Project description:The molecular mechanisms by which stem cell proliferation is precisely controlled during the course of regeneration are poorly understood. Namely, how a damaged tissue senses when to terminate the regeneration process, inactivates stem cell mitotic activity, and organizes ECM integrity remain fundamental unanswered questions. The Drosophila midgut intestinal stem cell (ISC) offers an excellent model system to study the molecular basis for stem cell inactivation. Here we show that a novel gene, CG6967 or dMOV10, is induced at the termination stage of midgut regeneration, and shows an inhibitory effect on ISC proliferation. dMOV10 encodes a putative component of the microRNA (miRNA) gene silencing complex (miRISC). Our data, along with previous studies on the mammalian MOV10, suggest that dMOV10 is not a core member of miRISC, but modulates miRISC activity as an additional component. Further analyses identified direct target mRNAs of dMOV10-containing miRISC, including Daughter against Dpp (Dad), a known inhibitor of BMP/TGF-β signaling. We show that RNAi knockdown of Dad significantly impaired ISC division during regeneration. We also identified miRNAs that are induced at the termination stage and their potential target transcripts. We propose that miRNA-mediated gene regulation contributes to the precise control of Drosophila midgut regeneration.
Project description:The molecular mechanisms by which stem cell proliferation is precisely controlled during the course of regeneration are poorly understood. Namely, how a damaged tissue senses when to terminate the regeneration process, inactivates stem cell mitotic activity, and organizes ECM integrity remain fundamental unanswered questions. The Drosophila midgut intestinal stem cell (ISC) offers an excellent model system to study the molecular basis for stem cell inactivation. Here we show that a novel gene, CG6967 or dMOV10, is induced at the termination stage of midgut regeneration, and shows an inhibitory effect on ISC proliferation. dMOV10 encodes a putative component of the microRNA (miRNA) gene silencing complex (miRISC). Our data, along with previous studies on the mammalian MOV10, suggest that dMOV10 is not a core member of miRISC, but modulates miRISC activity as an additional component. Further analyses identified direct target mRNAs of dMOV10-containing miRISC, including Daughter against Dpp (Dad), a known inhibitor of BMP/TGF-β signaling. We show that RNAi knockdown of Dad significantly impaired ISC division during regeneration. We also identified miRNAs that are induced at the termination stage and their potential target transcripts. We propose that miRNA-mediated gene regulation contributes to the precise control of Drosophila midgut regeneration.
Project description:The molecular mechanisms by which stem cell proliferation is precisely controlled during the course of regeneration are poorly understood. Namely, how a damaged tissue senses when to terminate the regeneration process, inactivates stem cell mitotic activity, and organizes ECM integrity remain fundamental unanswered questions. The Drosophila midgut intestinal stem cell (ISC) offers an excellent model system to study the molecular basis for stem cell inactivation. Here we show that a novel gene, CG6967 or dMOV10, is induced at the termination stage of midgut regeneration, and shows an inhibitory effect on ISC proliferation. dMOV10 encodes a putative component of the microRNA (miRNA) gene silencing complex (miRISC). Our data, along with previous studies on the mammalian MOV10, suggest that dMOV10 is not a core member of miRISC, but modulates miRISC activity as an additional component. Further analyses identified direct target mRNAs of dMOV10-containing miRISC, including Daughter against Dpp (Dad), a known inhibitor of BMP/TGF-β signaling. We show that RNAi knockdown of Dad significantly impaired ISC division during regeneration. We also identified miRNAs that are induced at the termination stage and their potential target transcripts. We propose that miRNA-mediated gene regulation contributes to the precise control of Drosophila midgut regeneration.
Project description:Identification of RNA targets of RNA-binding proteins (RBPs) is essential for complete understanding of their biological functions. However, it is still a challenge to identify the biologically relevant targets of RBPs through in vitro strategies of RIP-seq, HITS-CLIP, or GoldCLIP due to the potentially high background and complicated manipulation. In malaria parasites, RIP-seq and gene disruption are the few tools available currently for identification of RBP targets. Here, we have adopted the TRIBE (Targets of RNA binding proteins identified by editing) system to in vivo identify the RNA targets of PfDis3, a key exoribonuclease subunit of RNA exosome in Plasmodium falciparum. We generated a transgenic parasite line of Pfdis3-ADARcd, which catalyzes an adenosine (A)-to-inosine (I) conversion at the potential interacting sites of PfDis3-targeting RNAs. Most of PfDis3 target genes contain one edit site. The majority of the edit sites detected by PfDis3-TRIBE locate in exons and spread across the entire coding regions. The nucleotides adjacent to the edit sites contain ~ 75% of A+T. PfDis3-TRIBE target genes are biases toward higher RIP enrichment, suggesting that PfDis3-TRIBE preferentially detects stronger PfDis3 RIP targets. Collectively, PfDis3-TRIBE is a favorable tool to identify in vivo target genes of RBP with high efficiency and reproducibility. Additionally, the PfDis3-targeting genes are involved in stage-related biological processes during the blood-stage development. PfDis3 appears to shape the dynamic transcriptional transcriptome of malaria parasites through post-transcriptional degradation of a variety of unwanted transcripts from both strands in the asexual blood stage.
Project description:A midgut reference sample is compared to midgut samples taken at various stages before (18h, 4h), at (0h), and after (2h, 3h, 4h, 5h, 6h, 8h, 10h, 12h) puparium formation. Keywords = Drosophila, ecdysone, network, genomic, microarray, organogenesis, EcR, midgut, central nervous system, salivary gland, epidermis, imaginal disc, development Keywords: other