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The Trypanosoma cruzi nucleic acid binding protein Tc38 presents changes in the intramitochondrial distribution during the cell cycle.

Duhagon MA, Pastro L, Sotelo-Silveira JR, Pérez-Díaz L, Maugeri D, Nardelli SC, Schenkman S, Williams N, Dallagiovanna B, Garat B - BMC Microbiol. (2009)

Bottom Line: However, we found that Tc38 predominantly localizes into the mitochondrion.In epimastigotes, Tc38 is found only in association with kDNA in G1 phase.In non-replicating parasite stages such as trypomastigotes, the protein is found only surrounding the entire kinetoplast structure.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratorio de Interacciones Moleculares, Facultad de Ciencias, Montevideo, Uruguay. mduhagon@fcien.edu.uy

ABSTRACT

Background: Tc38 of Trypanosoma cruzi has been isolated as a single stranded DNA binding protein with high specificity for the poly [dT-dG] sequence. It is present only in Kinetoplastidae protozoa and its sequence lacks homology to known functional domains. Tc38 orthologues present in Trypanosoma brucei and Leishmania were proposed to participate in quite different cellular processes. To further understand the function of this protein in Trypanosoma cruzi, we examined its in vitro binding to biologically relevant [dT-dG] enriched sequences, its expression and subcellular localization during the cell cycle and through the parasite life stages.

Results: By using specific antibodies, we found that Tc38 protein from epimastigote extracts participates in complexes with the poly [dT-dG] probe as well as with the universal minicircle sequence (UMS), a related repeated sequence found in maxicircle DNA, and the telomeric repeat. However, we found that Tc38 predominantly localizes into the mitochondrion. Though Tc38 is constitutively expressed through non-replicating and replicating life stages of T. cruzi, its subcellular localization in the unique parasite mitochondrion changes according to the cell cycle stage. In epimastigotes, Tc38 is found only in association with kDNA in G1 phase. From the S to G2 phase the protein localizes in two defined and connected spots flanking the kDNA. These spots disappear in late G2 turning into a diffuse dotted signal which extends beyond the kinetoplast. This later pattern is more evident in mitosis and cytokinesis. Finally, late in cytokinesis Tc38 reacquires its association with the kinetoplast. In non-replicating parasite stages such as trypomastigotes, the protein is found only surrounding the entire kinetoplast structure.

Conclusions: The dynamics of Tc38 subcellular localization observed during the cell cycle and life stages support a major role for Tc38 related to kDNA replication and maintenance.

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Related in: MedlinePlus

Tc38 patterns in T. cruzi metacyclic trypomastigotes and amastigotes. Phase contrast, DAPI staining and Tc38 signal are indicated. For the merge images, Tc38-Alexa 488 signal is shown in green and DAPI nucleic acid staining in blue. "N" indicates the nucleus and "K" indicates the kinetoplast. Selected metacyclic trypomastigotes and amastigotes that show the most frequent patterns observed are presented. Bars = 5 μm. The dotted box in the phase contrast corresponds to the position of the fluorescent images.
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Figure 7: Tc38 patterns in T. cruzi metacyclic trypomastigotes and amastigotes. Phase contrast, DAPI staining and Tc38 signal are indicated. For the merge images, Tc38-Alexa 488 signal is shown in green and DAPI nucleic acid staining in blue. "N" indicates the nucleus and "K" indicates the kinetoplast. Selected metacyclic trypomastigotes and amastigotes that show the most frequent patterns observed are presented. Bars = 5 μm. The dotted box in the phase contrast corresponds to the position of the fluorescent images.

Mentions: To further understand the putative role of Tc38, we compared the labeling pattern of replicative epimastigotes with proliferative amastigotes and non-proliferative metacyclic trypomastigotes (Figure 7). In the non-replicative metacyclic form, Tc38 is always found surrounding the kinetoplast. However, in the replicative amastigote, Tc38 showed a dynamic localization; whereas some amastigotes exhibit a widespread distribution that is not restricted to the kDNA (as seen in epimastigotes during the late G2, M and C phases), others present a homogenous Tc38 signal associated with the kDNA. Taken together these results suggest that Tc38 changes the internal localization pattern only in the replicative stages of T. cruzi life cycle.


The Trypanosoma cruzi nucleic acid binding protein Tc38 presents changes in the intramitochondrial distribution during the cell cycle.

Duhagon MA, Pastro L, Sotelo-Silveira JR, Pérez-Díaz L, Maugeri D, Nardelli SC, Schenkman S, Williams N, Dallagiovanna B, Garat B - BMC Microbiol. (2009)

Tc38 patterns in T. cruzi metacyclic trypomastigotes and amastigotes. Phase contrast, DAPI staining and Tc38 signal are indicated. For the merge images, Tc38-Alexa 488 signal is shown in green and DAPI nucleic acid staining in blue. "N" indicates the nucleus and "K" indicates the kinetoplast. Selected metacyclic trypomastigotes and amastigotes that show the most frequent patterns observed are presented. Bars = 5 μm. The dotted box in the phase contrast corresponds to the position of the fluorescent images.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2654453&req=5

Figure 7: Tc38 patterns in T. cruzi metacyclic trypomastigotes and amastigotes. Phase contrast, DAPI staining and Tc38 signal are indicated. For the merge images, Tc38-Alexa 488 signal is shown in green and DAPI nucleic acid staining in blue. "N" indicates the nucleus and "K" indicates the kinetoplast. Selected metacyclic trypomastigotes and amastigotes that show the most frequent patterns observed are presented. Bars = 5 μm. The dotted box in the phase contrast corresponds to the position of the fluorescent images.
Mentions: To further understand the putative role of Tc38, we compared the labeling pattern of replicative epimastigotes with proliferative amastigotes and non-proliferative metacyclic trypomastigotes (Figure 7). In the non-replicative metacyclic form, Tc38 is always found surrounding the kinetoplast. However, in the replicative amastigote, Tc38 showed a dynamic localization; whereas some amastigotes exhibit a widespread distribution that is not restricted to the kDNA (as seen in epimastigotes during the late G2, M and C phases), others present a homogenous Tc38 signal associated with the kDNA. Taken together these results suggest that Tc38 changes the internal localization pattern only in the replicative stages of T. cruzi life cycle.

Bottom Line: However, we found that Tc38 predominantly localizes into the mitochondrion.In epimastigotes, Tc38 is found only in association with kDNA in G1 phase.In non-replicating parasite stages such as trypomastigotes, the protein is found only surrounding the entire kinetoplast structure.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratorio de Interacciones Moleculares, Facultad de Ciencias, Montevideo, Uruguay. mduhagon@fcien.edu.uy

ABSTRACT

Background: Tc38 of Trypanosoma cruzi has been isolated as a single stranded DNA binding protein with high specificity for the poly [dT-dG] sequence. It is present only in Kinetoplastidae protozoa and its sequence lacks homology to known functional domains. Tc38 orthologues present in Trypanosoma brucei and Leishmania were proposed to participate in quite different cellular processes. To further understand the function of this protein in Trypanosoma cruzi, we examined its in vitro binding to biologically relevant [dT-dG] enriched sequences, its expression and subcellular localization during the cell cycle and through the parasite life stages.

Results: By using specific antibodies, we found that Tc38 protein from epimastigote extracts participates in complexes with the poly [dT-dG] probe as well as with the universal minicircle sequence (UMS), a related repeated sequence found in maxicircle DNA, and the telomeric repeat. However, we found that Tc38 predominantly localizes into the mitochondrion. Though Tc38 is constitutively expressed through non-replicating and replicating life stages of T. cruzi, its subcellular localization in the unique parasite mitochondrion changes according to the cell cycle stage. In epimastigotes, Tc38 is found only in association with kDNA in G1 phase. From the S to G2 phase the protein localizes in two defined and connected spots flanking the kDNA. These spots disappear in late G2 turning into a diffuse dotted signal which extends beyond the kinetoplast. This later pattern is more evident in mitosis and cytokinesis. Finally, late in cytokinesis Tc38 reacquires its association with the kinetoplast. In non-replicating parasite stages such as trypomastigotes, the protein is found only surrounding the entire kinetoplast structure.

Conclusions: The dynamics of Tc38 subcellular localization observed during the cell cycle and life stages support a major role for Tc38 related to kDNA replication and maintenance.

Show MeSH
Related in: MedlinePlus