Dataset Information

The velvet protein Vel1 controls initial plant root colonization and conidia formation for xylem distribution in Verticillium wilt

ABSTRACT: Velvet domain proteins coordinate developmental processes and secondary metabolism in fungi. In this study, we characterized the role of all four velvet proteins in the life cycle of the plant pathogenic fungus Verticillium dahliae. From other fungi, the formation of different homo-and heterodimers as well as complexes with non-velvet proteins is known. We could show that V. dahliae Vel1 is important for initial plant root colonization. In planta, Vel1 together with Vel3 is involved in fungal distribution by conidia. Disease symptom induction in tomato plants also depends on Vel1. In the senescent plant formation of microsclerotia as resting structure ex planta is controlled by Vel1, Vel2 and Vel3. In vitro protein pull-downs were conducted to identify interaction partners of velvet proteins in V. dahliae. In this study, we were able to identify interactions between Vel1 and Vel2, Vel2 and Vos1, and Vel3 and Vos1 during vegetative hyphae growth, respectively. Similar interactions are known for A. nidulans. Further interacting candidates were identified for the velvet proteins except for Vel3, for which only interaction with Vos1 was found during the tested conditions. Velvet-interacting candidate proteins included redox-regulating proteins and metabolic enzymes, but also proteins involved in energy metabolism and transport.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Verticillium Dahliae Jr2

SUBMITTER: Oliver Valerius

LAB HEAD: Prof. Dr. Gerhard H. Braus

PROVIDER: PXD021861 | Pride | 2021-02-23

REPOSITORIES: Pride

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The velvet protein Vel1 controls initial plant root colonization and conidia formation for xylem distribution in Verticillium wilt.

Höfer Annalena M AM Harting Rebekka R Aßmann Nils F NF Gerke Jennifer J Schmitt Kerstin K Starke Jessica J Bayram Özgür Ö Tran Van-Tuan VT Valerius Oliver O Braus-Stromeyer Susanna A SA Braus Gerhard H GH

PLoS genetics 20210315 3

The conserved fungal velvet family regulatory proteins link development and secondary metabolite production. The velvet domain for DNA binding and dimerization is similar to the structure of the Rel homology domain of the mammalian NF-κB transcription factor. A comprehensive study addressed the functions of all four homologs of velvet domain encoding genes in the fungal life cycle of the soil-borne plant pathogenic fungus Verticillium dahliae. Genetic, cell biological, proteomic and metabolomic ...[more]

PMID: 33720931

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Project description:The in vivo labeling technique Split-Biotin identification (Split-BioID, De Munter et al., 2017, Schopp et al., 2017, Cho et al., 2020) was used to capture the common microenvironment of Bre5 and the ribosomal protein Rps2 (uS5). Biotinylated proteins were enriched from cell lysates using affinity purification. After SDS-PAGE, proteins were digested in-gel with trypsin and the resulting peptides were analyzed by liquid chromatography-mass spectrometry (LC-MS) for identification and relative quantification against controls. An aliquot of the cell lysate that was used for the affinity purification was taken as an input control and analyzed as well by LC-MS to account for possible differences in the input abundance of enriched proteins. Stable isotope labeling with amino acids in cell culture (SILAC) was used for relative quantification of proteins according to the 2nSILAC approach (Dannenmaier et al., 2018). To evaluate the labeling efficiency, an aliquot of each culture was harvested separately prior to the pooling of the cells for the Split-BioID experiment. Cell lysates from these samples were used for SDS-PAGE, and a part of each gel lane was used for an in-gel digest of proteins. These peptides were also analyzed with LC-MS and the percentages of SILAC-labeled peptides (based on MSMS counts) were calculated. The captured proteins belong to a common cellular microenvironment of Bre5 and Rps2. References: Cho, K.F., Branon, T.C., Rajeev, S., Svinkina, T., Udeshi, N.D., Thoudam, T., Kwak, C., Rhee, H.W., Lee, I.K., Carr, S.A., Ting, A.Y. (2020). Split-TurboID enables contact-dependent proximity labeling in cells. Proc Natl Acad Sci U S A. 117, 12143-12154. Dannenmaier, S., Stiller, S.B., Morgenstern, M., Lübbert, P., Oeljeklaus, S., Wiedemann, N., Warscheid, B. (2018). Complete Native Stable Isotope Labeling by Amino Acids of Saccharomyces cerevisiae for Global Proteomic Analysis. Anal Chem 90, 10501-10509. De Munter, S., Görnemann, J., Derua, R., Lesage, B., Qian, J., Heroes, E., Waelkens, E., Van Eynde, A., Beullens, M., Bollen, M. (2017). Split-BioID: a proximity biotinylation assay for dimerization-dependent protein interactions. FEBS Lett 591, 415-424. Schopp, I.M., Amaya Ramirez, C.C., Debeljak, J., Kreibich, E., Skribbe, M., Wild, K., Béthune, J. (2017). Split-BioID a conditional proteomics approach to monitor the composition of spatiotemporally defined protein complexes. Nat Commun 8,15690.

Project description:OBJECTIVE: To assess the safety and efficacy as well as determination of the molecular mechanism of action of topical allogeneic ADSC therapy in the treatment of ulcers in patients with neuropathic diabetic foot. RESEARCH DESIGN AND METHODS: We examined a total of 57 patients with neuropathic diabetic foot in grades IA and IIA according to the University of Texas classification. Finally 23 patients received treatment with standard therapy and fibrin glue (fibrin gel group) and 23 received treatment with standard therapy and ADSC suspended in tissue glue (fibrin gel + ADSC group). The primary outcome was determining the time needed to reduce the wound size by 50% from the initial size in both groups by assessing the wound surface, the evaluation of treatment safety, expressed as the occurrence of any adverse events; the evaluation of symptoms improvement (e.g. pain, itching, etc.) assessed by Visual Analogue Scale (VAS) and analysis of the expression of selected pro-angiogenic and immunomodulatory factors in the wounds treated in both study groups. RESULTS:The ADSC group benefited from the use of stem cells in terms of accelerated wound healing and greater likelihood of complete healing of the wound. In a second week after administration of therapy, the difference between the groups was already statistically significant (p = 0,04). The time to the 50% reduction of the size of the wound in the fibrin gel + ADSC group was significantly shorter (18.9 +/- 2.2 days ) than in the group receiving fibrin gel (39.8 +/- 6.9 days ), p = 0.0035. CONCLUSIONS: The results of the preliminary analysis show that the use of allogeneic ADSCs in the treatment of neuropathic diabetic foot ulcers is a safe method of therapy, which allows to improve the effects of wound healing. Our study indicates also the association of higher expression of selected proteins expression intensifying the healing process through a faster transition to the proliferation phase with a faster reduction in wound area, and long-term efficacy of allogeneic ADSC cells.

Dataset Information

The velvet protein Vel1 controls initial plant root colonization and conidia formation for xylem distribution in Verticillium wilt

Publications

The velvet protein Vel1 controls initial plant root colonization and conidia formation for xylem distribution in Verticillium wilt.

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