Dataset Information

Asrij maintains the stem cell niche and controls differentiation during Drosophila lymph gland hematopoiesis.

ABSTRACT: Several signaling pathways control blood cell (hemocyte) development in the Drosophila lymph gland. Mechanisms that modulate and integrate these signals are poorly understood. Here we report that mutation in a conserved endocytic protein Asrij affects signal transmission and causes aberrant lymph gland hematopoiesis. Mammalian Asrij (Ociad1) is expressed in stem cells of the blood vascular system and is implicated in several cancers. We found that Drosophila Asrij is a pan-hemocyte marker and localizes to a subset of endocytic vesicles. Loss of asrij causes hyperproliferation of lymph gland lobes coupled with increased hemocyte differentiation, thereby depleting the pool of quiescent hemocyte precursors. This co-relates with fewer Col+ cells in the hematopoietic stem cell niche of asrij mutants. Asrij null mutants also show excess specification of crystal cells that express the RUNX factor Lozenge (Lz), a target of Notch signaling. Asrij mutant lymph glands show increased N in sorting endosomes suggesting aberrant trafficking. In vitro assays also show impaired traffic of fluorescent probes in asrij null hemocytes. Taken together our data suggest a role for Asrij in causing increased Notch signaling thereby affecting hemocyte differentiation. Thus, conserved endocytic functions may control blood cell progenitor quiescence and differentiation.

SUBMITTER: Kulkarni V

PROVIDER: S-EPMC3215734 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Asrij maintains the stem cell niche and controls differentiation during Drosophila lymph gland hematopoiesis.

Kulkarni Vani V Khadilkar Rohan J RJ Magadi Srivathsa S SS Inamdar Maneesha S MS

PloS one 20111114 11

Several signaling pathways control blood cell (hemocyte) development in the Drosophila lymph gland. Mechanisms that modulate and integrate these signals are poorly understood. Here we report that mutation in a conserved endocytic protein Asrij affects signal transmission and causes aberrant lymph gland hematopoiesis. Mammalian Asrij (Ociad1) is expressed in stem cells of the blood vascular system and is implicated in several cancers. We found that Drosophila Asrij is a pan-hemocyte marker and lo ...[more]

PMID: 22110713

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Project description:Hematopoiesis occurs in two phases in Drosophila, with the first completed during embryogenesis and the second accomplished during larval development. The lymph gland serves as the venue for the final hematopoietic program, with this larval tissue well-studied as to its cellular organization and genetic regulation. While the medullary zone contains stem-like hematopoietic progenitors, the posterior signaling center (PSC) functions as a niche microenvironment essential for controlling the decision between progenitor maintenance versus cellular differentiation. In this report, we utilize a PSC-specific GAL4 driver and UAS-gene RNAi strains, to selectively knockdown individual gene functions in PSC cells. We assessed the effect of abrogating the function of 820 genes as to their requirement for niche cell production and differentiation. 100 genes were shown to be essential for normal niche development, with various loci placed into sub-groups based on the functions of their encoded protein products and known genetic interactions. For members of three of these groups, we characterized loss- and gain-of-function phenotypes. Gene function knockdown of members of the BAP chromatin-remodeling complex resulted in niche cells that do not express the hedgehog (hh) gene and fail to differentiate filopodia believed important for Hh signaling from the niche to progenitors. Abrogating gene function of various members of the insulin-like growth factor and TOR signaling pathways resulted in anomalous PSC cell production, leading to a defective niche organization. Further analysis of the Pten, TSC1, and TSC2 tumor suppressor genes demonstrated their loss-of-function condition resulted in severely altered blood cell homeostasis, including the abundant production of lamellocytes, specialized hemocytes involved in innate immune responses. Together, this cell-specific RNAi knockdown survey and mutant phenotype analyses identified multiple genes and their regulatory networks required for the normal organization and function of the hematopoietic progenitor niche within the lymph gland.

Project description:Hematological disorders result in perturbed homeostasis of the blood system. However, a comprehensive understanding of how physiological and genetic mechanisms regulate blood cell precursor maintenance and differentiation is lacking. Owing to simplicity and ease of genetic analysis, the Drosophila melanogaster lymph gland (LG) is an excellent model to study hematopoiesis. The LG is a multi-lobed structure compartmentalized into precursor and differentiation zones whose geography and identity is regulated by multiple signalling pathways. While additional molecular and functional subtypes are expected there is a paucity of information on gene expression and regulation of hemocyte homeostasis. Hence, we quantitatively analyzed the LG proteome under conditions that maintain precursors or promote their differentiation in vivo, by perturbing expression of Asrij, a conserved endosomal regulator of hematopoiesis. Although technically demanding, we pooled samples obtained from 1500 larval dissections per genotype and using iTRAQ quantitative proteomics, determined the relative expression levels of polypeptides in Asrij knockout (KO) and overexpressing (OV) LGs in comparison to wild type (control). Mass spectrometry data analysis showed that at least 6.5% of the Drosophila proteome is expressed in wild type LGs.Of 2,133 proteins identified, 780 and 208 proteins were common to the previously reported cardiac tube and hemolymph proteomes, respectively, resulting in the identification of 1238 proteins exclusive to the LG. Perturbation of Asrij levels led to differential expression of 619 proteins, of which 23% have human homologs implicated in various diseases. Proteins regulating metabolism, immune system, signal transduction and vesicle-mediated transport were significantly enriched. Immunostaining of representative candidates from the enriched categories and previous reports confirmed 75% of our results and validated the LG proteome. Our study provides, for the first time, an in vivo proteomics resource for identifying novel regulators of hematopoiesis that will also be applicable to understanding vertebrate blood cell development.

Dataset Information

Asrij maintains the stem cell niche and controls differentiation during Drosophila lymph gland hematopoiesis.

Publications

Asrij maintains the stem cell niche and controls differentiation during Drosophila lymph gland hematopoiesis.

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