Periodic Reporting for period 2 - UBIMOTIF (Short linear interaction motifs as specificity determinants in the ubiquitin system – discovery, mechanisms and therapeutic opportunities)
Período documentado: 2021-12-01 hasta 2024-05-31
The proliferation of cells and the regulation of cellular signaling relies on the proper control of protein function. A major regulatory mechanism of protein function is the ubiquitin system that can add or remove a small ubiquitin molecule to specific proteins. A major obstacle in understanding the ubiquitin system is how specific proteins are recognized by components of the system. An emerging theme is that short linear motifs (SLiMs) which are present in proteins are recognized by the ubiquitin system. However, identification of SLiMs and how they confer specificity is largely unexplored. Uncovering this will provide fundamental biological understanding of cell regulation and its deregulation in human malignancies. Furthermore, SLiMs are a powerful entry point for developing therapeutic agents that are able to either inhibit or hijack the ubiquitin system.
The UBIMOTIF network brought together researchers in a European network focusing on SLiMs in the ubiquitin system. A unique aspect of UBIMOTIF was that it merged researchers with expertise in SLiMs, the ubiquitin system and therapeutic approaches to allow breakthrough discoveries in the ubiquitin system. This diversity in experimental and knowledge background provided an ideal training setup for the early-stage researchers as they got exposed to different disciplines.
During the lifetime of UBIMOTIF the network successfully identified a large number of SLiMs experimentally and characterized these in cellular systems and using computational methods. Computational pipelines were developed that provide important tools for studying SLiMs in the future in different biological pathways. A number of ubiquitin ligases where also targeted in therapeutic strategies which allowed the identification of small molecule fragments and PROTAC scaffolds for further developments as therapeutic agents.
In the network there was a strong emphasis in training the ESRs with a focus on both scientific skills and transferable skill training. Furthermore, a strong network identity was established among the ESRs through the secondments, joint training activities and participation and presentation at annual meetings.
The UBIMOTIF network brought together researchers in a European network focusing on SLiMs in the ubiquitin system. A unique aspect of UBIMOTIF was that it merged researchers with expertise in SLiMs, the ubiquitin system and therapeutic approaches to allow breakthrough discoveries in the ubiquitin system. This diversity in experimental and knowledge background provided an ideal training setup for the early-stage researchers as they got exposed to different disciplines.
During the lifetime of UBIMOTIF the network successfully identified a large number of SLiMs experimentally and characterized these in cellular systems and using computational methods. Computational pipelines were developed that provide important tools for studying SLiMs in the future in different biological pathways. A number of ubiquitin ligases where also targeted in therapeutic strategies which allowed the identification of small molecule fragments and PROTAC scaffolds for further developments as therapeutic agents.
In the network there was a strong emphasis in training the ESRs with a focus on both scientific skills and transferable skill training. Furthermore, a strong network identity was established among the ESRs through the secondments, joint training activities and participation and presentation at annual meetings.
At a scientific level the work was distributed to three scientific work packages each with their unique focus on specific aspects in the ubiquitin system.
Overview of results:
One work package focused on the unique SLiM discovery tool pipeline allowing large scale discovery of motifs and integrating this with bioinformatic approaches as well as mass spectrometry-based approaches. During UBIMOTIF hundreds of protein domains from ubiquitin ligases and DUBs where screened which returned numerous novel motifs. Several computational platforms were developed suited for investigating SLiMs and have been made available to the research community. For a number of novel SLiMs the network analyzed these in a cellular context to elucidate their function in depth using dedicated cellular model systems.
To harness the new knowledge of SLiMs for therapeutic approaches several projects focused on small molecule development. Through evaluation of expression data specific members of the Cul3 ubiquitin ligases were identified as relevant targets for small molecule discovery and PROTAC development. Expression protocols for targets were developed and subsequently screened in fragment-based screening assays leading to hits that were further validated. Novel tool compounds for a Cul3 member have also been developed and characterized which constitutes an important step towards developing novel PROTACs.
A large effort of the network focused on understanding the cellular functions of SLiMs in the ubiquitin system. The focus has been on specific ubiquitin ligases and chain types. Novel insights into substrate recognition by Cul3 complexes and the hGID complex has been obtained hereby providing insight into how they regulate cellular processes. A unique system for ubiquitin replacement has also been established to understand how different ubiquitin chain types contribute to specificity in the system.
Exploitation of results:
The ubiquitin system is viewed as a highly relevant biological system for drug targeting. Particular the emergence of PROTACs, also known as molecular glues, constitute an important area of research with important clinical implications. An important unique element of the UBIMOTIF network was that SLiM binding pockets could constitute ideal targets for small molecule development that could be further developed into PROTACs. The establishment of expression and purification protocols for a large number of ubiquitin ligases and DUBs and the identification of SLiMs binding to these domains constitute an important resource for the scientific community that will allow exploitation of the ubiquitin system in the future for medical applications. The UBIMOTIF has already exploited this by developing small molecule compounds that bind to Cul3 family members, important ubiquitin ligases in the cell, that constitute an important first step in developing PROTACs.
Dissemination of results:
One of the main channels for dissemination has been through presentation at scientific meetings and through departmental seminars. The ESRs and their PIs have participated in more than 100 relevant meetings and presented their work either in the form of posters or through oral presentations with the target audience being the scientific community. All ESRs have also presented their work at departmental seminars at their institutes and at UBIMOTIF annual meetings. At annual meetings we have invited relevant researchers in the field so they became aware of the work in UBIMOTIF. The work on curating motifs as part of ESR journal clubs is integrated into the ProViz webpage (http://slim.icr.ac.uk/proviz/(se abrirá en una nueva ventana)) making this available to researchers across the world. As specific projects have finalized, they have been published in relevant journals making the results available to the scientific community and public. Computational tools such as FaSTPACE and xProtCAS that have been developed by UBIMOTIF are freely available to download.
Overview of results:
One work package focused on the unique SLiM discovery tool pipeline allowing large scale discovery of motifs and integrating this with bioinformatic approaches as well as mass spectrometry-based approaches. During UBIMOTIF hundreds of protein domains from ubiquitin ligases and DUBs where screened which returned numerous novel motifs. Several computational platforms were developed suited for investigating SLiMs and have been made available to the research community. For a number of novel SLiMs the network analyzed these in a cellular context to elucidate their function in depth using dedicated cellular model systems.
To harness the new knowledge of SLiMs for therapeutic approaches several projects focused on small molecule development. Through evaluation of expression data specific members of the Cul3 ubiquitin ligases were identified as relevant targets for small molecule discovery and PROTAC development. Expression protocols for targets were developed and subsequently screened in fragment-based screening assays leading to hits that were further validated. Novel tool compounds for a Cul3 member have also been developed and characterized which constitutes an important step towards developing novel PROTACs.
A large effort of the network focused on understanding the cellular functions of SLiMs in the ubiquitin system. The focus has been on specific ubiquitin ligases and chain types. Novel insights into substrate recognition by Cul3 complexes and the hGID complex has been obtained hereby providing insight into how they regulate cellular processes. A unique system for ubiquitin replacement has also been established to understand how different ubiquitin chain types contribute to specificity in the system.
Exploitation of results:
The ubiquitin system is viewed as a highly relevant biological system for drug targeting. Particular the emergence of PROTACs, also known as molecular glues, constitute an important area of research with important clinical implications. An important unique element of the UBIMOTIF network was that SLiM binding pockets could constitute ideal targets for small molecule development that could be further developed into PROTACs. The establishment of expression and purification protocols for a large number of ubiquitin ligases and DUBs and the identification of SLiMs binding to these domains constitute an important resource for the scientific community that will allow exploitation of the ubiquitin system in the future for medical applications. The UBIMOTIF has already exploited this by developing small molecule compounds that bind to Cul3 family members, important ubiquitin ligases in the cell, that constitute an important first step in developing PROTACs.
Dissemination of results:
One of the main channels for dissemination has been through presentation at scientific meetings and through departmental seminars. The ESRs and their PIs have participated in more than 100 relevant meetings and presented their work either in the form of posters or through oral presentations with the target audience being the scientific community. All ESRs have also presented their work at departmental seminars at their institutes and at UBIMOTIF annual meetings. At annual meetings we have invited relevant researchers in the field so they became aware of the work in UBIMOTIF. The work on curating motifs as part of ESR journal clubs is integrated into the ProViz webpage (http://slim.icr.ac.uk/proviz/(se abrirá en una nueva ventana)) making this available to researchers across the world. As specific projects have finalized, they have been published in relevant journals making the results available to the scientific community and public. Computational tools such as FaSTPACE and xProtCAS that have been developed by UBIMOTIF are freely available to download.
UBIMOTIF has used its unique screening approach to uncover new SLiMs in the ubiquitin system to move our understanding of specificity determinants beyond state of the art. Furthermore, the results will also have a wider impact for the research communities trying to understand how the intrinsically disordered regions of proteins control cellular functions. Given the large number of novel motifs discovered this facilitates the extraction of more general principles. The efforts of the network in developing small molecules targeting the ubiquitin system is expected to have a wider societal impact as it can lead to new treatments for human diseases which can support the growth of the European economy. By uniting researchers in the field of drug discovery with researchers focusing on SLiMs we created a unique constellation that paved the way for rethinking targeting of SLiMs in the ubiquitin system. Finally, the training provided by UBIMOTIF has resulted in highly skilled researchers that will support the knowledge based European economy. The combination of excellent scientific training combined with numerable transferable skills training makes the fellows highly attractive for the European job market where they can support several functions.