Our AIM– the basic ecotoxicological research towards understanding of:

● Molecular base and role of critical cellular defense and/or detoxification mechanisms in aquatic organisms;

● Interactions of cellular defense mechanisms with both, classical and emerging environmental contaminants.

The final goal of the basic investigations described above is the evaluation and positioning of the cellular defense mechanisms as important determinants of the ADME Tox characteristics of environmental contaminants. Finally, results of our studies contribute to the improvement and/or development of the early warning molecular biomarkers and high-throughput screening tools for evaluation of both single environmental contaminants and complex environmental samples.

Main research directions

1.Understanding of the ecotoxicological significance of:

a) Efflux transporters: ABC (ATP binding cassette) and MATE (Multidrug and Toxin extrusion) transport proteins

b) Uptake SLC (Solute carriers) transport proteins: SLC21/OATP and SLC21 protein families

c) GST (Glutathione S-transferase) enzymes: phase 2 of cellular ADME (Administration, Distribution, Metabolism, Elimination)

2.Development of the Effects-Directed Analyses (EDA) approach for identification of hazardous chemical contaminants.

2017- present – Group leader, Tenure-track Research Associate

Grant holder

 1. 2020-2023 ‘Structural characterization of factors involved in DNA-protein crosslink repair‘ - Slovenian-Croatian Bilateral Project (IPS-2020-01) (1.5 million HRK), Institute Ruder Boskovic, Croatia and National Chemistry Institute, Slovenia (300,000 EUR), total project value 500,000 EUR) . Collaborators: dr.sc Marjetka Podobnik (SLO) and dr.sc. Nives Ivić (IRB, HR)

 2. 2018-2023 ‘Deciphering DNA-Protein Crosslink Repair in vivo using CRISPR/Cas9 genome editing in zebrafish model ’Croatian National Science Foundation Installation project grant (1.86 million HRK), Institute Ruder Boskovic, Croatia.

Popovic lab webpage available at:

https://martafry.wixsite.com/popoviclab

Members:

Group leader: Marta Popovic, PhD

Research Associate: Cecile Otten, PhD

PhD student: Christine Supina, MSc molecular biology

PhD student: Ivan Anticevic, MSc biology and chemistry

PhD student: Marin Kutnjak, MSc molecular biology

Associate member: Research Associate Nives Ivić, PhD

Project title: Structural characterization of factors involved in DNA-protein crosslink repair (2020-2023)

DNA-protein crosslinks (DPCs) are severe DNA lesions which occur when a protein becomes irreversibly covalently linked to DNA. They have adverse effects on the organismal level including cancer, premature aging and neurodegenerative diseases. Due to their bulky nature, DPCs impair all DNA transactions (replication, transcription and repair) and DPC repair is therefore an essential cellular pathway. Despite the fact that DPCs are frequently occurring in the nucleus and cause severe damage on a cellular level, not much is known about their repair mechanisms at a molecular level. Our structural studies aim to solve near-atomic details of the SPRTN-dependent DPC repair complex, the SPRTN:p97 complex and a novel DPC factor, ACRC, for the first time. Obtained knowledge will be fundamentally important for a deeper understanding of the repair pathway and thus for the developing research on p97 and SPRTN inhibitors for the purpose of targeted clinical therapies in cancerogenesis and aging. In terms of methodology, proposed studies will offer novel approaches for investigating DNA repair complexes and disordered protein regions using cryo-EM. Cryo-EM (Cryogenic electron microscopy) is a powerful approach to solve structures of large proteins and protein complexes. The importance of the approach was further emphasized in 2017., when the Nobel Prize in Chemistry was awarded to J. Dubochet, J. Frank, R. Henderson and "for developing cryo-electron microscopy for the high-resolution structure determination of biomolecules in solution." (https://www.nature.com/news/cryo-electron-microscopy-wins-chemistry-nobel-1.22738;https://www.chemistryworld.com/news/explainer-what-is-cryo-electron-microscopy/3008091.article#/)

Project title: Deciphering DNA-Protein Crosslink Repair in vivo using zebrafish model (2018-2023)

DNA-protein crosslink (DPC) is a type of DNA lesion where a protein becomes irreversibly covalently bound to DNA upon exposure to endogenous or exogenous crosslink inducers. Endogenous DPC inducers are products of normal cellular metabolism such as reactive oxygen species, aldehydes and DNA helical alterations, while exogenous inducers include UV light, ionizing radiation and various chemicals. DNA-protein crosslinks are common DNA lesions which present a physical blockage to all DNA transactions: replication, transcription, recombination and repair. If not repaired, DPCs cause genomic instability and adverse phenotypes in humans including premature aging, neurodegeneration and cancer. Despite the frequency and severe outcomes of DPCs, DNA-protein crosslink repair (DPCR) has been sparsely studied, mostly because it has not been considered a separate DNA damage repair pathway until recently. In 2014 and 2016, several groups have identified novel proteases, Wss1 and SPRTN, which initiate the removal of DPCs through the proteolytic digestion of crosslinked proteins. The discovery of proteolysis-coupled DPC repair lead to recognition of the DNA-protein crosslink repair as a separate DNA damage repair pathway. However, we currently do not know how is the pathway orchestrated and which other factors are involved, while almost nothing is known of DPCR mechanism in vivo. Therefore, within this project we aim to unravel the orchestration of the DPCR pathway in vivo using zebrafish (Danio rerio) as a well-characterized vertebrate model. We will use CRISPR/Cas9 gene manipulation tools to knock-out or mutate specific genes in zebrafish which we suspect are involved in the removal of DNA-protein crosslinks. Contribution of each protein (and their combinations) to the DNA-protein crosslink repair will be quantified after DPC isolation from transgenic zebrafish embryos and adults. We will also generate a GFP reporter assay in cell lines and transgenic fish which will enable the quantification of DPCR efficiency in vitro and in vivo.

More information about the project: https://www.irb.hr/eng/Research/Divisions/Division-for-Marine-and-Environmental-Research/Laboratory-for-molecular-ecotoxicology/DNAPRO-Deciphering-DNA-Protein-Crosslink-Repair-in-vivo-using-zebrafish-model