Browsing by Subject "Rezeptor"

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Lokalisation von Pheromon-Rezeptoren und -Bindeproteinen in antennalen Sensillen von Insekten
(2007) Gohl, Thomas; Breer, Heinz
The remarkable reactivity of moth to specific pheromones is based on the extreme selectivity and sensitivity of sensory cells in the male antennae. This feature is supposed to be based on cells equipped with specific receptors. Only the sequencing of the genomes of Bombyx mori and Heliothis virescens provided the possibility to identify candidate genes of olfactory receptors in moths. Upon detailed inspection of candidate receptors it turned out that within the generally very heterogeneous group of receptor-genes a subfamily exists containing members of both species showing striking sequence homology. A conservation of the primary structure of receptors for pheromones has been postulated. For a continuing characterization in these studies different approaches were used to verify that these receptors are indeed expressed in cells of pheromone-sensitive sensilla (sensilla trichodea). By means of ?whole mount? in situ hybridization experiments the RNA of the receptor types BmOR1 and BmOR3 could be visualized in directly neighboring cells reflecting the topology of trichoid sensilla. Also some of the Heliothis receptor types (HR13, HR14, HR16) could be assigned to sensilla trichodea. In addition to the specific receptors, the pheromone binding proteins (PBPs) are expected to play an important role in the detection of hydrophobic pheromone molecules. PBPs are produced by glia-like cells surrounding the sensory neurons. In double in situ hybridization experiments it could be shown that HR13-cells are indeed surrounded by cells expressing HvirPBP1 and HvirPBP2. Analysis comparing the topology of different receptor-types showed that cells expressing HR13 can be assigned to sensilla trichodea type A, whereas HR14 and HR16 are expressed in cells of sensilla trichodea type C. This characteristic expression pattern is considered as a further indication that these candidate-receptors are indeed pheromone-receptors. The assignment of individual receptor-types to distinct sensilla-types provides the basis for investigating the functional implications of receptor-types for the registration of main or minor components of complex pheromone-blends. Further it turned out that HR13 shows coexpression with SNMP1 (sensory neuron membrane protein 1) which is considered as a ?marker?-protein for antennal sensory neurons. This is however not the case for receptor types HR14 and HR16. In search of further SNMP-types screening-experiments were carried out which led to the identification of a novel SNMP-type (SNMP2) of Heliothis virescens. Subsequent studies concerning the expression of SNMP2 showed that the topologic distribution of SNMP2-cells is comparable to SNMP1-cells, but they show a different morphology. Further experiments revealed that SNMP2 is in fact expressed in PBP-producing cells. These findings imply that the proposed putative function of SNMPs has to be reconsidered. One major goal of this study was the attempt, to identify receptor-relevant cells by visualization of mRNA via in situ hybridization but to visualize the localization of the receptor-protein via immunohistochemical approaches. Although the generation of antibodies for olfactory receptors is very difficult, it was possible to raise antibodies specific for receptor type HR13. Using these antibodies in immunohistochemical approaches allowed to also visualize HR13-receptor-protein. By means of double-staining experiments using HR13-specific antisense RNA-probes and anti-HR13 antibodies mRNA and protein were visualized in the same specific cells. Using confocal laserscanning microscopy, it was possible to document that receptor-protein was indeed located in the sensory dendrites. Further, the receptor-protein was also visualized in the axonal processes of sensory cells and the receptor-specific staining revealed that within the antennal nerve HR13-axons appear to be organized in fascicles. These HR13-immunolabeled fascicles were visible until they reach the ?sorting zone? of the antennal lobe; in contrast to mouse olfactory bulb, no receptor specific staining was visible in the antennal lobe.
Olfaktorische Rezeptoren mit speziellen topographischen Expressionsmustern
(2007) Feistel, Torben; Breer, Heinz
In the present study, olfactory receptors (ORs) featuring special expression patterns in chemosensory subsystems were analyzed. The data showed that out of a repertoire of about 1000 ORs a restricted group (about 50) was not only expressed in the olfactory epithelium but also in the vomeronasal organ, which is generally defined by the expression of characteristic V1R and V2R receptors. The ?ectopically? expressed OR genes represent different receptor families including genes from gene-clusters located on different chromosomes. However, in all individuals the same set of genes seemed to be expressed. The majority of the expressed ORs was present in only a small number of VNO cells, however some were found to be expressed in more than 100 cells. One distinct OR gene (mOR261-6) was expressed in many more cells of female VNOs than in males. The highest number of OR expressing cells was observed in a short postnatal, pre-pubertal period. Cells with mOR18-2-receptors were also found to express a V1R gene. In these cells the OR18-2 gene did not show a mono-allelic expression as compared to expression in the main olfactory system but rather was transcribed from both alleles (bi-allelic) in the vomeronasal neurons. The functional implication of receptor coexpression and bi-allelic OR-expression are unknown. Receptors of the OR37 gene family are exclusively expressed in the olfactory epithelium, in which they are expressed in a special pattern in a central area of the nasal turbinate. Detailed analysis of the spatial distribution of cells equipped with distinct OR37 subtypes revealed that these cells were positioned in specific sub-compartments within this central region. The high number of OR37 expressing cells resulted in the lower number of cells with receptors which are zonally distributed. This implies that cells in a small circumscribed area preferentially expressed OR37 genes. The mechanisms underlying this unique spatial expression pattern of OR37 genes in the olfactory epithelium are currently unknown. A newly generated mouse line was arranged in which even very transient transcription of a OR37 gene is visualized by permanent label in the expressing cells. The examination showed that cells outside of the OR37 cluster did express a OR 37 family member, yet transcription in these cells was rapidly terminated. These results suggest a mechanism which allows an initial transcription of these genes in different areas of the epithelium, but a sustained expression of OR37 genes is restricted exclusively to the central recess of the turbinate. The special features of the OR37 subtypes have led to the hypothesis that these receptors possibly react to chemical compounds relevant for mammalian species. Thus complex mixtures of volatile compounds from the habitat of mice were examined. Electroolfactograms from different regions of the olfactory epithelium showed that the ?head-space? of the embedding from mice cages elicited a stronger response in the central area of the turbinate IIb than in areas outside of this region. Single substances out of this complex mixture induced similar responses in different epithelial recesses. However, 6-hydro-6-methyl-3-heptanon elicited stronger responses within the central OR37 expressing region. This substance is considered to be a mouse pheromone. Taken together, these data on expression and ligand specificity of distinct olfactory receptors suggest, that the vomeronasal organ and the olfactory epithelium although structurally separated chemosensory systems, overlap in their response, spectrum and in their functions, particularly in detecting compounds with biological relevance.
PEP7 is a ligand for receptor kinase SIRK1 to regulate aquaporins and root growth
(2021) Wang, Jiahui; Schulze, Waltraud
Receptor kinases constitute the largest protein family in regulating various responses to external and internal biotic and abiotic signals. Functional characterization of this large protein family and particularly the identification of their ligands remains a major challenge in plant biology. Previously, we identified SIRK1 and QSK1 as a receptor / co-receptor pair involved in regulation of aquaporins in response to osmotic changes induced by sucrose. Here, we now identify a member of the Elicitor Peptide (PEP) family, namely PEP7, as a ligand to receptor kinase SIRK1. PEP7 was shown to bind to the extracellular domain of SIRK1 with a binding constant of 19 µM. PEP7 was secreted to the apoplasm specifically in response to sucrose. Formation of a signaling complex involving SIRK1, QSK1 as well as aquaporins as substrates was induced by sucrose or external PEP7 treatment. PEP7 induced aquaporin phosphorylation and water influx activity. The knock-out mutant of receptor SIRK1 was not responsive to external PEP7 treatment. Binding to receptor SIRK1 and induction of physiological responses was specific to PEP7, neither other members of the PEP-family (PEP6, PEP4), nor other small signaling peptides (CLEs, IDA, RALFs) induced SIRK1 kinase activity, aquaporin phosphorylation, or protoplast water influx activity.