Mapping the Interaction Network of Key Proteins Involved in Histone mRNA Generation: A Hydrogen/Deuterium Exchange Study.

Aleksandra Skrajna, Xiao-Cui Yang, Krzysztof Tarnowski, Kinga Fituch, William F Marzluff, Zbigniew Dominski, Michał Dadlez

Journal of molecular biology 2016 Mar 27

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Histone pre-mRNAs are cleaved at the 3' end by a complex that contains U7 snRNP, the FLICE-associated huge protein (FLASH) and histone pre-mRNA cleavage complex (HCC) consisting of several polyadenylation factors. Within the complex, the N terminus of FLASH interacts with the N terminus of the U7 snRNP protein Lsm11, and together they recruit the HCC. FLASH through its distant C terminus independently interacts with the C-terminal SANT/Myb-like domain of nuclear protein, ataxia-telangiectasia locus (NPAT), a transcriptional co-activator required for expression of histone genes in S phase. To gain structural information on these interactions, we used mass spectrometry to monitor hydrogen/deuterium exchange in various regions of FLASH, Lsm11 and NPAT alone or in the presence of their respective binding partners. Our results indicate that the FLASH-interacting domain in Lsm11 is highly dynamic, while the more downstream region required for recruiting the HCC exchanges deuterium slowly and likely folds into a stable structure. In FLASH, a stable structure is adopted by the domain that interacts with Lsm11 and this domain is further stabilized by binding Lsm11. Notably, both hydrogen/deuterium exchange experiments and in vitro binding assays demonstrate that Lsm11, in addition to interacting with the N-terminal region of FLASH, also contacts the C-terminal SANT/Myb-like domain of FLASH, the same region that binds NPAT. However, while NPAT stabilizes this domain, Lsm11 causes its partial relaxation. These competing reactions may play a role in regulating histone gene expression in vivo. Copyright © 2016 Elsevier Ltd. All rights reserved.

Citation

Aleksandra Skrajna, Xiao-Cui Yang, Krzysztof Tarnowski, Kinga Fituch, William F Marzluff, Zbigniew Dominski, Michał Dadlez. Mapping the Interaction Network of Key Proteins Involved in Histone mRNA Generation: A Hydrogen/Deuterium Exchange Study. Journal of molecular biology. 2016 Mar 27;428(6):1180-96