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Albany 2019: 20th Conversation - Abstracts

category image Albany 2019
Conversation 20
June 11-15 2019
Adenine Press (2019)

Multivalency Regulates Activity in an Intrinsically Disordered Transcription Factor

Transcription factors contain multiple regulatory sites for either post-translational modifications or binding partners, and their activity is thus tuned by the combined action of these components. Recent studies have revealed a high degree of intrinsic disorder in transcription factors, indicating that the inherent dynamical behavior harbored by these structures is critical for these regulatory events to take place. Our developing understanding suggests that these intrinsically disordered domains may provide a multivalent platform for the recruitment of regulatory binding partners. An example of a transcription factor with a long disordered domain is ASCIZ (ATMIN, ZNF822) which also has an unusually high number of multivalent sites for the product of its main target gene, the hub protein LC8 (1). ASCIZ has a folded N-terminal zinc finger domain for DNA binding and an unusually long disordered C-terminal domain, which binds multiple copies of LC8: 11 in human and 7 in Drosophila proteins. Here we integrate multiple approaches including NMR and single particle electron microscopy to elucidate the structure, dynamics, thermodynamics, and hydrodynamics of the large disordered ASCIZ-LC8 complexes, that together reveal a new model by which ASCIZ can maintain stable pools of the hub protein LC8. We tested the main features of this model in cells using transcription activity assays which show a trend wherein mutant ASCIZ constructs with lower LC8 occupancy display higher transcriptional activity, while constructs with higher LC8 occupancy have lower activity. We propose that a dynamic ensemble of complexes is important for fine-tuning ASCIZ transcriptional activity, where stable, low occupancy complexes function to maintain a basal buffering transcription rate for LC8. A change in LC8 cellular concentration would shift this dynamic equilibrium to a higher or lower occupancy state without dramatically altering the level of transcription. High LC8 occupancy shuts down transcription while low LC8 occupancy turns on transcription (2). As LC8 is an essential regulator of dozens of cellular processes, an ability to maintain an LC8 ‘buffer’ is likely important for cellular homeostasis. Although many other transcription factors are regulated by multisite phosphorylation or multiple binding events to different proteins we find no examples of activity tuned by multivalent binding to the gene product in a negative autoregulatory role, which underscores the novelty and potential impact of this study.

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A collage of NMR spectra, electron micrograph and LC8/ASCIZ structure showing three LC8 dimers and two chains of ASCIZ, while the rest of the complex remains disordered. Obtaining data of sufficient quality on a heterogeneous system of multiple conformations of IDP complexes was made possible by creative use of interdisciplinary approaches.

This research is supported by NSF MCB- 1617019 and NIH GM 084276 and HEI 1S10OD018518.

References

    1. Barbar, E. (2008) Dynein Light Chain LC8 Is a Dimerization Hub Essential in Diverse Protein Networks. Biochemistry 47, 503-508.

    2. Clark, S.A., Myers, J.B., King, A., Fiala, R., Novacek, J., Pearce, G., Heierhorst, J., Reichow, S.L., and Barbar, E. (2018) Multivalency Regulates Activity in an Intrinsically Disordered Transcription Factor. ELife, 1;7. pii: e36258.

Sarah Clark
Kayla Jara
Patrick Reardon
Elisar Barbar

Department of Biochemistry and Biophysics
Oregon State University
Corvallis, OR 97331

Ph: (541) 737-4143
Email: barbare@oregonstate.edu