Mapping the Interaction Network of Key Proteins Involved in Histone mRNA Generation: A Hydrogen/Deuterium Exchange Study. Issue 6 (27th March 2016)

• Record Type: Journal Article
• Authors: Skrajna, Aleksandra; Yang, Xiao-cui; Tarnowski, Krzysztof; Fituch, Kinga; Marzluff, William F.; Dominski, Zbigniew; Dadlez, Michał
• Is Part Of: Journal of molecular biology. Volume 428:Issue 6(2016:Mar. 27)
• Page Start: 1180
• Page End: 1196
• Publication Date: 2016-03-27
• Subjects: FLASH FLICE-associated huge protein -- NPAT nuclear protein, ataxia-telangiectasia locus -- YARP Yin Yang 1-associated protein-related protein -- H/D exchange hydrogen/deuterium exchange -- MS mass spectrometry; U7 snRNP -- FLASH -- Lsm11 -- NPAT -- Mass spectrometry; Molecular biology -- Periodicals; Biology -- Periodicals; Biochemistry -- Periodicals; Bacteriology -- Periodicals; Molecular Biology -- Periodicals; Biochemistry -- Periodicals; Biologie moléculaire -- Périodiques; Biologie -- Périodiques; Biochimie -- Périodiques; Moleculaire biologie; Biochemistry; Biology; Molecular biology; Periodicals; 572.805
• DOI: 10.1016/j.jmb.2016.01.031
• Languages: English
• ISSNs: 0022-2836

Abstract

Abstract: 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 . Graphical Abstract: Highlights: Proteins involved in processing of histone pre-mRNAs form a complex network of interactions. Several interactions within the complex were characterized by hydrogen/deuterium exchange. Our study reveals an unexpected interaction of Lsm11 with the C-terminal region of FLASH. This interaction may play a role in regulating histone gene expression in vivo.