemocyte is associated with the basement membrane of the stomach cells and apposed to their plasma membrane (Fig. 9C and D). In these observation fields, VCBP-A expression also is detected in the basal rim of the plasma membrane and in the adjacent cytoplasmic areas of the epithelial cells (Fig. 9C and D).


Expression of Ciona intestinalis variable region-containing chitin-binding proteins during development of the gastrointestinal tract and their role in host-microbe interactions.

Liberti A, Melillo D, Zucchetti I, Natale L, Dishaw LJ, Litman GW, De Santis R, Pinto MR - PLoS ONE (2014)

Immunogold localization of VCBP-A by anti-VCBP-A antibody in adult stomach sections.In stomach epithelium VCBP-A is detected in both the cytoplasm and within large vacuoles of the absorptive cells (A and B). In the lamina propria, gold particles are localized in the granular amoebocytes (C and D). In some instance, amoebocytes expressing VCBP-A are found associated with the basement membrane of the epithelium (C) and apposed to the plasma membrane of the epithelial cells (D); gold particles also are present in the basal rim of the plasma membrane of the stomach epithelium (D). (B) Higher magnification of the area marked in (A). bm, basement membrane; cy, cytoplasm; ga, granular amoebocyte; v, vacuole; arrowheads, gold particles; asterisk, lamina propria. (Scale bars: A, C and D, 1 µm; B, 0,5 µm).
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Related In: Results  -  Collection

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pone-0094984-g009: Immunogold localization of VCBP-A by anti-VCBP-A antibody in adult stomach sections.In stomach epithelium VCBP-A is detected in both the cytoplasm and within large vacuoles of the absorptive cells (A and B). In the lamina propria, gold particles are localized in the granular amoebocytes (C and D). In some instance, amoebocytes expressing VCBP-A are found associated with the basement membrane of the epithelium (C) and apposed to the plasma membrane of the epithelial cells (D); gold particles also are present in the basal rim of the plasma membrane of the stomach epithelium (D). (B) Higher magnification of the area marked in (A). bm, basement membrane; cy, cytoplasm; ga, granular amoebocyte; v, vacuole; arrowheads, gold particles; asterisk, lamina propria. (Scale bars: A, C and D, 1 µm; B, 0,5 µm).
Mentions: In order to clarify why VCBP-A protein could be detected in an as yet unidentified stomach cell, in which VCBP-A transcripts were not detected by ISH, IHC analysis was carried out at the ultrastructural level. VCBP-A protein can be localized to the cytoplasm and within the large vacuoles of the absorptive cells (Fig. 9A and B). VCBP-A also could be detected in the lamina propria, both as a secreted molecule and in the granular amoebocytes populating this compartment (Fig. 9C). Quite often this type of hemocyte is associated with the basement membrane of the stomach cells and apposed to their plasma membrane (Fig. 9C and D). In these observation fields, VCBP-A expression also is detected in the basal rim of the plasma membrane and in the adjacent cytoplasmic areas of the epithelial cells (Fig. 9C and D).

Bottom Line: The expression of VCBP-A and -C is detected first in discrete areas of larva endoderm and becomes progressively localized during differentiation in the stomach and intestine, marking the development of gut tracts.In "small adults" (1-2 cm juveniles) expression of VCBP-C persists and VCBP-A gradually diminishes, ultimately replaced by expression of VCBP-B.A potential role for VCBPs in gut-microbiota interactions and homeostasis is indicated.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Physiology and Evolution, Stazione Zoologica Anton Dohrn, Villa Comunale, Napoli, Italy.

ABSTRACT
Variable region-containing chitin-binding proteins (VCBPs) are secreted, immune-type molecules that have been described in both amphioxus, a cephalochordate, and sea squirt, Ciona intestinalis, a urochordate. In adult Ciona, VCBP-A, -B and -C are expressed in hemocytes and the cells of the gastrointestinal tract. VCBP-C binds bacteria in the stomach lumen and functions as an opsonin in vitro. In the present paper the expression of VCBPs has been characterized during development using in situ hybridization, immunohistochemical staining and quantitative polymerase chain reaction (qPCR) technologies. The expression of VCBP-A and -C is detected first in discrete areas of larva endoderm and becomes progressively localized during differentiation in the stomach and intestine, marking the development of gut tracts. In "small adults" (1-2 cm juveniles) expression of VCBP-C persists and VCBP-A gradually diminishes, ultimately replaced by expression of VCBP-B. The expression of VCBP-A and -C in stage 7-8 juveniles, at which point animals have already started feeding, is influenced significantly by challenge with either Gram-positive or -negative bacteria. A potential role for VCBPs in gut-microbiota interactions and homeostasis is indicated.

Show MeSH