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Movement is vital to all living organisms, from the transport of cellular organelles to the movement of entire organisms. Sarcomeres are the smallest contractile units of striated skeletal and heart muscles. The Z-disc is the attachment region for adjacent sarcomeres and plays a pivotal role not only in sustaining muscle architecture but also in signalling, mechanosensing and mechanotransduction, and protein turnover and autophagy.
We are interested in the molecular mechanisms underlying the architecture and assembly of the Z-discs to address the questions: how are Z-disc proteins assembled in highly ordered, albeit plastic complex able to transmit force along myofibrils? What is the hierarchy and cooperativity of interactions in sarcomeric biogenesis? What is the role of PTMs and how disease mutations underlying (cardio)myopathies disrupt this finely tuned assembly?
Other research lines focus on studies of metallo-enzymes involved in protection from chemical and oxidative damage.
In order to dissect the Z-disc structure and assembly at molecular level we are using a combination of biochemical, molecular biophysics and complementary structural biology methods on reconstituted complexes and their individual components. This blend of approaches depicts biochemical and biophysical properties of studied macromolecules and of their interactions, and gives structural information on different scales of resolution - from atomic and molecular to ultrastructural. These structural and functional data are in turn validated in vitro and in cellula, by structure informed structural, molecular and cell biophysics studies (the latter in collaboration). Finally, combining these diverse experimental data for integrative modelling to generate structural models of mini Z-disc assembles.
We are developing methods for results rectification, optimization of protein generation, and crystallization, and for analysis of intrinsically disordered proteins with solution scattering.
Kristina Djinovic Carugo studied chemistry at University of Ljubljana, Slovenia, before receiving her PhD in 1992 in Structural biochemistry in at university of Ljubljana with work performed at University of Pavia, Italy. In 1995 she moved to European Molecular Biology Laboratory in Heidelberg, first as a postdoc and then as a staff scientist. In 1999 she started her own lab as Head of the Structural Biology Laboratory at Elettra - Sincrotrone Trieste, Italy. Since 2004 she is a full professor of molecular structural biology at University of Vienna and since 2009 chair of the Department of structural and computational biology.
We determined the first complete structure of α-actinin from striated muscle and explored its functional implications on the biochemical and cellular level. The structure provides insight into the mechanism of α-actinin regulation by PIP2 and the intramolecular pseudoligand autoinhibition mechanism. It also offers a foundation to study interactions with Z-disc partners and effects of pathogenic mutations with molecular resolution and mechanistic insight. (Ribeiro et al., Cell, 2014).
Read more: soft link to paper - https://doi.org/10.1016/j.cell.2014.10.056
Actin is one of the most abundant proteins in eukaryotic cells. Actin filaments together with a large number of actin-binding proteins are critical players in many cellular functions, ranging from cell motility and muscle contraction to maintenance of cell shape and transcription regulation. α-Actinin—a member of the spectrin superfamily—is an archetypical F-actin–binding and –bundling protein. It is known that Ca2+ inhibits α-actinin capacity to bundle F-actin. We present a structure of a Ca2+ -regulated α-actinin and propose the mechanism for its regulation. We uncover that Ca2+ binding triggers an increase in protein rigidity, leading to reduced conformational flexibility and bundling activity. The proposed molecular mechanism is likely to be a blueprint for regulation of spectrin-like proteins (Pinotsis et al., PNAS, 2020).
Read more: soft link to paper - https://www.pnas.org/content/117/36/22101
In sarcomeres, α-actinin crosslinks actin filaments and anchors them to the Z-disk. FATZ proteins interact with α-actinin and five other core Z-disk proteins, contributing to myofibril assembly and maintenance as a protein interaction hub.
Here we report the first structure and its cellular validation of α-actinin-2 in complex with a Z-disk partner, FATZ-1, which is best described as a conformational ensemble. We show that FATZ-1 forms a tight fuzzy complex with α-actinin-2 and propose a molecular interaction mechanism via main molecular recognition elements and secondary binding sites. The obtained integrative model reveals a polar architecture of the complex which, in combination with FATZ-1 multivalent scaffold function, might organise interaction partners and stabilise α-actinin-2 preferential orientation in the Z-disk.
Finally, we uncover FATZ-1 ability to phase-separate and form biomolecular condensates with α-actinin-2, raising the intriguing question whether FATZ proteins can create an interaction hub for Z-disk proteins through membrane-less compartmentalization during myofibrillogenesis (Sponga et al., Sci Adv, 2021).
Read more: soft link to paper - https://www.science.org/doi/10.1126/sciadv.abg7653
The Z-disc is the boundary between adjacent sarcomeres, the basic contractile units of striated muscles. Myotilin is a Z-disc scaffold protein, providing structural integrity by multiple interactions, including F-actin and α-actinin-2. In our study, we provide the first integrative structural model of its complex with F-actin. We further show that myotilin displaces tropomyosin from F-actin, implying a novel role of myotilin in sarcomere biogenesis beyond an interaction hub for Z-disc partners (Kostan et al., PLOS Biol, 2021).
Read more: soft link to paper - https://doi.org/10.1371/journal.pbio.300114
The major bottleneck in macromolecular crystallography is the production of well-diffracting crystals. To increase the crystallization success, we developed a crystallization strategy customizable for each protein. The underlying concept is to use in crystallization cocktails compounds that increase the thermal stability of the protein. This customized strategy yielded not only twice as many crystal hits as the standard approach but also crystal hits for two additional proteins compared to the benchmark workflow. The inherently simple design and the modular and flexible nature of the platform makes it easy to modify, further develop, and optimize individual steps. The information gained can also be used to increase the monodispersity and stability of proteins, improving in this way their amenability for biochemical studies and eventually derive a possible function of not yet annotated proteins (Mlynek et al., Cryst Growth Des, 2020).
Read more: soft link to paper - https://dx.doi.org/10.1021/acs.cgd.9b01328
Stable anchoring of titin in muscle Z-disc is essential for muscle integrity during stretching. α-Actinin provides attachment points through binding to Z-repeats of titin. In collaboration with M. Rief, we used optical tweezers to study the mechanics of this interaction at a single-molecule level. We suggest a model where multiple weak α-actinin/Z-repeat interactions co-operate to ensure a stable titin anchoring while allowing for dynamic exchange of components (Grison et al., PNAS, 2017).
Read more: soft link to paper - https://www.embopress.org/doi/full/10.15252/embr.201439267
Cellular and organelle shape can remarkably modulate cellular function. Actin polymerization together with proteins that directly deform membranes, such as the BAR superfamily proteins have been implicated in regulation of membrane shape. Using a combination of structural (MX, cryo-EM), biochemical and cell biophysics approaches we showed that F-BAR domain of pacsin2 binds actin filaments using the same concave surface employed to also bind to membranes (Kostan et al., EMBO Rep, 2014).
Read more: soft link to paper - https://doi.org/10.15252/embr.201439267
Order from disorder in the sarcomere: FATZ forms a fuzzy but tight complex and phase-separated condensates with α-actinin.
Sponga, Antonio; Arolas, Joan L; Schwarz, Thomas C; Jeffries, Cy M; Rodriguez Chamorro, Ariadna; Kostan, Julius; Ghisleni, Andrea; Drepper, Friedel; Polyansky, Anton; De Almeida Ribeiro, Euripedes; Pedron, Miriam; Zawadzka-Kazimierczuk, Anna; Mlynek, Georg; Peterbauer, Thomas; Doto, Pierantonio; Schreiner, Claudia; Hollerl, Eneda; Mateos, Borja; Geist, Leonhard; Faulkner, Georgine; Kozminski, Wiktor; Svergun, Dmitri I; Warscheid, Bettina; Zagrovic, Bojan; Gautel, Mathias; Konrat, Robert; Djinovi?-Carugo, Kristina
Molecular basis of F-actin regulation and sarcomere assembly via myotilin.
Kostan, Julius; Pavsic, Miha; Puz, Vid; Schwarz, Thomas C; Drepper, Friedel; Molt, Sibylle; Graewert, Melissa Ann; Schreiner, Claudia; Sajko, Sara; van der Ven, Peter F M; Onipe, Adekunle; Svergun, Dmitri I; Warscheid, Bettina; Konrat, Robert; Fürst, Dieter O; Lenarcic, Brigita; Djinovic-Carugo, Kristina
Calcium modulates the domain flexibility and function of an α-actinin similar to the ancestral α-actinin.
Pinotsis, Nikos; Zielinska, Karolina; Babuta, Mrigya; Arolas, Joan L; Kostan, Julius; Khan, Muhammad Bashir; Schreiner, Claudia; Salmazo, Anita; Ciccarelli, Luciano; Puchinger, Martin; Gkougkoulia, Eirini A; Ribeiro, Euripedes de Almeida; Marlovits, Thomas C; Bhattacharya, Alok; Djinovic-Carugo, Kristina
Tailored Suits fit better: Customized Protein Crystallization Screens
Mlynek, Georg; Kostan, Julius; Leeb, Sarah; Djinovic-Carugo, Kristina
The Structure and Regulation of Human Muscle α-Actinin.
Ribeiro, Euripedes de Almeida; Pinotsis, Nikos; Ghisleni, Andrea; Salmazo, Anita; Konarev, Petr V; Kostan, Julius; Sjöblom, Björn; Schreiner, Claudia; Polyansky, Anton A; Gkougkoulia, Eirini A; Holt, Mark R; Aachmann, Finn L; Zagrović, Bojan; Bordignon, Enrica; Pirker, Katharina F; Svergun, Dmitri I; Gautel, Mathias; Djinović-Carugo, Kristina
Since 10/2021: Liquid-liquid Phase Separation in Biology – Ellipse, funded by the Austrian Science Fund FWF.
Building, learning and research capacities in the structural and functional analysis of biomolecules for the needs of biomedicine and biotechnology. Total grant Volume: 1.166.798,81 €
05/19 - 04/22
Since 6/2018: Exploring comammox: from molecules to ecology and applications, funded by University of Vienna.
03/2018 – 08/2021: Structure ZOOM
2016-2020: An integrated approach to the muscle Z-disk: from atomic structure to human disease (with M. Gautel, P. Elliott, H. Watkins, S. Raunser and K Gehmlich)
2016-2019: The Group Djinovic participates in the special Doctoral Program "Integrative Structural Biology" reviewed and funded by the Austrian Science Fund FWF. Kristina Djinovic-Carugo is Vice Speaker of the program.
2009-2013: Doctoral Program "Structure and Interaction of Biological Macromolecules" reviewed and funded by the Austrian Science Fund FWF.
2014-2016: Laura-Bassi Center of Expertise "COSS - Center for Optimized Structural Studies" funded by the Austrian Research Promotion Agency FFG (Co-ordinator)
2010-2013: Laura-Bassi Center of Expertise "COSS - Center for Optimized Structural Studies" funded by the Austrian Research Promotion Agency FFG (Co-ordinator)
2009-2013: Research Network funded by the GEN-AU Genome Research Program of the Federal Ministry of Science and Research
2014-2017: "Structure and regulation of the Myofibrillar Z-disc Interactome" funded by the German Research Foundation and the Austrian Science Fund
2010-2014: Research Unit 'Alpha Actinin, Filamin C and their Complexes with Binding Partners' funded by the German Research Foundation and the Austrian Science Fund
2009-2014: FP7 Marie-Curie Action ITN Muscle Z-disk Protein Complexes: from atomic structure to physiological function (MUZIC)
2014-2017: MORN “Structure and Function of the Trypanosoma brucei bilobe”
2010-2013: MORN repeat proteins and the Trypanosoma brucei bilobe (with G. Warren)
2010-2013: Structural Aspects of RAF-1:Rok-alpha Interaction (with M. Baccarini)