Nuclear Compartmentalization Contributes to Stage-Specific Gene Expression Control in Trypanosoma cruzi.

In the protozoan parasite Trypanosoma cruzi, as in other trypanosomatids, transcription of protein coding genes occurs in a constitutive fashion, producing large polycistronic transcription units. These units are composed of non-functionally related genes which are pervasively processed to yield each mRNA. Therefore, post-transcriptional processes are crucial to regulate gene expression. Considering that nuclear compartmentalization could contribute to gene expression regulation, we comparatively studied the nuclear, cytoplasmic and whole cell transcriptomes of the non-infective epimastigote stage of T. cruzi, using RNA-Seq. We found that the cytoplasmic transcriptome tightly correlates with the whole cell transcriptome and both equally correlate with the proteome. Nonetheless, 1,200 transcripts showed differential abundance between the nuclear and cytoplasmic fractions. For the genes with transcript content augmented in the nucleus, significant structural and compositional differences were found. The analysis of the reported epimastigote translatome and proteome, revealed scarce ribosome footprints and encoded proteins for them. Ontology analyses unveiled that many of these genes are distinctive of other parasite life-cycle stages. Finally, the relocalization of transcript abundance in the metacyclic trypomastigote infective stage was confirmed for specific genes. While gene expression is strongly dependent on transcript steady-state level, we here highlight the importance of the distribution of transcripts abundance between compartments in T. cruzi. Particularly, we show that nuclear compartmentation is playing an active role in the developmental stage determination preventing off-stage expression.

Invasive prostate cancer cells are tumor initiating cells that have a stem cell-like genomic signature.

Development of metastasis is a leading cause of cancer-induced death. Acquisition of an invasive tumor cell phenotype suggests loss of cell adhesion and basement membrane breakdown during a process termed epithelial-to-mesenchymal transition (EMT). Recently, cancer stem cells (CSC) were discovered to mediate solid tumor initiation and progression. Prostate CSCs are a subpopulation of CD44(+) cells within the tumor that give rise to differentiated tumor cells and also self-renew. Using both primary and established prostate cancer cell lines, we tested the assumption that CSCs are more invasive. The ability of unsorted cells and CD44-positive and -negative subpopulations to undergo Matrigel invasion and EMT was evaluated, and the gene expression profiles of these cells were analyzed by microarray and a subset confirmed using QRT-PCR. Our data reveal that a subpopulation of CD44(+) CSC-like cells invade Matrigel through an EMT, while in contrast, CD44(-) cells are non-invasive. Furthermore, the genomic profile of the invasive cells closely resembles that of CD44(+)CD24(-) prostate CSCs and shows evidence for increased Hedgehog signaling. Finally, invasive cells from DU145 and primary prostate cancer cells are more tumorigenic in NOD/SCID mice compared with non-invasive cells. Our data strongly suggest that basement membrane invasion, an early and necessary step in metastasis development, is mediated by these potential cancer stem cells.

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

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.