Limits...
Participation of actin on Giardia lamblia growth and encystation.

Castillo-Romero A, Leon-Avila G, Perez Rangel A, Cortes Zarate R, Garcia Tovar C, Hernandez JM - PLoS ONE (2009)

Bottom Line: All drugs caused a growth reduction (27 to 45%) and changes on the distribution of actin.Furthermore, the drugs caused a significant reduction of the intensity of fluorescence-labeled CWP1 on ESV and on cyst wall, this was coincident with a reduction of CWP1 gene expression (34%).All our results, indicated an important role of actin in the morphology, growth and encystation and indirectly suggested an actin role in gene expression.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biologia Celular, CINVESTAV-IPN, Mexico Distrito Federal, Mexico.

ABSTRACT

Background: Microfilaments play a determinant role in different cell processes such as: motility, cell division, phagocytosis and intracellular transport; however, these structures are poorly understood in the parasite Giardia lamblia.

Methodology and principal findings: By confocal microscopy using TRITC-phalloidin, we found structured actin distributed in the entire trophozoite, the label stand out at the ventral disc, median body, flagella and around the nuclei. During Giardia encystation, a sequence of morphological changes concurrent to modifications on the distribution of structured actin and in the expression of actin mRNA were observed. To elucidate whether actin participates actively on growth and encystation, cells were treated with Cytochalasin D, Latrunculin A and Jasplakinolide and analyzed by confocal and scanning electron microscopy. All drugs caused a growth reduction (27 to 45%) and changes on the distribution of actin. Besides, 60 to 80% of trophozoites treated with the drugs, exhibited damage at the caudal region, alterations in the flagella and wrinkles-like on the plasma membrane. The drugs also altered the cyst-yield and the morphology, scanning electron microscopy revealed diminished cytokinesis, cysts with damages in the wall and alterations in the size and on the intermembranal space. Furthermore, the drugs caused a significant reduction of the intensity of fluorescence-labeled CWP1 on ESV and on cyst wall, this was coincident with a reduction of CWP1 gene expression (34%).

Conclusions and significance: All our results, indicated an important role of actin in the morphology, growth and encystation and indirectly suggested an actin role in gene expression.

Show MeSH

Related in: MedlinePlus

Localization of actin in Giardia lamblia trophozoites and cysts.The cells were labeled using TRITC-phalloidin and analyzed by confocal microscopy. A, DIC image of trophozoites. B, Trophozoites stained with TRITC-phalloidin (d = ventral disc; m = median body; n = nuclei; and, f = flagella). C, DIC image of cysts. D, Cysts stained with TRITC-phalloidin. E and F, optical slices of D. The B inset represents an optical slice. The nuclei were labelled with DAPI.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2743995&req=5

pone-0007156-g001: Localization of actin in Giardia lamblia trophozoites and cysts.The cells were labeled using TRITC-phalloidin and analyzed by confocal microscopy. A, DIC image of trophozoites. B, Trophozoites stained with TRITC-phalloidin (d = ventral disc; m = median body; n = nuclei; and, f = flagella). C, DIC image of cysts. D, Cysts stained with TRITC-phalloidin. E and F, optical slices of D. The B inset represents an optical slice. The nuclei were labelled with DAPI.

Mentions: In order to detect actin in both stages of Giardia, we first analyzed its distribution using TRITC-phalloidin –a well accepted specific tool for structured actin identification–. Confocal microscopy images of trophozoites (Figure 1A) demonstrated actin distributed like small spots in the entire trophozoites and in nuclei. However an intense stain was evident, in median body, on the periphery of the ventral disc and flagella (Figure 1B). In cysts (Figure 1C), actin was distributed as thick ribbons forming a compact oval structure, also some patches were evidents (Figure 1D–F).


Participation of actin on Giardia lamblia growth and encystation.

Castillo-Romero A, Leon-Avila G, Perez Rangel A, Cortes Zarate R, Garcia Tovar C, Hernandez JM - PLoS ONE (2009)

Localization of actin in Giardia lamblia trophozoites and cysts.The cells were labeled using TRITC-phalloidin and analyzed by confocal microscopy. A, DIC image of trophozoites. B, Trophozoites stained with TRITC-phalloidin (d = ventral disc; m = median body; n = nuclei; and, f = flagella). C, DIC image of cysts. D, Cysts stained with TRITC-phalloidin. E and F, optical slices of D. The B inset represents an optical slice. The nuclei were labelled with DAPI.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2743995&req=5

pone-0007156-g001: Localization of actin in Giardia lamblia trophozoites and cysts.The cells were labeled using TRITC-phalloidin and analyzed by confocal microscopy. A, DIC image of trophozoites. B, Trophozoites stained with TRITC-phalloidin (d = ventral disc; m = median body; n = nuclei; and, f = flagella). C, DIC image of cysts. D, Cysts stained with TRITC-phalloidin. E and F, optical slices of D. The B inset represents an optical slice. The nuclei were labelled with DAPI.
Mentions: In order to detect actin in both stages of Giardia, we first analyzed its distribution using TRITC-phalloidin –a well accepted specific tool for structured actin identification–. Confocal microscopy images of trophozoites (Figure 1A) demonstrated actin distributed like small spots in the entire trophozoites and in nuclei. However an intense stain was evident, in median body, on the periphery of the ventral disc and flagella (Figure 1B). In cysts (Figure 1C), actin was distributed as thick ribbons forming a compact oval structure, also some patches were evidents (Figure 1D–F).

Bottom Line: All drugs caused a growth reduction (27 to 45%) and changes on the distribution of actin.Furthermore, the drugs caused a significant reduction of the intensity of fluorescence-labeled CWP1 on ESV and on cyst wall, this was coincident with a reduction of CWP1 gene expression (34%).All our results, indicated an important role of actin in the morphology, growth and encystation and indirectly suggested an actin role in gene expression.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biologia Celular, CINVESTAV-IPN, Mexico Distrito Federal, Mexico.

ABSTRACT

Background: Microfilaments play a determinant role in different cell processes such as: motility, cell division, phagocytosis and intracellular transport; however, these structures are poorly understood in the parasite Giardia lamblia.

Methodology and principal findings: By confocal microscopy using TRITC-phalloidin, we found structured actin distributed in the entire trophozoite, the label stand out at the ventral disc, median body, flagella and around the nuclei. During Giardia encystation, a sequence of morphological changes concurrent to modifications on the distribution of structured actin and in the expression of actin mRNA were observed. To elucidate whether actin participates actively on growth and encystation, cells were treated with Cytochalasin D, Latrunculin A and Jasplakinolide and analyzed by confocal and scanning electron microscopy. All drugs caused a growth reduction (27 to 45%) and changes on the distribution of actin. Besides, 60 to 80% of trophozoites treated with the drugs, exhibited damage at the caudal region, alterations in the flagella and wrinkles-like on the plasma membrane. The drugs also altered the cyst-yield and the morphology, scanning electron microscopy revealed diminished cytokinesis, cysts with damages in the wall and alterations in the size and on the intermembranal space. Furthermore, the drugs caused a significant reduction of the intensity of fluorescence-labeled CWP1 on ESV and on cyst wall, this was coincident with a reduction of CWP1 gene expression (34%).

Conclusions and significance: All our results, indicated an important role of actin in the morphology, growth and encystation and indirectly suggested an actin role in gene expression.

Show MeSH
Related in: MedlinePlus