Bam and Bgcn antagonize
            <i>Nanos</i>
            -dependent germ-line stem cell maintenance

Li, Yun; Minor, Nicole T.; Park, Joseph K.; McKearin, Dennis M.; Maines, Jean Z.

doi:10.1073/pnas.0901452106

Cited by 148 publications

(219 citation statements)

References 48 publications

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“…GSCs express the translational repressor Nanos, a factor essential for GSC maintenance. On displacement away from the cap cell niche, GSC daughters express Bam, which represses the translation of nanos in differentiating cysts (30). Costaining control and Lsd1 mutant germaria for Nanos and Bam revealed that less than 1.5% of Lsd1 mutant germaria expressed detectable levels of Bam (Fig.…”

Section: Resultsmentioning

confidence: 99%

Loss of lysine-specific demethylase 1 nonautonomously causes stem cell tumors in the Drosophila ovary

Eliazer

Shalaby

2011

Proc. Natl. Acad. Sci. U.S.A.

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Specialized microenvironments called niches keep stem cells in an undifferentiated and self-renewing state. Dedicated stromal cells form niches by producing a variety of factors that act directly on stem cells. The size and signaling output of niches must be finely tuned to ensure proper tissue homeostasis. Although advances have been made in identifying factors that promote niche cell fate, the mechanisms that restrict niche cell formation during development and limit niche signaling output in adults remain poorly understood. Here, we show that the histone lysine-specific demethylase 1 (Lsd1) regulates the size of the germline stem cell (GSC) niche in Drosophila ovaries. GSC maintenance depends on bone morphogenetic protein (BMP) signals produced by a small cluster of cap cells located at the anterior tip of the germarium. Lsd1 null mutant ovaries carry small germline tumors containing an expanded number of GSC-like cells with round fusomes that display ectopic BMP signal responsiveness away from the normal niche. Clonal analysis and cell type-specific rescue experiments demonstrate that Lsd1 functions within the escort cells (ECs) that reside immediately adjacent to cap cells and prevents them from ectopically producing niche-specific signals. Temporally restricted gene knockdown experiments suggest that Lsd1 functions both during development, to specify EC fate, and in adulthood, to prevent ECs from forming ectopic niches independent of changes in cell fate. Further analysis shows that Lsd1 functions to repress decapentaplegic (dpp) expression in adult germaria. The role of Lsd1 in regulating niche-specific signals may have important implications for understanding how disruption of its mammalian homolog contributes to cancer and metastasis.

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Section: Resultsmentioning

confidence: 99%

Loss of lysine-specific demethylase 1 nonautonomously causes stem cell tumors in the Drosophila ovary

Eliazer

Shalaby

2011

Proc. Natl. Acad. Sci. U.S.A.

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“…1C). To express sgRNA in vivo, we selected three different regulatory sequences: U6a (CR31379) and U6b (CR32867) (26,27), both of which direct transcription of Drosophila U6 snRNAs, and nosmini, which contains the nos promoter and a minimal 5′ UTR (28,29). We used the 404-bp upstream and 93-bp downstream regulatory sequences of U6a to construct U6a-sgRNA-short and U6a-sgRNA-long.…”

Section: Methodsmentioning

confidence: 99%

Optimized gene editing technology for Drosophila melanogaster using germ line-specific Cas9

Greenleaf

Sun

Housden

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

382

402

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The ability to engineer genomes in a specific, systematic, and costeffective way is critical for functional genomic studies. Recent advances using the CRISPR-associated single-guide RNA system (Cas9/sgRNA) illustrate the potential of this simple system for genome engineering in a number of organisms. Here we report an effective and inexpensive method for genome DNA editing in Drosophila melanogaster whereby plasmid DNAs encoding short sgRNAs under the control of the U6b promoter are injected into transgenic flies in which Cas9 is specifically expressed in the germ line via the nanos promoter. We evaluate the off-targets associated with the method and establish a Web-based resource, along with a searchable, genome-wide database of predicted sgRNAs appropriate for genome engineering in flies. Finally, we discuss the advantages of our method in comparison with other recently published approaches.nanos-Cas9 | HRMA M uch of our knowledge of the mechanisms underlying biological processes relies on genetic approaches, whereby gene activity is perturbed and the phenotypic consequences of perturbation are analyzed in detail. In recent years, several major advances have been made in the design of methods for specifically and efficiently perturbing genomes. Arguably, the most exciting advances rely on the ability to induce double-strand breaks (DSBs) by targeting a nuclease to a specific genomic sequence. Repair of DSBs by the error-prone nonhomologous endjoining (NHEJ) mechanism allows for the recovery of small deletions; moreover, repair of DSBs by homologous recombination (HR) in the presence of a donor template opens the door to a wide range of specifically engineered changes at the targeted site (1).Two nuclease-based systems, the zinc-finger nuclease (ZFN) and transcription activator-like effector nuclease (TALEN) systems, work effectively in a number of organisms (2-7). But because these approaches require the production of a construct encoding a unique DNA-binding protein fused to the nuclease domain, they can be both cumbersome and costly. In contrast, the recent approach based on the bacterial CRISPR-associated single-guide RNA (Cas9/sgRNA) system does not require production of specific fusion proteins for each targeted sequence (8-10).Cas9 was first identified in type II Streptococcus pyogenes as an RNA-guided defense system against invading viruses and plasmids (11-13). This adaptive immune-like system contains three components: CRISPR RNA (crRNA), trans-activating CRISPR RNA (tracrRNA), and Cas9. The tracrRNA triggers Cas9 nuclease activity and the crRNA guides Cas9 to cleave the specific foreign dsDNA sequence via base-pairing between the crRNA and the target DNA. Importantly, a single-guide RNA (sgRNA, also known as chiRNA), comprising the minimal crRNA and tracrRNA, can function similarly to the crRNA and tracrRNA, thereby providing a simplified method for genome editing (8)(9)(10)(14)(15)(16)(17)(18)(19)(20).Given the great promise of the Cas9/sgRNA method for genome engineering, we set out to test the sys...

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“…Nanos promotes GSC fate, whereas Bam promotes differentiation. These antagonistic activities were shown to depend on the repression of nanos mRNA translation by the Bam/Bgcn complex, possibly via nanos 3′UTR (9). In this model, Bam would thus promote differentiation by inhibiting stem cell-promoting factors, which is different from inhibiting self-renewal by stabilizing differentiation factors, such as CycA.…”

mentioning

confidence: 99%

“…Here, it is important to have in mind in which cell types these proteins are endogenously expressed to distinguish gain-vs. loss-of-function experiments. Bam and Nanos proteins are expressed in nonoverlapping reciprocal domains, with Nanos expressed in GSCs and 16-cell cysts, whereas Bam protein is found exclusively during the four mitotic divisions (9). Bgcn and Otu are expressed throughout the germarium.…”

mentioning

confidence: 99%