Human lipocalin-1 (Lcn-1, also called tear lipocalin), a member of the lipocalin structural superfamily, is produced by a number of glands and tissues and is known to bind an unusually large array of hydrophobic ligands. Apart from its specific function in stabilizing the lipid film of human tear fluid, it is suggested to act as a physiological scavenger of potentially harmful lipophilic compounds, in general. To characterize proteins involved in the reception, detoxification, or degradation of these ligands, a cDNA phage-display library from human pituitary gland was constructed and screened for proteins interacting with Lcn-1. Using this method an Lcn-1 interacting phage was isolated that expressed a novel human protein. Molecular cloning and analysis of the entire cDNA indicated that it encodes a 55-kDa protein, lipocalin-1 interacting membrane receptor (LIMR), with nine putative transmembrane domains. The cell membrane location of this protein was confirmed by immunocytochemistry and Western blot analysis of membrane fractions of human NT2 cells. Independent biochemical investigations using a recombinant N-terminal fragment of LIMR also demonstrated a specific interaction with Lcn-1 in vitro. Based on these data, we suggest LIMR to be a receptor of Lcn-1 ligands. These findings constitute the first report of cloning of a lipocalin interacting, plasma membranelocated receptor, in general. In addition, a sequence comparison supports the biological relevance of this novel membrane protein, because genes with significant nucleotide sequence similarity are present in Takifugu rubripes, Drosophila melanogaster, Caenorhabditis elegans, Mus musculus, Bos taurus, and Sus scrofa. According to data derived from the human genome sequencing project, the LIMR-encoding gene has to be mapped on human chromosome 12, and its intron/exon organization could be established. The entire LIMR-encoding gene consists of about 13.7 kilobases in length and contains 16 introns with a length between 91 and 3438 base pairs.The protein superfamily of lipocalins consists of small, mainly secretory proteins defined on the basis of conserved amino acid sequence motifs and their common structure. Functionally they share several properties including the ability to bind/transport a remarkable array of small hydrophobic molecules, the formation of macromolecular complexes, and the binding to specific cell surface receptors (1-3). Whereas a large number of various lipophilic ligands able to bind to lipocalins are known, only limited data are available concerning the identity of lipocalin receptors. There is clear evidence of a specific receptor for plasma retinol-binding protein (RBP) 1 (4 -7) and more indirect evidence for receptors for ␣1-microglobulin (8), major urinary protein (9), ␤-lactoglobulin (10), olfactory-binding protein (11), ␣1-acid glycoprotein (12), and glycodelin (13), but with the exception of megalin, which seems to be an endocytic receptor for a variety of soluble macromolecules including several lipocalins, no specific lipocal...

Follicular B (FoB) and marginal zone B (MZB) cells are functionally and spatially distinct mature B cell populations in the spleen, originating from a Notch2-dependent fate decision after splenic influx of immature transitional B cells. In the B cell follicle, a Notch2-signal is provided by DLL-1-expressing fibroblasts. However, it is unclear whether FoB cells, which are in close contact with these DLL-1 expressing fibroblasts, can also differentiate to MZB cells if they receive a Notch2-signal. Here, we show induced Notch2IC-expression in FoB cells re-programs mature FoB cells into bona fide MZB cells as is evident from the surface phenotype, localization, immunological function and transcriptome of these cells. Furthermore, the lineage conversion from FoB to MZB cells occurs in immunocompetent wildtype mice. These findings demonstrate plasticity between mature FoB and MZB cells that can be driven by a singular signaling event, the activation of Notch2.

The upstream binding factor UBF, an activator of RNA polymerase I transcription, is posttranslationally modified by phosphorylation and acetylation. We found that in NIH3T3 cells, UBF is acetylated in S-phase but not in G1-phase. To assess the role of acetylation in regulation of UBF activity, we have established an NIH3T3 cell line that inducibly overexpresses HDAC1. Both in vivo and in vitro, HDAC1 efficiently hypoacetylates UBF. Immunoprecipitation with antibodies against the Pol I-associated factor PAF53 co-precipitated UBF in mock cells but not in cells overexpressing HDAC1. Pull-down experiments showed that acetylation of UBF augments the interaction with Pol I. Consistent with acetylation of UBF being important for association of PAF53 and recruitment of Pol I, the level of Pol I associated with rDNA and pre-rRNA synthesis were reduced in cells overexpressing HDAC1. The results suggest that acetylation and deacetylation of UBF regulate rRNA synthesis during cell cycle progression.

There is increasing experimental evidence demonstrating that many lipocalins bind to specific cell surface receptors. However, whereas the binding of lipocalins to their lipophilic ligands has now been characterized in much detail, there is a lack of knowledge about the nature of lipocalin receptors, the physiological role of receptor binding, and the molecular mechanism of ligand delivery. We previously identified a novel human membrane protein (lipocalin-1-interacting membrane receptor (LIMR)), which interacts with lipocalin-1 (Wojnar, P., Lechner, M., Merschak, P., and Redl, B. (2001) J. Biol. Chem. 276, 20206 -20212). In the present study, we investigated the physiological role of LIMR and found this protein to be essential for mediating internalization of lipocalin-1 (Lcn-1) in NT2 cells, leading to its degradation. Whereas control NT2 cells rapidly internalized 125 I-Lcn-1 or fluorescein isothiocyanate-labeled Lcn-1, NT2 cells that were made LIMR deficient by cDNA antisense expression greatly accumulated Lcn-1 in the culture medium but did not internalize it. Because sequence and structure analysis indicated that proteins similar to LIMR are present in several organisms and at least two closely related orthologues are found in human and mouse, we suggest LIMR to be the prototype of a new family of endocytic receptors, which are topographically characterized by nine putative transmembrane domains and a characteristic large central cytoplasmic loop.Lipocalins were found to be important extracellular carriers of lipophilic compounds in vertebrates, invertebrates, plants, and bacteria (1-4). There is increasing evidence that this group of proteins is involved in a variety of physiological processes including retinoid, fatty acid, and pheromone signaling; immunomodulation; inflammation; detoxification; modulation of growth and metabolism; tissue development; apoptosis; and even behavior processes (1, 5-7). Whereas the structural basis of lipocalin-ligand binding is now well understood (8), there is a major lack of knowledge regarding the mechanisms by which lipocalins exert their biological effects. It is well accepted that many, if not all, of these proteins are able to bind to specific cell receptors (9), although only two of these receptors have been identified thus far (10, 11). Due to limited data concerning the structure of the lipocalin receptors themselves, the molecular mechanisms beyond this receptor binding are very unclear at the moment. One hypothesis is that the holo-lipocalin releases its ligand upon receptor binding, and this ligand diffuses through the cell membrane to interact with an intracellular fatty acid-binding protein or an intracellular receptor. There is also some evidence that lipocalins undergo internalization by receptor-mediated endocytosis. Another plausible mechanism might be that the lipocalin-receptor interaction creates a direct signal inducing various physiological processes (9).We have recently identified and characterized LIMR, 1 a novel human 57-kDa cell membrane protein ...