Molekulare Verankerung von Resilin in der elastischen Kutikula der Insekten
Zusammenfassung der Projektergebnisse
Resilin is a highly resilient extracellular protein matrix in insects. Within specific regions of the body cuticle it contributes to their stability and function during locomotion including flight and walking as well as for yet undefined movements. The diversity of resilin incidences in an insect detected thanks to its autofluorescence property suggests that each distinct function necessitates qualitative differences between the regions. In order to address this hypothesis, we used the model insect Drosophila melanogaster (fruit fly) in a genetic approach. To begin with, we knocked-out the gene coding for the reported protein unit of resilin named pro-resilin by Crispr/Cas9 generating two mutant alleles. We showed that elimination of Pro-Resilin function is not lethal in the laboratory but precludes flight without visibly affecting other body functions. Among others, this weak phenotype suggests the presence of redundant proteins in resilin matrices compensating lack of Pro-Resilin function. Based on published predictions, we considered the chitin-binding protein CPR56F as such a candidate. Observing a partial colocalisation of the proteins in histological experiments with tagged versions, we identified CPR56F as a resilin matrix protein that could potentially contribute to the physical properties of resilin matrices. The partial non-co-localisation of Pro-Resilin and CPR56F supports the notion that the biological function of cuticle regions with resilin may depend on the composition of the matrix. As the knock-out of cpr56f is lethal reflecting its function before the adult stage, in the present project, we were unable to further test this hypothesis. A second question that we followed in the frame of this project concerns the evolution of resilin matrices. To tackle this issue, we first examined the phenotype of pro-resilin knock-out by Crispr/Cas9 in the spotted wing fruit fly D. suzukii. We obtained an identical flight-less phenotype also in this species indicating conserved function of Pro-Resilin in flies. The work on resilin in fruit flies yielded two important publications. Using our extensive knowledge on the matter in fruit flies, we next decided to study resilin matrix incidences in other also distantly related insect species. For this purpose a detailed protocol for fluorescence microscopy of resilin matrices was developed. Using two filters for overlapping emission spectra outside the emission peak of resilin, the resilin autofluorescence signal was separated from background and unspecific information thereby enhancing especially weak signals. Applying this protocol, we found, for instance, differences of resilin incidences in the trochanter segment of the leg of a number of insect species possibly reflecting functional differences. The impact of these differences remain to be studied in locomotion assays. This work will be submitted for publication, soon. Together, these data have served to formulate a new project on resilin in the frame of the SPP 2416 (CodeChi) that has been accepted and is currently being implemented.
Projektbezogene Publikationen (Auswahl)
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Resilin matrix distribution, variability and function in Drosophila. BMC Biology, 18(1).
Lerch, Steven; Zuber, Renata; Gehring, Nicole; Wang, Yiwen; Eckel, Barbara; Klass, Klaus-Dieter; Lehmann, Fritz-Olaf & Moussian, Bernard
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Resilin is needed for wing posture in Drosophila suzukii. Archives of Insect Biochemistry and Physiology, 111(1).
Lerch, Steven; Yang, Yang; Flaven‐Pouchon, Justin; Gehring, Nicole & Moussian, Bernard
