Limits...
Evidence for a "wattle and daub" model of the cyst wall of entamoeba.

Chatterjee A, Ghosh SK, Jang K, Bullitt E, Moore L, Robbins PW, Samuelson J - PLoS Pathog. (2009)

Bottom Line: Here we show chitin, which was detected with wheat germ agglutinin, is made in secretory vesicles prior to its deposition on the surface of encysting Ei.Jacob lectins, which have tandemly arrayed chitin-binding domains (CBDs), and chitinase, which has an N-terminal CBD, were each made early during encystation.These findings provide evidence for a "wattle and daub" model of the Entamoeba cyst wall, where the wattle or sticks (chitin fibrils likely cross-linked by Jacob lectins) is constructed prior to the addition of the mortar or daub (Jessie3 lectins).

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA, USA.

ABSTRACT
The cyst wall of Entamoeba invadens (Ei), a model for the human pathogen Entamoeba histolytica, is composed of fibrils of chitin and three chitin-binding lectins called Jacob, Jessie3, and chitinase. Here we show chitin, which was detected with wheat germ agglutinin, is made in secretory vesicles prior to its deposition on the surface of encysting Ei. Jacob lectins, which have tandemly arrayed chitin-binding domains (CBDs), and chitinase, which has an N-terminal CBD, were each made early during encystation. These results are consistent with their hypothesized roles in cross-linking chitin fibrils (Jacob lectins) and remodeling the cyst wall (chitinase). Jessie3 lectins likely form the mortar or daub of the cyst wall, because 1) Jessie lectins were made late during encystation; 2) the addition to Jessie lectins to the cyst wall correlated with a marked decrease in the permeability of cysts to nucleic acid stains (DAPI) and actin-binding heptapeptide (phalloidin); and 3) recombinant Jessie lectins, expressed as a maltose-binding proteins in the periplasm of Escherichia coli, caused transformed bacteria to agglutinate in suspension and form a hard pellet that did not dissociate after centrifugation. Jessie3 appeared as linear forms and rosettes by negative staining of secreted recombinant proteins. These findings provide evidence for a "wattle and daub" model of the Entamoeba cyst wall, where the wattle or sticks (chitin fibrils likely cross-linked by Jacob lectins) is constructed prior to the addition of the mortar or daub (Jessie3 lectins).

Show MeSH

Related in: MedlinePlus

Three-dimensional high-resolution fluorescence microscopy shows that coincident with the addition of Jessie lectins (red), cyst walls become impermeable to DAPI (blue) and phalloidin (green).(A to C) After 72 hrs encystation, some Ei are still ameboid in appearance, some have relatively little Jessie3 lectin in the wall, and some protists have abundant Jessie3. DAPI and phalloidin stain well the immature cysts (Imm.) but fail to penetrate mature cysts. (D to F) In contrast, DAPI and phalloidin penetrate all cysts that have been frozen and thawed prior to staining with these reagents. Bar is 10 microns. Each image is a single optical section.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2698119&req=5

ppat-1000498-g004: Three-dimensional high-resolution fluorescence microscopy shows that coincident with the addition of Jessie lectins (red), cyst walls become impermeable to DAPI (blue) and phalloidin (green).(A to C) After 72 hrs encystation, some Ei are still ameboid in appearance, some have relatively little Jessie3 lectin in the wall, and some protists have abundant Jessie3. DAPI and phalloidin stain well the immature cysts (Imm.) but fail to penetrate mature cysts. (D to F) In contrast, DAPI and phalloidin penetrate all cysts that have been frozen and thawed prior to staining with these reagents. Bar is 10 microns. Each image is a single optical section.

Mentions: Unlike encysting Ei in Figs. 1, 2, and 4, protists here were labeled with antibodies to Jessie3 prior to fixation, so that only the surfaces of the encysting parasites are labeled. While there is very little Jessie3 lectin on the parasite surface after 36 hrs (A), Jessie3 appears in increasing number of punctate spots, sometimes in linear arrays (arrows), in the cyst wall at 48 hrs (B) and 60 hrs (C). In contrast, at 72 hrs (D), Jessie3 has a swirling appearance in the Ei cyst wall. After Jessie3 is fully incorporated into the wall (D), the cyst becomes impermeable to DAPI, so that nuclei are no longer visible or are weakly visible. Bar is 10 microns. Each image is the composite of multiple optical sections.


Evidence for a "wattle and daub" model of the cyst wall of entamoeba.

Chatterjee A, Ghosh SK, Jang K, Bullitt E, Moore L, Robbins PW, Samuelson J - PLoS Pathog. (2009)

Three-dimensional high-resolution fluorescence microscopy shows that coincident with the addition of Jessie lectins (red), cyst walls become impermeable to DAPI (blue) and phalloidin (green).(A to C) After 72 hrs encystation, some Ei are still ameboid in appearance, some have relatively little Jessie3 lectin in the wall, and some protists have abundant Jessie3. DAPI and phalloidin stain well the immature cysts (Imm.) but fail to penetrate mature cysts. (D to F) In contrast, DAPI and phalloidin penetrate all cysts that have been frozen and thawed prior to staining with these reagents. Bar is 10 microns. Each image is a single optical section.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2698119&req=5

ppat-1000498-g004: Three-dimensional high-resolution fluorescence microscopy shows that coincident with the addition of Jessie lectins (red), cyst walls become impermeable to DAPI (blue) and phalloidin (green).(A to C) After 72 hrs encystation, some Ei are still ameboid in appearance, some have relatively little Jessie3 lectin in the wall, and some protists have abundant Jessie3. DAPI and phalloidin stain well the immature cysts (Imm.) but fail to penetrate mature cysts. (D to F) In contrast, DAPI and phalloidin penetrate all cysts that have been frozen and thawed prior to staining with these reagents. Bar is 10 microns. Each image is a single optical section.
Mentions: Unlike encysting Ei in Figs. 1, 2, and 4, protists here were labeled with antibodies to Jessie3 prior to fixation, so that only the surfaces of the encysting parasites are labeled. While there is very little Jessie3 lectin on the parasite surface after 36 hrs (A), Jessie3 appears in increasing number of punctate spots, sometimes in linear arrays (arrows), in the cyst wall at 48 hrs (B) and 60 hrs (C). In contrast, at 72 hrs (D), Jessie3 has a swirling appearance in the Ei cyst wall. After Jessie3 is fully incorporated into the wall (D), the cyst becomes impermeable to DAPI, so that nuclei are no longer visible or are weakly visible. Bar is 10 microns. Each image is the composite of multiple optical sections.

Bottom Line: Here we show chitin, which was detected with wheat germ agglutinin, is made in secretory vesicles prior to its deposition on the surface of encysting Ei.Jacob lectins, which have tandemly arrayed chitin-binding domains (CBDs), and chitinase, which has an N-terminal CBD, were each made early during encystation.These findings provide evidence for a "wattle and daub" model of the Entamoeba cyst wall, where the wattle or sticks (chitin fibrils likely cross-linked by Jacob lectins) is constructed prior to the addition of the mortar or daub (Jessie3 lectins).

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA, USA.

ABSTRACT
The cyst wall of Entamoeba invadens (Ei), a model for the human pathogen Entamoeba histolytica, is composed of fibrils of chitin and three chitin-binding lectins called Jacob, Jessie3, and chitinase. Here we show chitin, which was detected with wheat germ agglutinin, is made in secretory vesicles prior to its deposition on the surface of encysting Ei. Jacob lectins, which have tandemly arrayed chitin-binding domains (CBDs), and chitinase, which has an N-terminal CBD, were each made early during encystation. These results are consistent with their hypothesized roles in cross-linking chitin fibrils (Jacob lectins) and remodeling the cyst wall (chitinase). Jessie3 lectins likely form the mortar or daub of the cyst wall, because 1) Jessie lectins were made late during encystation; 2) the addition to Jessie lectins to the cyst wall correlated with a marked decrease in the permeability of cysts to nucleic acid stains (DAPI) and actin-binding heptapeptide (phalloidin); and 3) recombinant Jessie lectins, expressed as a maltose-binding proteins in the periplasm of Escherichia coli, caused transformed bacteria to agglutinate in suspension and form a hard pellet that did not dissociate after centrifugation. Jessie3 appeared as linear forms and rosettes by negative staining of secreted recombinant proteins. These findings provide evidence for a "wattle and daub" model of the Entamoeba cyst wall, where the wattle or sticks (chitin fibrils likely cross-linked by Jacob lectins) is constructed prior to the addition of the mortar or daub (Jessie3 lectins).

Show MeSH
Related in: MedlinePlus