Intermediate filaments as signaling determinants
Intermediate filaments (IFs) form structures that are essential for providing mechanical strength to cells. Interestingly, recent data point to an important function of IFs in cell signaling as scaffolding of critical signaling molecules. Our lab focuses on studying the signaling-mediated regulation of IFs as well as the role of IFs as signaling modulators in differentiation, cell stress and cancer.
Funding: Academy of Finland and Sigrid Jusélius Foundation
Vimentin in cell size, growth and migration
Vimentin is a type III intermediate filament protein expressed abundantly in mesenchymal cells, and it is known to be a marker for epithelial to mesenchymal transition. Our current research is focusing on vimentin’s role in physiological and pathological conditions, using both in vitro and in vivo models. We have recently published that “Vimentin regulates EMT signaling and fibroblast proliferation during the wound healing process” in PNAS 2016. We found that vimentin plays role in wound healing mechanism, by regulating cell proliferation, EMT, growth signaling and migration.
Cell proliferation and growth are two coordinated processes that are essential for proper development and body homeostasis. These processes directly influence wound healing and cancer. We have recently found that vimentin regulates cell size through the insulin/Akt/mTOR signaling and, currently, we are trying to understand the molecular mechanism by which vimentin regulates this pathway. Cell migration is a critical step during wound healing processes and we found that vimentin plays a role in regulating migration mechanisms. Major in vivo vimentin phosphorylation sites have been already identified by our lab in 2004 and the physiological functions of these sites are being studied. However, their role in cell migration is still poorly understood, so we are now focused on understanding which phosphosites are related to cell migration and the underlying molecular mechanisms regulated by them.
Post-translational modifications (PTMs) as regulators of IF structure and function
Vimentin is one of the most abundant of intermediate filaments, being mostly expressed in various cells of mesenchymal origin. Previous research has established that phosphorylation is a major regulator of dynamics, assembly, organization and functions of vimentin. However, it has been unclear which of the multiple in vivo phosphorylation sites are the most crucial for the assembly and organization of vimentin intermediate filament networks. To address this question, we analyzed phosphomimetic aspartate and phosphorylation-deficient alanine point mutations corresponding to the phosphorylated or dephosphorylated states of previously identified vimentin serine and threonine interphase in vivo phosphorylation sites. The analysis shows that multiple sites in the far N-terminal serines cluster of Ser 6, 7, 8, 9 have the most prominent, dominant, effects on the filamentous structure of vimentin while Ser 71, 72 has smaller and other phosphorylation sites do not seem to have major structural roles. A phospho-mimicking mutant of vimentin Ser 4, 6,7,8,9 broke vimentin down to tetrameric soluble state with aggregates in part of the cells. A phosphorylation-deficient mutant of these sites was able to form a limited amount of filaments but mostly aggregates indicating that some phosphorylation on these sites may be needed to stabilize network and may offer a new layer of regulation. The phosphorylation state of these sites also affected vimentin cytoskeletal associations. Based on our results and previous research it is likely that vimentin nonapeptide motif binds to rod domain 2B and phosphorylation of Ser(6,)7,8,9 has a critical effect on this interaction and completely or partially limit vimentin assembly to soluble tetrameric state. Future goal is to study the functional consequence of these phosphorylation sites in cell growth, cell migration and other cellular processes.
Vimentin as a signaling gatekeeper in injury
We propopse that vimentin, the type III cytoskeletal intermediate filament and a universal mesenchymal marker, may behave as a signaling scaffold in response to stress and upon injury. By collaboration with other intermediate filaments, for instance keratin upon the epithelium injury, nestin and desmin upon muscle injury and GFAP, nestin during the neuron damage, vimentin modify various cell fate and cellular activities and consequently lead to re-epithelialization, muscle regeneration or gliosis to maintain tissue integrity. Our current research is focusing on vimentin’s role in epithelium injury under physiological and pathological conditions using a variety of in vitro and in vivo model systems.
Characterization of Vim and Viml KO zebrafish phenotypes in terms of wound healing
Zebrafish is an interesting model organism to study the molecular mechanisms underlying wound healing using imaging, thanks to the transparency and fast developmental stages of zebrafish embryos. In collaboration with Ilkka Paatero of Ivaska Lab, we are employing knock – out zebrafish lines of Vim and Viml genes to unravel yet to be understood phenotypes associated with wound healing. Focus is given on macrophage and zebrafish migration and as well as endothelial sprouting in response to embryo, tail wound healing. Using high-resolution microscopy the migration pattern of macrophages, fibroblasts and endothelial cells in terms of speed and directionality are studied. Apart from that, the extent and speed of regeneration of cut or amputated embryo tail is also studied.
Responsible Team Members
- Hend Abdelkader
- Ponnuswamy Mohanasundaram
- Arun Venu
- Leila Rato
- Emilia Holm
- Mayank Modi
cFLIP mediated signal transduction in receptor-mediated apoptosis and cell survival
The cellular FLICE-inhibitory protein (cFLIP) is a key regulator of the death receptor apoptotic pathway. The protein has three known isoforms, cFLIP long (cFLIPL), short (cFLIPS) and Raji (cFLIPR). While the short isoform has a distinct role of inhibiting death receptor apoptosis, cFLIPL exhibits a dichotomy in its function between promoting cell death and activating survival pathways. Understanding the mechanisms that serve to determine this crosstalk which results in different cell fates is the central theme of our research.
Regulation of cFLIP by post-translational modifications
The activity of cFLIP, like most cellular proteins, can be regulated at the protein level by post-translational modifications (PTMs). We previously demonstrated that stability of the short isoforms is regulated by classical PKC-mediated phosphorylation on a serine in the prodomain of the protein.
Ongoing research focuses on pinpointing the cFLIPL isoform-specific relevance of two phosphorylation sites we have previously identified. Current findings reiterate the predominantly nuclear localization of cFLIPL and provide evidence of a phosphorylation-dependent subcellular redistribution of the protein in response to signals originating from activated death receptors.
Furthermore, we aim to investigate the relevance of this finding within the context of minimising resistance and improving sensitivity to TRAIL.
cFLIP regulation of cell population size
Our recent findings show that the cFLIP phosphorylation on certain amino acid residues is increased during mitosis, and that the phosphorylated pool of cFLIP proteins is accumulated at the centrosome throughout the cell cycle. In addition, we observe that overexpression of wild type cFLIPL significantly increases the cell population size, whereas overexpression of certain phospho-deficient mutants of cFLIPL causes a decrease in cell population.
Moreover using a pseudo in vivo system (chicken egg model) increased tumour growth is observed in cFLIPL overexpressed cells. These results indicate that by regulating the levels and phosphorylation status of cFLIPL, it is possible to control tumour growth.
We are currently investigating the signalling pathway behind this observation as well as the possible involvement of cell cycle.
Modeling dead and alive: in silico model of cell fate
The research here focuses on iteratively combining mathematical modelling and experiments to understand the role of cFLIP phosphorylation in cell fate decisions.
Our current modelling tasks involve the construction of a single cell and cell population based models to investigate and analyse the dynamics of isoform-specific cFLIP phosphorylation in receptor-mediated cell death signaling and cell cycle progression.
Other related research involves application of systems identification methods for model parameter estimation from flow cytometry data.
The work is done in collaboration with the research groups of Prof Henrik Saxen (Dept of Chemical Engineering ) & Prof H. Toivonen (Information Technology) at ÅÅU.
Responsible Team Members
- Alia Joko
- Erik Nimela
- Preethy Paul
- Michael Silva
Natural compounds and their derivatives as anti-HPV agents
Human papilloma virus (HPV) is the leading cause of cervical cancer, other epithelial cancers, including oral cancer, various head and neck cancers and other diseases like common and genital warts. Cervical cancer causes ~260,000 deaths annually, and every year ~500,000 new diagnoses are being made. Head and neck cancers (HNSCC) cause more than ~750,000 deaths annually, and about 1 million new cases are detected every year. There are currently no specific treatments for patients with HPV-driven cancers. We have discovered few plant-derived compounds and synthetic small molecules that specifically target E6 & E7 oncoproteins, the root cause of HPV-associated diseases. We have had a project for the development and commercialization of this chemo-therapeutic strategy of small molecule derivatives by the Finnish Funding Agency for Innovation, Business Finland and Novo Nordisk Foundation. We have also developed a largescale production procedure of the active natural compounds for the treatment of HPV-mediated warts. In addition, the group is also developing a clear and accelerated pathway for the development of these natural active compounds individually or as combination treatment for warts, in order to create and execute a plan to commercialize this technology in Finland.
- Senthilkumar Rajendran
- Preethy Paul
- Michael Santos Silva
Sensitization of prostate cancer cells to TRAIL-induced cell death
Prostate cancer cells frequently develop resistance toward androgen-deprivation and chemotherapy. Prostate cancer is the most common malignancy and the second leading cause of cancer mortality in men. Prostate tumors express frequently TRAIL receptors on the cell surface, but these receptors are often not able to trigger TRAIL-induced apoptosis because of increased pro-survival signaling. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been shown to be selectively pro-apoptotic in cancer cells, with minimal toxicity to normal tissues. Although this feature makes TRAIL a promising anticancer agent, not all cancer cell types are sensitive to TRAIL-induced apoptosis despite abundant expression of TRAIL receptors. Thus, combinatorial treatments to sensitize tumor cells to TRAIL-induced apoptosis have been in the focus of extensive research. Dietary lignans have shown cancer preventive and antitumorigenic activity, but the mechanisms behind these effects are poorly known. We have identified few plant lignans that can sensitize prostate cancer cells to TRAIL-mediated apoptosis. We have also to certain extent characterized the key structural features for their function and defined their mechanism of action.
- Preethy Paul
Nanoparticles as carriers of antitumor drugs
To be able to target cells of interest is one of the key aims in medicine today, because it could minimize side effects of drugs. This is of special importance in cancer therapies, were the treatment has serious side-effects on healthy cells. Nanomedicine seeks to overcome this limitation by utilizing nanoparticles as targeted drug carrier systems. Together with Jessica Rosenholm at the Pharmaceutical Sciences Laboratory at Åbo Akademi University and Cecilia Sahlgren at the Cell Fate Laboratory at Turku Centre for Biotechnology, we have shown that specific targeting of nanoparticles to cancer cells is possible and, furthermore, that the nanoparticles can be used as
carriers for antitumor drugs with minimal off targeting effect. The collaboration aims at developing this concept further in order to get better specificity, and also to employ the particles as carriers for
other types of compounds such as anti-inflammatory drugs that could be beneficial e.g. ineurodegenerative diseases.
- Erik Niemelä
Functionnal materials at Biological interfaces
Functional Materials at Biological Interfaces (FunMat-consortium) aims to develop innovative materials and functionalized surfaces for the control and sense of cell functions. By temporal and spatial control of cell function we will be able to design novel biological structures to investigate fundamental aspects and applications of tissue engineering and regeneration therapies, tissue monitoring, biotechnology, biomaterials design, drug screening and development.
- Hend Abdelkader
- Arun Venu
- Emilia Holm
- Mayank Modi