Reference number : GC-2017-Q1-02988, amendment number : , agreement type : G, recipient legal name : Saridakis, Vivian (York University)|Saridakis, Vivian (Université York), recipient country : CA, recipient province : nan, recipient city : Ontario, recipient postal code : nan, program name english : Discovery Grants Program - Individual, program name french : Programme de subventions à la découverte - individuelles, program purpose english : Ubiquitin signalling is critical in many cellular processes. The covalent attachment of ubiquitin to substrate proteins affects their biological functions. The best characterized function is 26S mediated proteasomal degradation of the substrate protein. Ubiquitin is attached to substrate proteins through an enzymatic cascade consisting of E1, E2 and E3 enzymes. Ubiquitin can also be cleaved from substrate proteins by deubiquitinases (DUBs). Deubiquitination activity must be highly regulated otherwise proteins would be targeted by these enzymes non-specifically resulting in deregulated cellular processes. Pathogenic organisms have the ability to manipulate ubiquitin signalling in host organisms to their advantage. This is a critical step in invasion by the pathogen. A variety of pathogenic organisms including bacteria and viruses possess proteins involved in ubiquitin signalling including bacterial/viral E3 ligases and DUBs that are used to dampen the immune response to enable infection. 
The long-term goals of my lab are to investigate the function of bacterial, viral and human deubiquitinases to provide in-depth understanding of their roles in cellular processes. The objective of this research program is to characterize a bacterial and several viral DUBs by elucidating their function, substrates and mechanism of action in host organisms. We are now expanding our research theme to include DUBs from various pathogenic organisms. The project is divided into several short term objectives: 
 
[1] Structural and Functional Characterization of the E. coli deubiquitinase ElaD. Very little is known about E. coli ElaD. We will identify host substrates and characterize the structural basis of interaction between ElaD and its host substrates. 
[2] Characterization of viral deubiquitinases BPLF1, ORF64, UL36 and UL48. BPLF1, ORF64, UL36 and UL48 are deubiquitinases from different herpesviruses. There is very little known about the function of these proteins. For instance, few host substrate proteins have been identified for three of the four viral DUBs. BPLF1 interacts with and deubiquitinates PCNA, TRAF6 and NEMO, UL36 deubiquitinates TRAF3 and ORF64 deubiquitinates RIG-1. 
[3] Identification of BPLF1, ORF64, UL36 and UL48 host substrate(s) and interacting proteins. There are very few host substrates known for viral DUBS. In this part of the proposal we will identify novel host substrates using affinity purification-mass spectrometry. We will follow up by validating the identified viral DUB substrates in human cells with a variety of experiments. 
 Significance: Pathogenic organisms utilize a variety of methods to overcome the immune response during infection. The proposed studies will enhance our knowledge of the role of pathogenic deubiquitinases during infection by identifying substrates and pathways that are manipulated by the bacteria or viruses., program purpose french : Ubiquitin signalling is critical in many cellular processes. The covalent attachment of ubiquitin to substrate proteins affects their biological functions. The best characterized function is 26S mediated proteasomal degradation of the substrate protein. Ubiquitin is attached to substrate proteins through an enzymatic cascade consisting of E1, E2 and E3 enzymes. Ubiquitin can also be cleaved from substrate proteins by deubiquitinases (DUBs). Deubiquitination activity must be highly regulated otherwise proteins would be targeted by these enzymes non-specifically resulting in deregulated cellular processes. Pathogenic organisms have the ability to manipulate ubiquitin signalling in host organisms to their advantage. This is a critical step in invasion by the pathogen. A variety of pathogenic organisms including bacteria and viruses possess proteins involved in ubiquitin signalling including bacterial/viral E3 ligases and DUBs that are used to dampen the immune response to enable infection. 
The long-term goals of my lab are to investigate the function of bacterial, viral and human deubiquitinases to provide in-depth understanding of their roles in cellular processes. The objective of this research program is to characterize a bacterial and several viral DUBs by elucidating their function, substrates and mechanism of action in host organisms. We are now expanding our research theme to include DUBs from various pathogenic organisms. The project is divided into several short term objectives: 
 
[1] Structural and Functional Characterization of the E. coli deubiquitinase ElaD. Very little is known about E. coli ElaD. We will identify host substrates and characterize the structural basis of interaction between ElaD and its host substrates. 
[2] Characterization of viral deubiquitinases BPLF1, ORF64, UL36 and UL48. BPLF1, ORF64, UL36 and UL48 are deubiquitinases from different herpesviruses. There is very little known about the function of these proteins. For instance, few host substrate proteins have been identified for three of the four viral DUBs. BPLF1 interacts with and deubiquitinates PCNA, TRAF6 and NEMO, UL36 deubiquitinates TRAF3 and ORF64 deubiquitinates RIG-1. 
[3] Identification of BPLF1, ORF64, UL36 and UL48 host substrate(s) and interacting proteins. There are very few host substrates known for viral DUBS. In this part of the proposal we will identify novel host substrates using affinity purification-mass spectrometry. We will follow up by validating the identified viral DUB substrates in human cells with a variety of experiments. 
 Significance: Pathogenic organisms utilize a variety of methods to overcome the immune response during infection. The proposed studies will enhance our knowledge of the role of pathogenic deubiquitinases during infection by identifying substrates and pathways that are manipulated by the bacteria or viruses., agreement title english : Structural and Functional Analysis of Bacterial and Viral Deubiquitinases, agreement title french : Structural and Functional Analysis of Bacterial and Viral Deubiquitinases, agreement number : RGPIN, agreement value : 130000.0, agreement start date : 2017-05-10, agreement end date : nan, description english : nan, description french : nan, expected results english : nan, expected results french : nan, additional information english : Grant or Award spanning more than one fiscal year. (2017-2018 to 2022-2023), additional information french : Subvention ou bourse octroyée s'appliquant à plus d'un exercice financier. (2017-2018 à 2022-2023), owner organization : nserc-crsng, owner organization title : Natural Sciences and Engineering Research Council of Canada | Conseil de recherches en sciences naturelles et en génie du Canada