Limb development membrane protein-1 (LMBR1)/lipocalin-interacting membrane receptor (LIMR)-type proteins are putative ninetransmembrane receptors that are evolutionarily conserved across metazoans. However, their biological function is unknown. Here, we show that the fly family member Lilipod (Lili) is required for germline stem cell (GSC) self-renewal in the Drosophila ovary where it enhances bone morphogenetic protein (BMP) signaling. lili mutant GSCs are lost through differentiation, and display reduced levels of the Dpp transducer pMad and precocious activation of the master differentiation factor bam. Conversely, overexpressed Lili induces supernumerary pMad-positive bamP-GFP-negative GSCs. Interestingly, differentiation of lili mutant GSCs is bam-dependent; however, its effect on pMad is not. Thus, although it promotes stem cell self-renewal by repressing a bam-dependent process, Lilipod enhances transduction of the Dpp signal independently of its suppression of differentiation. In addition, because Lili is still required by a ligand-independent BMP receptor, its function likely occurs between receptor activation and pMad phosphorylation within the signaling cascade. This first, to our knowledge, in vivo characterization of a LMBR1/LIMR-type protein in a genetic model reveals an important role in modulating BMP signaling during the asymmetric division of an adult stem cell population and in other BMP signaling contexts.T he human lipocalin-interacting membrane receptor LIMR (also known as LMBR1L) and the closely related limb development membrane protein-1 (LMBR1) share a predicted multitransmembrane (TM) structure that is strikingly different from other well-characterized integral membrane receptors. LIMR was originally isolated by phage display through its interaction with lipocalin-1 (Lcn-1), a secreted lipid-binding carrier protein (1). Cell culture studies showed that LIMR could mediate the internalization of Lcn-1, suggesting a role in cell signaling (2). However, because the physiological roles of both Lcn-1 and LIMR are unknown, the significance of this observation is unclear. Less is known about the LMBR1 protein. Initially, the human locus was genetically linked to multiple congenital limb malformations. However, further studies of the human and mouse loci showed that the original association with limb defects was incidental because of the disruption of a long-range SHH enhancer located within an intron of LMBR1 (3). To date, no loss-of-function or gain-of-function analyses of LIMR or LMBR1 have been reported in any model system, and studies in vertebrates may be complicated by functional redundancy between the two family members.Drosophila contains a single uncharacterized LIMR-like protein, CG5807, which is the shared ortholog of both LIMR and LMBR1. We have investigated the fly gene in vivo and show here that it functions in the germ-line stem cells (GSC) of the Drosophila ovary. In the fruit fly, oocytes are continually produced by GSCs that are housed within a structure called germarium (Fig....
BackgroundBMP signaling is involved in myriad metazoan developmental processes, and study of this pathway in Drosophila has contributed greatly to our understanding of its molecular and genetic mechanisms. These studies have benefited not only from Drosophila’s advanced genetic tools, but from complimentary in vitro culture systems. However, the commonly-used S2 cell line is not intrinsically sensitive to the major BMP ligand Dpp and must therefore be augmented with exogenous pathway components for most experiments.ResultsHerein we identify and characterize the responses of Drosophila ML-DmD17-c3 cells, which are sensitive to Dpp stimulation and exhibit characteristic regulation of BMP target genes including Dad and brk. Dpp signaling in ML-DmD17-c3 cells is primarily mediated by the receptors Put and Tkv, with additional contributions from Wit and Sax. Furthermore, we report complex regulatory feedback on core pathway genes in this system.ConclusionsNative ML-DmD17-c3 cells exhibit robust transcriptional responses to BMP pathway induction. We propose that ML-DmD17-c3 cells are well-suited for future BMP pathway analyses.Electronic supplementary materialThe online version of this article (10.1186/s12861-019-0181-0) contains supplementary material, which is available to authorized users.
Background: Replication requires coordinated interactions among proteins/DNA. Results: Mutational analysis of T4 helicase loader, gp59, identifies domains that interact with DNA and replication factors. Conclusion: The high mobility group motif of gp59 interacts with the DNA fork; gp59 C-terminal surfaces interact with helicase and single-stranded binding protein.Significance: Processive replication may involve switching of gp59 between interactions with helicase and the single-stranded binding protein.
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