Early phase of development of the assembly complex during cytomegalovirus infection

Funded by the Croatian Science Foundation (Grant: IP-2014-09-9564)

PI: Professor Pero Lučin, M.D., PhD
Department for Physiology and Immunology

University of Rijeka Faculty of Medicine

 

Research team

Professor Pero Lučin, M.D., PhD • UniRi portfolio • Google Scholar
Associate professor Hana Mahmutefendić, PhD • UniRi portfolio • Google Scholar
Assistant professor Gordana Blagojević Zagorac, M.D., PhD • UniRi portfolio • Google Scholar
Assistant professor Kristina Grabušić, PhD • UniRi portfolio • Google Scholar
PhD students
Ljerka Karleuša • UniRi portfolio • Google Scholar
Valentino Pavišić, M.D. • UniRi portfolio • Google Scholar
Natalia Jug • UniRi portfolio • Google Scholar
Silvija Lukanović Jurić, M.D. • UniRi portfolio • Google Scholar
Marina Marcelić, mag. pharm. inv• UniRi portfolio • Google Scholar
Technical supporti
Jelene Đirlić
Ksenija Tulić
Igor Štimac, mag. san.
 

Research summary

Beta-herpesviruses massively reorganize the intracellular compartments in order to create an environment for virion production, known as assembly complex (AC). This structure develops gradually from the earliest stages of infection and involves stepwise reorganization of membrane organelles. Although the final structure is relatively well characterized during human cytomegalovirus infection, the sequence of events that leads to the development of the AC is poorly understood. The goal of this project is to characterize processes proceeding to the formation of the AC in cells infected with murine CMV (MCMV). MCMV starts with remodeling of the endosomal system very early in the infection, which we characterized as development of the early-phase endosomal retention compartment (EPERC). There is strong indication that the EPERC is predecessor of AC. It is generally accepted that endosomal remodeling is driven by small GTPases from the Rab and Arf family and phosphoinositides (PI).

In the proposed project, we plan to reveal sequence of membranous organelle reorganization events in the early phase of infection that presumably lead to the formation of the AC. With that purpose, we will determine the role of small GTPases of the Rab and Arf family and their effector proteins in membrane organelle perturbation events during the early phase of MCMV infection. Finally, we plan to characterize membrane domains of the EPERC during the entire early phase regarding PI content, which include the study of PI effectors and major enzymes that drive PI conversion and thereby change membrane identity. To achieve these goals, we will use triple immunofluorescence staining of EPERC, genetically engineered fluorescent probes, RNA silencing and pharmacological inhibitors. Outcome of the proposed research will be a reconstruction of the mechanism of development of the EPERC and identification of cellular proteins that are targeted by MCMV in the early phase of infection.

AsCo hypothesis

Working hypothesis – a model of development of the EPERC. MCMV initiates development of the EPERC by (left) intrusion of early endosomal vesicles into juxtanuclear area and displacement of the Golgi towards periphery, and by (right down) inhibition of the recycling route from early endosomes to plasma membrane, trafficking route from early endosome to trans-Golgi and from early endosomes to late endosomes (dashed arrows).

 

Achievements

 

Publikacije

  1. Mahmutefendić H., Blagojević Zagorac G., Grabušić K., Karleuša Lj., Maćešić S., Momburg F., Lučin P. (2017). Late Endosomal Recycling of Open MHC-I Conformers. Journal of Cellular Physiology, 232(4), 872-887. doi: 10.1002/jcp.25495.

  2. Gordana Blagojević Zagorac, Hana Mahmutefendić, Senka Maćešić, Ljerka Karleuša, Pero Lučin (2017). Quantitative analysis of endocytic recycling of membrane proteins by monoclonal antibody-based recycling assays. Journal of Cellular Physiology, 232(3), 463-476. doi: 10.1002/jcp.2550.

  3. Karleuša Lj., Mahmutefendić H., Ilić Tomaš M., Blagojević Zagorac G., Lučin P. (2018). Landmarks of endosomal remodeling in the early phase of cytomegalovirus infection. Virology, 515, 108-122. (DOI: 10.1016/j.virol.2017. 12.001).

  4. Mahmutefendić H., Blagojević Zagorac G., Maćešić S., Lučin P. (2018). Chapter 6: Rapid Endosomal Recycling. Open access peer-reviewed Edited volume: Peripheral Membrane Proteins. Edited by Shihori Tanabe. Published: July 25th 2018 by IntechOpen (DOI: 10.5772/intechopen.71809; ISBN: 978-1-78923-513-5; Print ISBN: 978-1-78923-512-8), pp83-104. DOI: 10.5772/intechopen.75685

  5. Kuharić J., Grabušić K., Sotošek Tokmadžić V., Štifter S., Tulić K., Shevchuk O., Lučin P., Šustić A. (2018). Severe traumatic brain injury induces early changes in the physical properties and protein composition of intracranial extracellular vesicles. Journal of Neurotrauma, (https://doi.org/10.1089/neu.2017.5515).

Prezentacije

  1. Karleuša Lj., Grabušić K., Mahmutefendić H., Blagojević Zagorac G., Lučin P. Endosomal remodeling during early phase of Murine Cytomegalovirus infection (Robinskon College), Biochemical Society/FEBS Focused Meeting, Signalling 2015, Cellular Functions of Phosphoinositides and Inositol Phosphates. Cambridge, 1-4 Sepetember 2015. Abstract book, page 29.
  2. Karleuša, Lj., Grabušić, K.,  Mahmutefendić, H., Blagojević Zagorac, G., Ilić Tomaš, M., Lučin, P. Remodelling of endosomal system during murine cytomegalovirus infection. Annual Meeting of Croatian Phyiological Society with international Participation, September 18th - 20th2015. Abstract book pages 37-38
  3. Blagojević Zagorac G., Mahmutefendić H., Mačešić S., Karleuša Lj., Lučin P.: Chasing recycled MHC class I molecules. 4th European Congress of Immunology - ECI 2015, Vienna, Austria, September 6-9, 2015. Abstract book, p246.
  4. Mahmutefendić H., Blagojević Zagorac G., Grabušić K., Karleuša Lj., Momburg F., Lučin P. Late endosomal trafficking and endosecretory recycling route of open MHC-I conformers. 4th European Congress of Immunology - ECI 2015 - Vienna, Vienna, Austria, September 6-9, 2015. Abstract book, pp246-247.

  5. Lučin P., Mahmutefendić H., Blagojević Zagorac G. Pathways of endosomal recycling of MHC-I proteins. Congress of the Croatian Society for Biochemistry and Molecular Biology. Split, June 1-4, 2016. Book of abstracts, p. 37.

  6. Lukanović Jurić S., Jug N., Pavišić V., Karleuša Lj., Mahmutefendić H., Blagojević Zagorac G., Volarević S., Lučin P. Autophagy in MCMV infected cells. 11. Godišnji simpozij Hrvatskog društva fiziologa s međunarodnim sudjelovanjem „Homeostaza-od stanice do organizma“ Zagreb, 17.-18. rujna 2016.

  7. Jug N., Mahmutefendic H., Blagojević Zagorac G., Karleruša Lj., Lučin P. IFNγ influence the endocytic pathways of surface glycoproteins. The 11th Annual CPS Symposium with International Participation. Zagreb, 17-18. 09. 2016.

  8. Pavišić V., Blagojević Zagorac G., Mahmutefendić H., Karleuša Lj., Jug N., Lukanović Jurić S., Lučin P. Role of the ARF proteins during the MCMV infection. The 11th Annual CPS Symposium with International Participation. Zagreb, 17-18. 09. 2016.

  9. Karleuša Lj., Mahmutefendić H., Blagojević Zagorac G., Lučin P. Disruption of proteasomal function, endosomal acidification and actin integration influence IE1 expression in MCMV infected cells. 2016 Annual Meeting of Croatian Immunological Society, Ogulin 14.-15.10.2016.

  10. Blagojević Zagorac G, Mahmutefendić H, Maćešić S, Karleuša Lj, Lučin P. Chasing Recycled Molecules by Monoclonal Antibody-Based Recycling Assays. 2016 Annual Meeting of Croatian Immunological Society, Ogulin 14.-15.10.2016.

  11. Mahmutefendić H, Karleuša L, Ilić Tomaš M, Blagojević Zagorac G, Jug N, Lukanović Jurić S, Pavišić V, Marcelić M, Lučin P. Murine cytomegalovirus restrains endocytic trafficking in sorting endosomes to initiate remodeling of membranous system for its egress. 4th Congress of Croatian Physiological Society and 2nd Regional Congress of the Physiological Societies, September 21st – 24th 2017, Dubrovnik, Croatia.

  12. Marcelić M, Mahmutefendić H, Jurak Begonja A, Blagojević Zagorac G, Karleuša L, Jug N, Lukanović Jurić S, Pavišić V, Lučin P. PI3P is important for early phase endosomal retention compartment (EPERC) formation in MCMV infected fibroblasts. 4th Congress of Croatian Physiological Society and 2nd Regional Congress of the Physiological Societies, September 21st – 24th 2017, Dubrovnik, Croatia.

  13. Jug N, Mahmutefendic H, Lukanović Jurić S, Karleuša L, Blagojević Zagorac G, Pavišić V, Marcelić M, Lučin P. Cellular serine proteases are important for inhibition of transferrin recycling after MCMV infected cells. 4th Congress of Croatian Physiological Society and 2nd Regional Congress of the Physiological Societies, September 21st – 24th 2017, Dubrovnik, Croatia.

  14. Pavišić V, Blagojević Zagorac G, Mahmutefendić H, Karleuša L, Lučin P. ARF5 protein and the MCMV infection. 4th Congress of Croatian Physiological Society and 2nd Regional Congress of the Physiological Societies, September 21st – 24th 2017, Dubrovnik, Croatia.

  15. Blagojevic Zagorac G., Mahmutefendic H, Pavisic V, Jug N, Karleuša L, Lucin P. Influence of different dynamin inhibitors on transferrin/transferrin receptor internalization and recycling. 4th Congress of Croatian Physiological Society and 2nd Regional Congress of the Physiological Societies, September 21st – 24th 2017, Dubrovnik, Croatia.

  16. Lukanović Jurić S, Jug N, Pavišić V, Karleuša L, Blagojević Zagorac G, Marcelić M, Lučin P, Mahmutefendić H. Contribution of cellular proteases in infection process of murine cytomegalovirus. 4th Congress of Croatian Physiological Society and 2nd Regional Congress of the Physiological Societies, September 21st – 24th 2017, Dubrovnik, Croatia.

  17. Lučin P., Mahmutefendić H., Blagojević Zagorac G. Remodeling of endosomal system by viral infection. 4th Congress of Croatian Physiological Society & 2nd Regional Congress of the Physiological Societies. 21-24 September, 2017, Dubrovnik, Croatia.