Unraveling client selectivity and pattern recognition of DNAJ molecular chaperones inhibiting intracellular polyglutamine aggregation in neurodegenerative models

• Applicant: Dr. Gaetano Calabrese
• Subject Area: Biochemistry ; Cell Biology
• Term: from 2020 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 446508933

• Final Report Year: 2024

Final Report Abstract

This research represents a significant step forward in understanding the roles of DNAJB6 and DNAJB8 co-chaperones in protein folding, identifying the dient proteins that depend an these co-chaperones and uncovering specific motifs or regions within clients by employing cutting-edge mass spectrometry techniques. Notably, despite their similar domain organization, DNAJB6 and DNAJB8 possess interactions with distinct dient proteins, displaying minimal overlap in their dient repertoires. This investigation demonstrates that the dient repertoires of DNAJB6 and DNAJB8 exhibit distinctive characteristics and specificities, enhancing our understanding of their roles in cellular proteostasis. The research also unveils intriguing insights, indicating that DNAJB6 potentially plays a crucial role in the regulation of proteins within cellular condensates, while highlighting the significance of DNAJB8 in the formation of mitochondria sheaths during late spermatogenesis. lt reveals that DNAJB6 engages with a diverse array of potential dient proteins, predominantly located in the nucleus, suggesting its involvement in maintaining the solubility of nuclear proteins undergoing phase separation. Moreover, the research points to unexplored avenues, such as the tissue-specificity of DNAJB6 interactomes and the identification of dient binding motifs, which offer promising prospects for future investigations in this field. This study also advances scientific knowledge and practical applications through the introduction of an innovative method that combines pulse-SILAC and BiolD techniques, allowing for the precise differentiation of newly synthesized and long-lived dient proteins, thereby refining the categorization of chaperone-client interactions. An unexpected result emerges when this approach is applied to the study of DNAJA2, uncovering a remarkable difference between newly synthesized and long-lived dient proteins, with the former displaying an elevated ß-sheet content and an increased propension to aggregation in the absence of DNAJA2, offering valuable insights into the realms of protein folding and stability.

Publications

• Protein interaction networks in neurodegenerative diseases: From physiological function to aggregation. Journal of Biological Chemistry, 298(7), 102062.
  Calabrese, Gaetano; Molzahn, Cristen & Mayor, Thibault
  
• Pulse labeling reveals the tail end of protein folding by proteome profiling. Cell Reports, 40(3), 111096.
  Zhu, Mang; Kuechler, Erich R.; Wong, Ryan W.K.; Calabrese, Gaetano; Sitarik, Ian M.; Rana, Viraj; Stoynov, Nikolay; O.’Brien, Edward P.; Gsponer, Jörg & Mayor, Thibault
  
• Identification of newly translated thermo-sensitive proteins using pulse SILAC mass spectrometry and the GAL promoter system. STAR Protocols, 4(1), 102059.
  Zhu, Mang; Calabrese, Gaetano; Wong, Ryan W.K. & Mayor, Thibault
  
• Real-Time Monitoring of Hydrogen Peroxide Levels in Yeast and Mammalian Cells. Methods in Molecular Biology, 149-165. Springer US.
  Calabrese, Gaetano; Jacobs, Lianne J. H. C. & Riemer, Jan