A mosaic analysis in Drosophila fat body cells of the control of antimicrobial peptide genes by the Rel proteins Dorsal and DIF.

Expression of the gene encoding the antifungal peptide Drosomycin in Drosophila adults is controlled by the Toll signaling pathway. The Rel proteins Dorsal and DIF (Dorsal-related immunity factor) are possible candidates for the transactivating protein in the Toll pathway that directly regulates the drosomycin gene. We have examined the requirement of Dorsal and DIF for drosomycin expression in larval fat body cells, the predominant immune-responsive tissue, using the yeast site-specific flp/FRT recombination system to generate cell clones homozygous for a deficiency uncovering both the dorsal and the dif genes. Here we show that in the absence of both genes, the immune-inducibility of drosomycin is lost but can be rescued by overexpression of either dorsal or dif under the control of a heat-shock promoter. This result suggests a functional redundancy between both Rel proteins in the control of drosomycin gene expression in the larvae of Drosophila. Interestingly, the gene encoding the antibacterial peptide Diptericin remains fully inducible in the absence of the dorsal and dif genes. Finally, we have used fat body cell clones homozygous for various mutations to show that a linear activation cascade Spaetzle--> Toll-->Cactus-->Dorsal/DIF leads to the induction of the drosomycin gene in larval fat body cells.     

Yantar, a conserved arginine-rich protein is involved in Drosophila hemocyte development

To identify novel factors involved in Drosophila hematopoiesis, we screened a collection of lethal recessive mutations that also affected normal hemocyte composition in larvae. We present the characterization of the gene yantar (ytr) for which we isolated null and hypomorphic mutations that were associated with severe defects in hemocyte differentiation and proliferation; ytr is predominantly expressed in the hematopoietic tissue during larval development and encodes an evolutionary conserved protein which is predominantly localized in the nucleus. The hematopoietic phenotype in ytr mutants is consistent with a defect or block in differentiation of precursor hemocytes: mutant larvae have enlarged lymph glands (LGs) and have an excess of circulating hemocytes. In addition, many cells exhibit both lamellocyte and crystal cell markers. Ytr function has been preserved in evolution as hematopoietic specific expression of the Drosophila or mouse Ytr proteins rescue the differentiation defects in mutant hemocytes.     

Orbit, a novel microtubule-associated protein essential for mitosis in Drosophila melanogaster

We describe a Drosophila gene, orbit, that encodes a conserved 165-kD microtubule-associated protein (MAP) with GTP binding motifs. Hypomorphic mutations in orbit lead to a maternal effect resulting in branched and bent mitotic spindles in the syncytial embryo. In the larval central nervous system, such mutants have an elevated mitotic index with some mitotic cells showing an increase in ploidy. Amorphic alleles show late lethality and greater frequencies of hyperploid mitotic cells. The presence of cells in the hypomorphic mutant in which the chromosomes can be arranged, either in a circular metaphase or an anaphase-like configuration on monopolar spindles, suggests that polyploidy arises through spindle and chromosome segregation defects rather than defects in cytokinesis. A role for the Orbit protein in regulating microtubule behavior in mitosis is suggested by its association with microtubules throughout the spindle at all mitotic stages, by its copurification with microtubules from embryonic extracts, and by the finding that the Orbit protein directly binds to MAP-free microtubules in a GTP-dependent manner.     

The Drosophila homolog of NTF-2, the nuclear transport factor-2, is essential for immune response

Nuclear transport factor-2 (NTF-2) functions in yeast and mammalian cell culture in targeting proteins into the nucleus. The Drosophila homolog, DNTF-2, is an essential component of the nuclear import machinery, since ntf mutants are lethal. Interestingly, hypomorphic alleles show specific phenotypes. Some are viable, but the number of omatidia in the eye is severely reduced. The immune response in the Drosophila larval fat body is also affected; the three NF-κB/Rel proteins Dorsal, Dif and Relish do not target to the nucleus after infection, and, consequently, the expression of the anti-microbial peptide genes drosomycin, attacin and drosocin is severely impaired. Hence, in spite of its general requirement in many developmental processes, DNTF-2 has a higher specific requirement in the development of the eye and in the immune response. We also found that DNTF-2 interacts directly with Mbo/DNup88, which does not contain phenylalanine-glycine-rich repeats, but has been shown to function in the import of Rel proteins.     

Functional characterization of the Drosophila MRP (mitochondrial RNA processing) RNA gene

MRP RNA is a noncoding RNA component of RNase mitochondrial RNA processing (MRP), a multi-protein eukaryotic endoribonuclease reported to function in multiple cellular processes, including ribosomal RNA processing, mitochondrial DNA replication, and cell cycle regulation. A recent study predicted a potential Drosophila ortholog of MRP RNA (CR33682) by computer-based genome analysis. We have confirmed the expression of this gene and characterized the phenotype associated with this locus. Flies with mutations that specifically affect MRP RNA show defects in growth and development that begin in the early larval period and end in larval death during the second instar stage. We present several lines of evidence demonstrating a role for Drosophila MRP RNA in rRNA processing. The nuclear fraction of Drosophila MRP RNA localizes to the nucleolus. Further, a mutant strain shows defects in rRNA processing that include a defect in 5.8S rRNA processing, typical of MRP RNA mutants in other species, as well as defects in early stages of rRNA processing.     

Regulation of larval hematopoiesis in Drosophila melanogaster: a role for the multi sex combs gene

Drosophila larval hematopoietic organs produce circulating hemocytes that ensure the cellular host defense by recognizing and neutralizing non-self or noxious objects through phagocytosis or encapsulation and melanization. Hematopoietic lineage specification as well as blood cell proliferation and differentiation are tightly controlled. Mutations in genes that regulate lymph gland cell proliferation and hemocyte numbers in the body cavity cause hematopoietic organ overgrowth and hemocyte overproliferation. Occasionally, mutant hemocytes invade self-tissues, behaving like neoplastic malignant cells. Two alleles of the Polycomb group (PcG) gene multi sex combs (mxc) were previously isolated as such lethal malignant blood neoplasm mutations. PcG genes regulate Hox gene expression in vertebrates and invertebrates and participate in mammalian hematopoiesis control. Hence we investigated the need for mxc in Drosophila hematopoietic organs and circulating hemocytes. We show that mxc-induced hematopoietic hyperplasia is cell autonomous and that mxc mainly controls plasmatocyte lineage proliferation and differentiation in lymph glands and circulating hemocytes. Loss of the Toll pathway, which plays a similar role in hematopoiesis, counteracted mxc hemocyte proliferation but not mxc hemocyte differentiation. Several PcG genes tested in trans had no effects on mxc hematopoietic phenotypes, whereas the trithorax group gene brahma is important for normal and mutant hematopoiesis control. We propose that mxc provides one of the regulatory inputs in larval hematopoiesis that control normal rates of plasmatocyte and crystal lineage proliferation as well as normal rates and timing of hemocyte differentiation.     

Signaling mechanisms in the antimicrobial host defense of Drosophila

Drosophila has appeared in recent years as a powerful model to study innate immunity. Several papers published in the past year shed light on the role of the three Rel proteins Dorsal, Dif and Relish in the regulation of antimicrobial peptide expression. In addition, the discovery that a blood serine protease inhibitor is involved in the control of the antifungal response indicates that Toll is activated upon triggering of a proteolytic cascade and does not function as a Drosophila pattern recognition receptor.     

The peripheral nervous system supports blood cell homing and survival in the Drosophila larva

Interactions of hematopoietic cells with their microenvironment control blood cell colonization, homing and hematopoiesis. Here, we introduce larval hematopoiesis as the first Drosophila model for hematopoietic colonization and the role of the peripheral nervous system (PNS) as a microenvironment in hematopoiesis. The Drosophila larval hematopoietic system is founded by differentiated hemocytes of the embryo, which colonize segmentally repeated epidermal-muscular pockets and proliferate in these locations. Importantly, we show that these resident hemocytes tightly colocalize with peripheral neurons and we demonstrate that larval hemocytes depend on the PNS as an attractive and trophic microenvironment. atonal (ato) mutant or genetically ablated larvae, which are deficient for subsets of peripheral neurons, show a progressive apoptotic decline in hemocytes and an incomplete resident hemocyte pattern, whereas supernumerary peripheral neurons induced by ectopic expression of the proneural gene scute (sc) misdirect hemocytes to these ectopic locations. This PNS-hematopoietic connection in Drosophila parallels the emerging role of the PNS in hematopoiesis and immune functions in vertebrates, and provides the basis for the systematic genetic dissection of the PNS-hematopoietic axis in the future.     

家蚕免疫稳态调控分子的鉴定和表达模式分析

昆虫免疫稳态的维持有赖于准确地激活和有效地抑制Toll或IMD信号通路中的关键转录因子-- Dorsal/Dif或Relish。在果蝇等昆虫中, 已报道了多种降低转录因子稳定性和活性的免疫稳态调控分子, 突变或敲除这类分子导致免疫系统的过度激活。对家蚕Bombyx mori免疫信号通路的研究中, 至今为止尚无对这类分子的探索。本研究通过比较基因组学, 在家蚕基因组中鉴定了多个可能参与免疫稳态调控的分子, 包括Wnt家族成员、 Ubc9、 FAF和POSH等; 并通过检测家蚕被微生物感染后这些分子在多种免疫器官中的诱导表达模式, 发现这些分子的表达水平在微生物感染后普遍呈下降趋势, 虽然在某些组织中表达量有明显的升高（>1.5倍）, 但此高表达水平均不能维持且迅速下降; 而且免疫稳态调控分子和受其调控的信号通路的对应关系在不同组织中表现出差异。本研究是首次对家蚕免疫稳态调控分子的报道, 为深入研究家蚕免疫负调控的分子机制提供了参考。