CPSF30 and Wdr33 directly bind to AAUAAA in mammalian mRNA 3′ processing

Chan, Serena Leong; Huppertz, Ina; Yao, Chengguo; Weng, Lingjie; Moresco, James J.; Yates, John R.; Ule, Jernej; Manley, James L.; Shi, Yongsheng

doi:10.1101/gad.250993.114

Cited by 195 publications

(203 citation statements)

References 48 publications

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“…Instead, CPSF30 and Wdr33 were identified as the proteins involved in AU-hexamer recognition (17,18). These findings are intriguing in light of evidence that the H1N1 human influenza virus protein NS1A targets CPSF30 to obstruct cellular mRNA processing (19)(20)(21), suggesting that the link between NS1A and cellular mRNA processing is RNA recognition by CPSF30.…”

Section: Significancementioning

confidence: 77%

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Cleavage and polyadenylation specificity factor 30: An RNA-binding zinc-finger protein with an unexpected 2Fe–2S cluster

Shimberg

Michalek

Oluyadi

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Cleavage and polyadenylation specificity factor 30 (CPSF30) is a key protein involved in pre-mRNA processing. CPSF30 contains five Cys 3 His domains (annotated as "zinc-finger" domains). Using inductively coupled plasma mass spectrometry, X-ray absorption spectroscopy, and UV-visible spectroscopy, we report that CPSF30 is isolated with iron, in addition to zinc. Iron is present in CPSF30 as a 2Fe-2S cluster and uses one of the Cys 3 His domains; 2Fe-2S clusters with a Cys 3 His ligand set are rare and notably have also been identified in MitoNEET, a protein that was also annotated as a zinc finger. These findings support a role for iron in some zinc-finger proteins. Using electrophoretic mobility shift assays and fluorescence anisotropy, we report that CPSF30 selectively recognizes the AU-rich hexamer (AAUAAA) sequence present in pre-mRNA, providing the first molecular-based evidence to our knowledge for CPSF30/RNA binding. Removal of zinc, or both zinc and iron, abrogates binding, whereas removal of just iron significantly lessens binding. From these data we propose a model for RNA recognition that involves a metaldependent cooperative binding mechanism.zinc | iron-sulfur | RNA | protein | binding

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Section: Significancementioning

confidence: 77%

“…CPSF73 is the endonuclease that cleaves the RNA; the roles of the other CPSF proteins are less clear (12,13). Initially, CPSF160 was identified as the protein within the CPSF complex that recognizes the AU-hexamer (14-16); however, two recent studies using cellbased methods found that CPSF160 does not play this role (17,18).…”

mentioning

confidence: 99%

Cleavage and polyadenylation specificity factor 30: An RNA-binding zinc-finger protein with an unexpected 2Fe–2S cluster

Shimberg

Michalek

Oluyadi

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…The U7snRNP recruits processing factors via an interaction between Lsm11 and FLASH (Burch et al 2011;Sabath et al 2013). The highly conserved AAUAAA characteristic of canonical poly(A) mRNAs binds CPSF30 and WDR33 (Chan et al 2014;Schönemann et al 2014) and CstF64 contacts the less conserved G-or G/U-rich element (Takagaki and Manley 1997). While both types of mRNAs utilize unique proteins, cleavage and polyadenylation specificity factor (CPSF)73, CPSF100, and Symplekin are essential for proper processing of all metazoan mRNAs (Sullivan et al 2009).…”

Section: Introductionmentioning

confidence: 99%

In vivo characterization of the Drosophila mRNA 3′ end processing core cleavage complex

Michalski

Steiniger

2015

RNA

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A core cleavage complex (CCC) consisting of CPSF73, CPSF100, and Symplekin is required for cotranscriptional 3′ end processing of all metazoan pre-mRNAs, yet little is known about the in vivo molecular interactions within this complex. The CCC is a component of two distinct complexes, the cleavage/polyadenylation complex and the complex that processes nonpolyadenylated histone pre-mRNAs. RNAi-depletion of CCC factors in Drosophila culture cells causes reduction of CCC processing activity on histone mRNAs, resulting in read through transcription. In contrast, RNAi-depletion of factors only required for histone mRNA processing allows use of downstream cryptic polyadenylation signals to produce polyadenylated histone mRNAs. We used Dmel-2 tissue culture cells stably expressing tagged CCC components to determine that amino acids 272-1080 of Symplekin and the C-terminal approximately 200 amino acids of both CPSF73 and CPSF100 are required for efficient CCC formation in vivo. Additional experiments reveal that the C-terminal 241 amino acids of CPSF100 are sufficient for histone mRNA processing indicating that the first 524 amino acids of CPSF100 are dispensable for both CCC formation and histone mRNA 3 ′ end processing. CCCs containing deletions of Symplekin lacking the first 271 amino acids resulted in dramatic increased use of downstream polyadenylation sites for histone mRNA 3 ′ end processing similar to RNAi-depletion of histone-specific 3 ′ end processing factors FLASH, SLBP, and U7 snRNA. We propose a model in which CCC formation is mediated by CPSF73, CPSF100, and Symplekin C-termini, and the N-terminal region of Symplekin facilitates cotranscriptional 3 ′ end processing of histone mRNAs.

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“…The 3 ′ end processing of most mRNAs relies on the polyadenylation signal (PAS) located 5 ′ to the cleavage site in mRNAs' 3 ′ untranslated regions (UTRs) (Chan et al 2011;Proudfoot 2011). Whereas the CPSF component WDR33 associates via the PAS, FIP1L1 binds flanking U-rich sequences (Kaufmann et al 2004;Chan et al 2014;Schonemann et al 2014).…”

mentioning

confidence: 99%

The Y3** ncRNA promotes the 3′ end processing of histone mRNAs

et al. 2015

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We demonstrate that the Y3/Y3 * * noncoding RNAs (ncRNAs) bind to the CPSF (cleavage and polyadenylation specificity factor) and that Y3 * * associates with the 3 ′ untranslated region (UTR) of histone pre-mRNAs. The depletion of Y3 * * impairs the 3 ′ end processing of histone premRNAs as well as the formation and protein dynamics of histone locus bodies (HLBs), the site of histone mRNA synthesis and processing. HLB morphology is also disturbed by knockdown of the CPSF but not the U7-snRNP components. In conclusion, we propose that the Y3 * * ncRNA promotes the 3 ′ end processing of histone pre-mRNAs by enhancing the recruitment of the CPSF to histone pre-mRNAs at HLBs.

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CPSF30 and Wdr33 directly bind to AAUAAA in mammalian mRNA 3′ processing

Cited by 195 publications

References 48 publications

Cleavage and polyadenylation specificity factor 30: An RNA-binding zinc-finger protein with an unexpected 2Fe–2S cluster

Cleavage and polyadenylation specificity factor 30: An RNA-binding zinc-finger protein with an unexpected 2Fe–2S cluster

In vivo characterization of the Drosophila mRNA 3′ end processing core cleavage complex

The Y3** ncRNA promotes the 3′ end processing of histone mRNAs

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