DAAD-LSM doctoral projects 2018
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- The offered projects of this GSSP-application cover most areas of natural and life sciences from Anthropology and Human Biology, Cell and Developmental Biology, Genetics, Ecology, Microbiology, Molecular Biology, Biochemistry, Cell and Developmental Biology, Ecology, Evolutionary Biology, Plant Sciences, Pharmacology, Systematics.
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Project 1 (DAAD-01/18)
Supervisor: PD Dr. Bettina Bölter (Plant Sciences)
Acclimation in chloroplast protein import.
Short term acclimation of chloroplasts to various environmental triggers requires drastic changes in their proteome composition. For example, the amount of photosynthetic complexes needs to be adapted in response to varying light intensities or temperature to ensure optimal performance. Since the majority of chloroplast proteins is encoded in the nucleus and translated as preproteins in the cytosol, posttranslational targeting is a first crucial step towards providing these organelles with the required proteins. To enter the chloroplast, preproteins have to cross the double envelope membrane, which is facilitated by two complex translocation machineries responsible for preprotein recognition and transport. The import process itself is highly regulated at different levels from the outside as well as the inside of the organelle where distinct signalling cascades lead to dynamic acclimation of import activity.
In the proposed project we want to address the following questions:
I. How is thiol and redox mediated import acclimation integrated into cell signalling?
II. Which novel components participate in modulating the transport processes?
To this end, we will combine classical biochemistry and cell biology methods with novel genetic approaches.
Bölter B, Soll J, Schwenkert S. Redox meets protein trafficking. Biochim Biophys Acta (2015) 1847(9):949-56. doi: 10.1016/j.bbabio.2015.01.010
Project 2 (DAAD-02/18)
Supervisor: Prof. Dr. Angelika Böttger (Cell Biology)
Title: Developmental signalling pathways and stem cell behaviour in the cnidarian Hydra vulgaris.
Little is known about molecular mechanisms that govern stem cell proliferation and differentiation in multi-cellular animals outside the bilaterian clade. This also concerns mechanisms of tumor formation and the key question in tumor biology about the role of cancer stem cells for the initiation, maintenance and growth of tumors. Important signalling pathways involved in stem cell homeostasis have already been found in basal metazoans including Hydra, which have an evolutionary origin that is older than that of bilaterians. Therefore, one might expect to find tumors in these very simple metazoans. A phylostratigraphic approach (see link 1) has predicted that the origin of cancer lies in pre-bilaterian metazoans.
The Notch- and Wnt.-pathways belong to the most important developmental signalling pathways in animals. Canonical Wnt signalling is conserved in Hydra (see link 2). The existence of a Notch homologue, two putative Notch-ligands, the target gene Hes, Su(H) and the co-repressor groucho are also known. Further studies have shown that inhibition of the Notch-pathway in different ways leads to hyperplasia of certain cell-types in the interstitial stem cell lineage (see link 3, 4, 5). However, in these studies the pharmacological inhibitors could only be applied for short periods of time. We suggest that a permanent impairment of the Notch pathway may lead to the formation of tumor-like abnormalities. In addition, the effects of Wnt-signalling on stem cells in Hydra are completely unknown to date, but they are predicted to play a role assuming that the pathway is functionally conserved between higher animals and cnidarians.
With the suggested project we aim to induce neoplasia in the interstitial stem cell lineage of Hydra vulgaris by impairing the Notch and the Wnt-signalling pathways. Therefore, we will express and inhibit Notch and Wnt in certain cell types and at certain differentiation stages. Inducing hyperplasia in one of the conditions will give us insight into the exact cell type and differentiation stage in and at which impairment of these pathways leads to abnormal accumulations. This research will enable us to study the involvement of Notch and Wnt in the regulation of proliferation, apoptosis and differentiation of interstitial stem cells. Finally, inducing stable tumor-like abnormalities will give us the opportunity to study tumor-formation not only in its most simple form but also in a basal animal with an evolutionary origin of 1300 million years, perhaps providing us with insights about the origin of cancer.
- Link: http://www.nature.com/ncomms/2014/140624/ncomms5222/full/ncomms5222.html
- Link: http://www.nature.com/nature/journal/v407/n6801/full/407186a0.html
- Link: http://www.sciencedirect.com/science/article/pii/S0012160613004478
- Link: http://www.sciencedirect.com/science/article/pii/S0012160613004478
- Link: http://www.sciencedirect.com/science/article/pii/S0012160606013777
Domazet-Lošo T., Klimovich A., Anokhin B., Anton-Erxleben F. A., Hamm M. J., Lange C., Bosch T. C. G., Naturally occuring tumours in the basal metazoan Hydra, Nature Communications, (2014), DOI:10.1038/ncomms5222
Käsbauer T., Towb P., Alexandrova O., David C. N., Dall’Armi E., Staudigl A., Stiening B., Böttger A., The Notch signalling pathway in the cnidarian Hydra, Developmental Biologie, (2007), 303:376-390
Münder, S., Tischer, S., Grundhuber, M., Buchels, N., Bruckmeier, N., Eckert, S., Seefeldt, C.A., Prexl, A., Kasbauer, T., Böttger, A., (2013). Notch-signalling is required for head regeneration and tentacle patterning in Hydra. Developmental Biology, 383 (1): 146-57
Münder, S., Käsbauer,T., Prexl, A., Aufschnaiter, R., Zhang, X., Towb, P. and Böttger, A. (2010). Notch signalling defines critical boundary during budding in Hydra, Developmental Biology, 344(1):331-45
Tina Käsbauer, Par Towb, Olga Alexandrova, Charles N. David, Ekaterina Dall’Armi, Andrea Staudigl, Beate Stiening, and Angelika Böttger (2007): The Notch signalling pathway in the cnidarian Hydra, Developmental Biology 303(1):376-90
Project 3 (DAAD-03/18)
Supervisor: Prof. Dr. Nicolas Gompel (Evolutionary Ecology)
Title: The role of DNA shape in transcriptional regulation.
cis-regulatory elements control where, when, and how much genes are expressed, in response to the binding of transcription factors at specific DNA sites. They play a central role in defining cell identity, organizing cell and tissue interactions during embryonic development, or adapting the physiological response of an organism to changing environment. They are the nodes holding together the myriad of gene regulatory networks that rule the life of a cell. Yet, their mode of action and the determinants of their activity remain very poorly understood.
The present project aims at understanding the clockwork mechanism of a well-defined regulatory element controlling the expression of a pigmentation gene on the wing of Drosophila. The dissection of this element will rely on the use of reporter constructs, biochemistry, and bioinformatics. Two dimensions of the DNA sequence will particularly be scrutinized during the project: the chemical information given by the primary DNA sequence, as well as the structural information given by the local variation in DNA shape. These two properties of the double helix are largely independent, and the role of DNA shape in gene expression has been overlooked.
Abe, N. et al. Deconvolving the Recognition of DNA Shape from Sequence. Cell 161, 307–318 (2015).
Arnoult, L. et al. Emergence and diversification of fly pigmentation through evolution of a gene regulatory module. Science 339, 1423-1426 (2013).
Gompel, N., Prud'homme, B., Wittkopp, P. J., Kassner, V. A. & Carroll, S. B. Chance caught on the wing: cis-regulatory evolution and the origin of pigment patterns in Drosophila. Nature 433, 481-487 (2005).
Levine, M., Cattoglio, C. & Tjian, R. Looping back to leap forward: transcription enters a new era. Cell 157, 13-25 (2014)
Prud'homme, B. et al. Repeated morphological evolution through cis-regulatory changes in a pleiotropic gene. Nature 440, 1050-1053 (2006).
Rohs, R., West, S. M., Liu, P. & Honig, B. Nuance in the double-helix and its role in protein-DNA recognition. Current opinion in structural biology 19, 171-177 (2009)
Zhou, T. et al. DNAshape: a method for the high-throughput prediction of DNA structural features on a genomic scale. Nucleic acids research 41, W56-62 (2013).
Project 4 (DAAD-04/18)
Supervisor: PD Dr. Dr. Christian Grimm (Pharmacology, Molecular Biology)
Title: Endolysosomal cation channels in health and disease.
Our group is interested in the analysis of cation channels of the TRP (transient receptor potential) superfamily within the trafficking network of the endolysosomal system. Lysosomes are cell organelles involved in the breakdown of proteins, lipids, and other macromolecules.
Lysosomal dysfunction can result in endolysosomal storage disorders (LSDs) such as mucolipidoses or mucopolysaccharidoses but is also implicated in metabolic diseases, the development of neurodegenerative disorders such as Alzheimer’s and Parkinson’s disease, retinal diseases
and pigmentation disorders, trace metal deficiencies such as iron deficiency, and even cancer. Highly critical for the proper function of lysosomes, endosomes, and lysosome-related organelles (LROs) is the tight regulation of various fusion and fission processes and the regulation of proton and other cation concentrations within the endolysosomal system (ES). TRPML cation channels (TRPML1, 2 and 3) and Two-pore channels (TPCs) have recently emerged as important regulators of such processes within the ES and appear to be essential for a proper communication between the various endolysosomal vesicles. We use lysosomal patch-clamp techniques, molecular and cell biology techniques as well as knockout mouse models to study the physiological roles and activation mechanisms of these ion channels in-depth.
Grimm C, Jörs S, Saldanha SA, Obukhov AG, Pan B, Oshima K, Cuajungco MP, Chase P, Hodder P, Heller, S (2010) Small molecule activators of TRPML3. Cell Chem. Biol., 17: 135-148
Grimm C, Holdt LM, Chen C-C, Hassan S, Müller C, Jörs S, Cuny H, Kissing S, Schröder B, Butz E, Northoff B, Castonguay J, Luber CA, Moser M, Spahn S, Lüllmann-Rauch R, Fendel C, Klugbauer N, Griesbeck O, Haas A, Mann M, Bracher F, Teupser D, Saftig P, Biel M, Wahl-Schott C (2014) High susceptibility to fatty liver disease in two-pore channel 2-deficient mice. Nature Commun, 5:4699
Chen, C-C, Keller, M, Hess, M, Schiffmann, R, Urban, N, Wolfgardt, A, Schaefer, M, Bracher, F, Biel, M, Wahl-Schott, C, Grimm, C (2014) A small molecule restores function to TRPML1 mutant isoforms responsible for mucolipidosis type IV. Nature Commun, 5: 4681
Sakurai, Y, Kolokoltsov, AA, Chen, C-C, Tidwell, MW, Bauta, WE, Klugbauer, N, Grimm, C, Wahl-Schott, C, Biel, M, Davey, RA (2015) Two pore channels control Ebolavirus host cell entry and are drug targets for disease treatment, Science, 347: 995-998 top
Project 5 (DAAD-05/18)
Supervisor: PD Dr. Ralf Heermann (Microbiology)
Title: Role of AHL-LuxR solos for the life cycle of Photorhabdus bacteria.
Bacterial communication to mediate group-coordinated behavior is commonly referred to as quorum sensing. The prototypical quorum sensing system of Gram-negative bacteria consists of a LuxI-type autoinducer synthase that produces acyl-homoserine lactones (AHLs) as signals and a LuxR-type receptor that detects the AHLs to control expression of specific genes. However, many bacteria possess LuxR homologs but lack a cognate LuxI-type AHL synthase. Those LuxR-type receptors are designated as LuxR orphans or solos. Entomopathogenic bacteria of the genus Photorhabdus all harbor an extreme high number of LuxR solos. Two novel quorum sensing systems based on LuxR solos were recently found in Photorhabdus species to regulate cell clumping and therefore pathogenicity. In P. luminescens and P. temperata the LuxR solo PluR senses α-pyrones named photopyrones instead of AHLs, which are produced by the pyrone synthase PpyS. In contrast, P. asymbiotica, a closely related insect and human pathogen, has the PluR homolog PauR, which senses dialkylresorcinols produced by the DarABC pathway to regulate pathogenicity. The plenty of so called PAS4-LuxR receptors are believed to be involved in host sensing. Furthermore, all three Photorhabdus species harbour at least one LuxR solo with an intact AHL-binding motif, which might also allow sensing of exogenous AHLs. Since Photorhabdus bacteria all have a great potential in degrading AHLs, these AHL-LuxR solos could be involved in quorum quenching to block communication of other bacteria. The role of those AHL-LuxR solos for the life cycle of Photorhabdus bacteria, especially for symbiosis and pathogenicity is unknown to date.
Project 6 (DAAD-06/18)
Supervisor: Prof. Dr. Kirsten Jung (Microbiology)
Title: The role of the pyruvate-responsive BtsS/BtsR system in Escherichia coli, Salmonella typhimurium and other enteric bacteria.
Experience and expertise in microbiological, molecular and bioinformatic techniques as well as a strong interest for microscopic techniques, especially high-resolution microscopy are required.
Ude S, Lassak J, Starosta AL, Kraxenberger T, Wilson DN, Jung K (2013) Translation elongation factor EF-P alleviates ribosome stalling at polyproline stretches. Science 339: 82–85
Doerfel LK, Wohlgemuth I, Kothe C, Peske F, Urlaub H, Rodnina MV (2013) EF-P is essential for rapid synthesis of proteins containing consecutive proline residues. Science 339:85-88
Gutierrez E, Shin BS, Woolstenhulme CJ, Kim JR, Saini P, Buskirk AR, Dever TE (2013) eIF5A promotes translation of polyproline motifs. Mol Cell 51: 35–45
Lassak J, Wilson DN, Jung K (2016) Stall no more at polyproline stretches with the translation elongation factors EF-P and IF-5A, Mol Microbiol 99: 219–235
Project 7 (DAAD-07/18)
Supervisor: Prof. Dr. Andreas Klingl ( Plant Development and Electron Microscopy)
Title: Identification and localization of the S-layer proteins of two Acidithiobacillus strains SP5/1 and HV2/2
In course of several studies we could show that the Acidithiobacillus isolates SP5/1 and HV2/2, which belong to the –proteobacteria and are mesophilic and moderate thermophilic, respectively, are covered by a stripe like S-layer protein with p2-symmetry (Klingl et al., 2011). A surface or so called S-layer is a protein, which covers the cell surface and arranges itself in a regular lattice with pseudo-crystalline areas exhibiting either p1-, p2-, p3-, p4- or p6-symmetry driven by a most likely entropic process (Sleytr et al., 2007). Whereas the phylogenetic position of strain SP5/1 as a member of the species Acidithiobacillus ferrooxidans could be clarified (Kelly and Wood, 2000), the status of the isolate HV2/2 is still unclear. It might belong to the species Acidithiobacillus caldus but its ability to oxidize Fe2+ and pyrite might also lead to the description of a new species „Candidatus Acidithiobacillus striatothermus“ angebracht. The main focus of this project will not be the specific phylogenetic status of SP5/1 and HV2/2 but on the common feature of both strains which they also share with other Acidithiobacillus species: the stripe-like S-layer (Fig. 1). Within the project, the protein should be isolated, purified and localized via specific antibodies. The identification of the S-layer protein itself should finally if it shows similarities to Afe_2303 which is mentioned as a potential S-layer protein in the type strain of A. ferrooxidans.
Fig. 1: Transmission electron micrographs of freeze fractured and freeze etched cells of strain Sp5/1 (A-C), HV2/2 (E) and an S-layer deficient lab strain ΔSlp90 (D). Scale bars: 500 nm (A, D, E) and 200 nm (B, C). Also see Klingl et al. (2011).
Kelly D.P. and Wood A.P. (2000). Reclassification of some Thiobacillus to the newly designated genera Acidithiobacillus gen. nov., Halothiobacillus gen. nov. and Thermithiobacillus gen. nov. International Journal of Systematic and Evolutionary Microbiology 50: 511-516.
Klingl A., Moissl-Eichinger C., Wanner G., Zweck J., Huber H., Thomm M. and Rachel R. (2011). Analysis oft he surface proteins of Acidithiobacillus ferrooxidans strain SP5/1 and the new, pyrite-oxidizing Acidithiobacillus isolate HV2/2, and their possible involvement in pyrite oxidation. Archives of Microbiology 193: 867-882. Doi: 10.1007/s00203-011-0720-y
Sleytr U.B., Huber C., Ilk N., Pum D., Schuster B. and Egelseer E.M. (2007). S-layers as a tool kit for nanobiotechnical applications. FEMS Microbiological Letters 267: 131-144.
Project 8 (DAAD-08/18)
Supervisor: Dr. Macarena Marin (Cell biology, Biochemistry, Genetics)
Title: Symbiotic reprogramming of plant cell by rhizobia effectors.
Effectors of plant-associated bacteria are proteins secreted directly into the plant cell cytoplasm. Here they interact with host targets thereby subverting host-signaling cascades to promote their own fitness, for example by undermining the plant immune system. The molecular functions of a sweet of phytopathogenic effectors have been investigated. However our understanding of symbiotic effectors is limited, although the vast majority of all known symbiotic bacteria harbor effector secretion systems.
The aim of this project is to study how rhizobia effectors target pathways in the host cell leading to symbiotic reprogramming. The selected student will use molecular, cell-biological and biochemical methods to elucidate the molecular function of effectors inside of the plant cell and characterize their interactions with host target proteins that have been identified in our group. Applicants should ideally have expertise in biochemistry and molecular biology and a strong interest in organismic interactions and plant sciences.
1. Oldroyd, G. E., et al (2011). "The rules of engagement in the legume-rhizobial symbiosis." Annu Rev Genet 45: 119-144.
2. Miwa, H. and Okazaki, S. (2017). “How effectors promote beneficial interactions.” Curr Op Plant Sci 38:148-154.
For further information see: http://www.genetik.bio.lmu.de/research/marin/index.html
Project 9 (DAAD-09/18)
Supervisor: PD.Dr. Stylianos Michalakis (Gene Therapy)
Title: Novel gene therapy approaches for inherited eye diseases.
The group works on the development of recombinant adeno-associated virus (AAV) vector-based gene therapies for eye diseases and is part of the RD-CURE consortium that successfully initiated the first ocular gene therapy trial on CNGA3-linked achromatopsia and currently prepares for the initiation of the first German retinitis pigmentosa gene therapy trial. AAV vectors have become the “gold standard” gene delivery tool for targeting retinal cells. AAV-based vectors have been tested in a number of human clinical trials and have proven safe for the use in the human eye. However, there is still unmet need for improvement of the AAV vector platform regarding tropism and transduction efficiency.
Specific aims and methodology. The overarching goal of this project is the development of novel optimized AAV vector-based gene therapy vectors for gene-specific and/or gene independent treatment approaches. The novel vectors will be further developed for optimized efficacy in relevant animal models of inherited blinding diseases. The methodology will include state-of-the-art in vitro and in vivo biochemical, genetic, cell biological and viral gene transfer methods in relevant mouse models. Potential candidates should have a strong interest and background in mouse physiology and anatomy (focus on the eye), molecular biology and gene therapy.
Further information and selected literature:
R. Mühlfriedel, N. Tanimoto, C. Schön, V. Sothilingam, M. Garcia Garrido, S. C. Beck, G. Huber, M. Biel, M. W. Seeliger, S. Michalakis, Front Neurosci 2017, 11, 292.
S. Koch, V. Sothilingam, M. Garcia Garrido, N. Tanimoto, E. Becirovic, F. Koch, C. Seide, S. C. Beck, M. W. Seeliger, M. Biel, R. Mühlfriedel, S. Michalakis, Hum Mol Genet 2012, 21, 4486-4496.
S. Michalakis, R. Mühlfriedel, N. Tanimoto, V. Krishnamoorthy, S. Koch, M. D. Fischer, E. Becirovic, L. Bai, G. Huber, S. C. Beck, E. Fahl, H. Büning, F. Paquet-Durand, X. Zong, T. Gollisch, M. Biel, M. W. Seeliger, Mol Ther 2010, 18, 2057-2063
Project 10 (DAAD-09/18)
Supervisor: PD Dr. Stylianos Michalakis (Epigenetics)
Title: Role of TET-mediated 5mC oxidation for neuronal differentiation and plasticity.
Scientific background. Neuronal networks show a remarkable degree of plasticity during physiological and pathophysiological processes. This plasticity goes along with major adjustments in the expression of key genes. The mechanisms controlling gene expression and neuronal plasticity are not well understood, but it is suggested that epigenetic mechanisms such as DNA methylation contribute crucially to these biological processes. Methylation of the DNA base cytosine is catalyzed by DNA methyltransferases (DNMT) and occurs at the C-5 position of the cytosine base resulting in 5-methylcytosine (5mC). Removal of the methyl group involves oxidation by TET methylcytosine dioxygenases. The overarching goal of this project is to help improving our understanding on how TET enzymes and 5mC oxidation products shape the epigenome of neurons and influence CNS function.
Specific aims and methodology. The functional significance of TET proteins and their enzymatic products in the CNS has not been characterized and will be addressed in this project with specific genetic mouse and cellular models. TET enzymes act in concert with chromatin remodeling proteins and transcription factors. We identified intriguing novel TET interaction partners in mouse retina and induced pluripotent stem cell (iPSC)-derived bipolar neurons. The potential of these proteins to engage with TET3 and modulate its enzymatic activity will be assessed in this proposal. The methodology will include state-of-the-art in vitro and in vivo biochemical, genetic, cell biological and viral gene transfer methods in relevant mouse models. Potential candidates should have a strong interest and background in neurobiology, molecular biology and epigenetics.
Further information and selected literature:
M. Wagner, J. Steinbacher, T. F. Kraus, S. Michalakis, B. Hackner, T. Pfaffeneder, A. Perera, M. Müller, A. Giese, H. A. Kretzschmar, T. Carell, Angew Chem Int Ed 2015, 54, 12511-12514.
A. Perera, D. Eisen, M. Wagner, S. K. Laube, A. F. Künzel, S. Koch, J. Steinbacher, E. Schulze, V. Splith, N. Mittermeier, M. Müller, M. Biel, T. Carell, S. Michalakis, Cell Rep 2015, 11, 283-294.
T. Pfaffeneder, F. Spada, M. Wagner, C. Brandmayr, S. K. Laube, D. Eisen, M. Truss, J. Steinbacher, B. Hackner, O. Kotljarova, D. Schuermann, S. Michalakis, O. Kosmatchev, S. Schiesser, B. Steigenberger, N. Raddaoui, G. Kashiwazaki, U. Müller, C. G. Spruijt, M. Vermeulen, H. Leonhardt, P. Schar, M. Müller, T. Carell, Nat Chem Biol 2014, 10, 574-581
Project 11 (DAAD-10/18)
Supervisor: Dr. Tamara Mikeladze-Dvali (Cell Biology)
Title: The function of centrioles/basal bodies for neurite anchoring during C. elegans mophogenesis.
Centrioles carry two main functions: in proliferating cells centrioles recruit peri-centriolar material and act as microtubule organizing centers, which mediate bipolar spindle formation during mitosis; in differentiating cells centrioles become basal bodies and are essential for the formation cilia and flagella (Fig. 1. and 2.). Abnormalities in centriole biogenesis are associated with multiple disorders ranging from microcephaly to ciliopathies and tumor formation. Thus, it is essential that centrioles and basal bodies are tightly regulated according to their cellular context. Our major objective is to uncover molecular mechanisms, by which centrioles and centrosomes are regulated in a tissue specific manner during the development of a multicellular organism.
Sensory neurons in the head of C. elegans, which are born during morphogenesis, form their neurites by a mechanism called retrograde extension. During retrograde extension the ciliary transition zone (or basal body) attaches the tips of the cells at the mouth region of the worm and thereby anchors the neurons (Fig. 3.). The cell body moves away from the anchorage point and consequently the neurite extends (1,2).
The aim of this project is to elucidate the molecular mechanism by which centrioles/basal bodies are targeted to the prospective site of neurite anchoring. Our preliminary data suggest that junctional complexes and polarity factors might be involved in this process. The main goal of the PhD project will be to understand which of these factors are involved in the process of centriole targeting and how exactly centrioles are positioned in prospective location of anchoring. The project will involve a variety of molecular and genetic techniques, as well as imaging using state of the art microscopes.
1. Heiman, M. G. and Shaham, S. DEX-1 and DYF-7 establish sensory dendrite length by anchoring dendritic tips during cell migration. Cell (2009); 137(2)
2. Schouteden, C., Serwas, D., Palfy, M., and Dammermann, A.. The ciliary transition zone functions in cell adhesion but is dispensable for axoneme assembly in C. elegans. J Cell Biol. (2015); 210(1)
Project 12 (DAAD-11/18)
Supervisor: Prof. Dr. Jörg Nickelsen (Plant Sciences, Biochemistry, Genetics)
Title: Chlorophyll Incorporation into Photosynthetic Complexes
Thylakoids mediate photosynthetic electron transfer and represent one of the most elaborate energy-transducing membrane systems. Despite our detailed knowledge of its structure and function, much remains to be learned about how the machinery is put together. The concerted synthesis and assembly of lipids, proteins and low-molecular-weight cofactors like pigments and transition metal ions requires a high level of spatiotemporal coordination. While increasing numbers of assembly factors are being functionally characterized, the principles that govern how thylakoid membrane maturation is organized in space are just starting to emerge. In both cyanobacteria and chloroplasts, distinct subcellular production lines for the fabrication of photosynthetic complexes, in particular photosystem II (PSII), have been identified.
In background work, the Nickelsen group has identified OHP2, a member of the LHC superfamily in the green alga Chlamydomonas reinhardtii, as a likely candidate involved in delivering Chl during assembly and/or repair of PSII, in particular to its D1 subunit.
By applying advanced molecular genetics, imaging and biochemical approaches, the suborganellar localization of OHP2, its function during photoprotection, putative interactions partners, and evolutionary conservation will be addressed. Pigment binding and structure of OHP2 will be determined together with international cooperation partners.
Fig.1: PSII assembly and repair in chloroplasts. A possible function of OHP2 in chlorophyll insertion is indicated.
Stengel, A., Gügel, I., Hilger, D., Rengstl, B., Jung, H. and Nickelsen, J. (2012) Initial steps in photosystem II assembly and preloading with manganese take place in biogenesis centers in Synechocystis. Plant Cell 24, 660-675.
Nickelsen, J. and Rengstl, B. (2013) Photosystem II assembly: From cyanobacteria to plants. Annu. Rev. Plant Biol. 64, 609-635.
Bohne, A.-V., Schwarz, C., Schottkowski, M., Lidschreiber, M., Piotrowski, M., Zerges, W. and Nickelsen, J. (2013) Reciprocal regulation of protein synthesis and carbon metabolism for thylakoid membrane biogenesis. PLoS Biol. 11, e1001482.
Rast, A., Heinz, S., Nickelsen, J. (2015) Biogenesis of thylakoid membranes. Biochim. Biophys. Acta, DOI: 10.1016/j.bbabio.2015.01.007. [Epub ahead of print].
Project 13 (DAAD-12/18)
Supervisor: Prof. Dr. Kai Papenfort (Microbiology)
Title: Autoinducer-controlled riboswitches in commensal and pathogenic bacteria.
Quorum sensing (QS) is the process of cell-cell communication among microbes (1). QS involves the production, release, and detection of extracellular signaling molecules called autoinducers, and allows bacteria to collectively regulate gene expression in response to changes in cell density and species composition of the vicinal community. Many, if not all, bacteria use QS to regulate gene expression, and this process is intimately linked to pathogenesis and biofouling (2). Understanding QS is fundamental to all of microbiology, and ultimately could prove to be key to understanding other collective behaviors, including those of higher organisms.
Autoinducers bind to cognate protein receptors, which are either transcription factors or signaling proteins. In this project, we aim to study the role of the recently discovered DPO-mediated QS system in Vibrio cholerae (3). Specifically, we will explore if and how DPO controls the expression of virulence-related gene expression, and which protein and RNA factors participate in this process.
In this project, the doctoral student will employ state-of-the-art RNA sequencing technology (4) to determine the role of DPO on gene expression in V. cholerae. Follow-up experiments will aim at the understanding of the molecular mechanisms underlying DPO-mediated gene control focusing on the role of transcription factors and regulatory RNAs in this process.
1. Papenfort, K. and Bassler, B.L. (2016) Quorum sensing signal-response systems in Gram-negative bacteria. Nat Rev Micro, 14, 576-588.
2. Dong, Y.H., Wang, L.H., Xu, J.L., Zhang, H.B., Zhang, X.F. and Zhang, L.H. (2001) Quenching quorum-sensing-dependent bacterial infection by an N-acyl homoserine lactonase. Nature, 411, 813-817.
3. Papenfort, K., Silpe, J.E., Schramma, K.R., Cong, J.P., Seyedsayamdost, M.R. and Bassler, B.L. (2017) A Vibrio cholerae autoinducer-receptor pair that controls biofilm formation. Nat Chem Biol, 13, 551-557.
4. Papenfort, K., Forstner, K.U., Cong, J.P., Sharma, C.M. and Bassler, B.L. (2015) Differential RNA-seq of Vibrio cholerae identifies the VqmR small RNA as a regulator of biofilm formation. Proc Natl Acad Sci U S A, 112, E766-775.
Project 14 (DAAD-13/18)
Supervisor: Dr Arne Weiberg (Genetics, Biochemistry, Plant Sciences)
Title: The role of small RNAs in shaping the plant microbiome.
Probiotic microbial communities are valuable determinants of health and performance in animals and plants. Phylogenomics analysis of the Arabidopsis microbiome revealed probabilistic composition of communities that is associated with the occurrence of key species that are proposed to act as microbial hubs influencing the synthesis of plant-associated bacterial communities (1,2). However, how communication in plant-microbe and microbe-microbe interaction establishes and maintains these plant microbiome communities and how this leads to plant beneficial traits is currently not understood. Deepen our knowledge of the molecular mechanisms will open new opportunities of synthetic communities to promote plant health and robustness.
Small RNAs are non-coding RNAs that regulate gene expression in a mechanism called RNA interference (RNAi). RNAi plays a pivotal role in plant-microbe interaction, contributing to the activation of host immunity as well as pathogen virulence. Recent discoveries demonstrated that small RNAs are exchanged bi-directionally in pathogenic fungus Botrytis-plant interaction to induce cross-kingdom RNAi (3,4). While pathogen small RNAs attack the host plant immunity, synthetic RNAs can be engineered to target pathogen genes that confer plant resistance.
The aim of this study is to disentangle the role of small RNAs in shaping the plant phyllosphere microbiome. In particular, we are interested in the function of small RNA-based inter-kingdom communication during plant-microbe and microbe-microbe interaction. We propose to study RNA communication using Arabidopsis as our model plant in combination with two distinct Arabidopsis pathogens, Botrytis cinerea and the oomycete pathogen Hyaloperonospora arabidopsidis, both use small RNAs to suppress plant immunity. The doctoral candidate will study the influence of deconstruction and reconstruction of microbiome structures on the disease outcome. By profiling small RNAs using high-throughput sequencing methods we will learn how microbes try to influence each other and their host plant via cross-kingdom RNAi, and how this channel of communication shapes the plant microbiome.
1. Hacquard S, Spaepen S, Garrido-Oter R, Schulze-Lefert P.: Interplay Between Innate Immunity and the Plant Microbiota. Annu Rev Phytopathol, 2017, 55:565-589.
2. Agler MT, Ruhe J, Kroll S, Morhenn C, Kim ST, Weigel D, Kemen EM.: Microbial Hub Taxa Link Host and Abiotic Factors to Plant Microbiome Variation. PLoS Biol, 2016, doi: 10.1371/journal.pbio.1002352
3. Wang M, Weiberg A, Lin FM, Thomma BP, Huang HD, Jin H.: Bidirectional cross-kingdom RNAi and fungal uptake of external RNAs confer plant protection. Nat Plants, 2016, doi: 10.1038/nplants.2016.151
4. Weiberg A, Wang M, Lin FM, Zhao H, Zhang Z, Kaloshian I, Huang HD, Jin H.: Fungal small RNAs suppress plant immunity by hijacking host RNA interference pathways. Science, 2013, 342:118-23.
Project 15 (DAAD-14/18)
Supervisor: Dr. Esther Zanin (Cell Biology)
Title: Molecular mechanisms controlling polar body cytokinesis.
During meiosis the diploid genome is segregated into haploid gametes. If meiosis fails, the embryo will inherit an abnormal chromosome number and die. Infertility in humans is often caused by defects in meiosis and therefore understanding the molecular mechanisms of meiosis will help to develop novel therapeutic strategies to treat infertility. Female meiotic divisions are highly asymmetric generating two small polar bodies and a large ovum1. One great model system to study female meiosis is the small nematode C. elegans since fertilized embryos can be easily obtained for live-cell microscopy and many molecular and genetic tools are well established in this model organism.
Like in vertebrates, in C. elegans the meiotic spindle aligns perpendicular to the plasma membrane during metaphase (Fig. 1). Firstly, the cell cortex adjacent to the spindle pole is remodeled and a premature contractile ring assembles2. Secondly, in anaphase the contractile ring ingresses from the cell surfaces along the meiotic spindle. Than contractile ring ingression stops half way between the segregating chro-mosomes at the spindle midzone. Finally the contractile ring constricts and the polar body is abscised. To ensure the correct segregation of the genetic content the ingression and constriction of the contractile ring as well as the abscission of the polar body has to be spatially and temporally coordinated with chromosome segregation and cell cycle progression. How this complex control is achieved on the molecular level is only poorly understood at present and the PhD project will address this question. During mitotic cytokinesis the molecular pathways that control contractile ring formation are well established3. Mitotic and meiotic cytokinesis share many similarities but they also differ: for example, in mitotic cytokinesis contractile ring formation is induced in anaphase but not in metaphase. Further the mitotic anaphase spindle has centrosomal microtubule asters whereas the meiotic spindle is acentriolar. It has been shown that some mitotic cytokinesis regulators are required for polar body cytokinesis2,4 however at which step and how they function has not been determined. Due to the morphological differences between meiotic and mitotic cytokinesis it is likely that the communication between the spindle and the cell cortex also differs. The goal of our laboratory is to reveal the regulatory network that controls cytokinesis during cell division. Towards this goal we combine several complementary approaches in the nematode C. elegans and human tissue culture cells. We aim to identify signaling complexes involved in cytokinesis by biochemical approaches as well as by genome-wide microscopy-based screens. Signaling complexes involved in cytokinesis often have multiple functions during the cell division. To disrupt their function specifically during cytokinesis or in a specific region of the cell we will also develop light-controllable compounds in collaboration with the group of H. Dube. The function of the identified signaling complexes is than investigated in vivo by quantitative live-cell microscopy. The proposed PhD project will use a variety of microscopy-based, biochemical and screening techniques to dissect the molecular pathways controlling polar body cytokinesis.
1. Maddox, A. S., Azoury, J. & Dumont, J. Polar body cytokinesis. Cytoskeleton (Hoboken) 69, 855–868 (2012).
2. Dorn, J. F. et al. Actomyosin Tube Formation in Polar Body Cytokinesis Requires Anillin in C. elegans. Current Biology 20, 2046–2051 (2010).
3. Green, R. A., Paluch, E. & Oegema, K. Cytokinesis in Animal Cells. Annu. Rev. Cell Dev. Biol. 28, 29–58 (2012).
4. Fabritius, A. S., Flynn, J. R. & McNally, F. J. Initial diameter of the polar body contractile ring is minimized by the centralspindlin complex. Developmental Biology 359, 137–148 (2011).top