Fip1 is a multivalent interaction scaffold for processing factors in human mRNA 3' end biogenesis

Abstract
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Abstract

3' end formation of most eukaryotic mRNAs is dependent on the assembly of a ~1.5 megadalton multiprotein complex, that catalyzes the coupled reaction of pre-mRNA cleavage and polyadenylation. In mammals, the cleavage and polyadenylation specificity factor (CPSF) constitutes the core of the 3' end processing machinery onto which the remaining factors, including cleavage stimulation factor (CstF) and poly(A) polymerase (PAP), assemble. These interactions are mediated by Fip1, a CPSF subunit characterized by high degree of intrinsic disorder. Here, we report two crystal structures revealing the interactions of human Fip1 (hFip1) with CPSF30 and CstF77. We demonstrate that CPSF contains two copies of hFip1, each binding to the zinc finger (ZF) domains 4 and 5 of CPSF30. Using polyadenylation assays we show that the two hFip1 copies are functionally redundant in recruiting one copy of PAP, thereby increasing the processivity of RNA polyadenylation. We further show that the interaction between hFip1 and CstF77 is mediated via a short motif in the N-terminal 'acidic' region of hFip1. In turn, CstF77 competitively inhibits CPSF-dependent PAP recruitment and 3' polyadenylation. Taken together, these results provide a structural basis for the multivalent scaffolding and regulatory functions of hFip1 in 3' end processing.

Data availability

X-ray diffraction data (atomic coordinates and structure factors) have been submitted to the PDB and will be released upon publication.

The following data sets were generated

1. Lena Muckenfuss
2. Martin Jinek
(2022) Crystal structure of human CPSF30 in complex with hFip1
Protein Data Bank ID: 7ZYH.

https://dx.doi.org/10.2210/pdb7zyh/pdb
1. Lena Muckenfuss
2. Martin Jinek
(2022) Crystal structure of human CstF77 in complex with hFip1
Protein Data Bank ID: 7ZY4.

https://dx.doi.org/10.2210/pdb7zy4/pdb

The following previously published data sets were used

1. Sun
2. Y.
3. Zhang
4. Y.
5. Walz
6. T.
7. Tong
8. L.
(2019) Cryo-EM structure of human CPSF160-WDR33-CPSF30-PAS RNA-CstF77 complex
Potein Data Bank ID: 6URO.

https://www.rcsb.org/structure/6URO
1. Sun
2. Y.
3. Zhang
4. Y.
5. Walz
6. T.
7. Tong
8. L.
(2019) Cryo-EM structure of human CPSF160-WDR33-CPSF30-PAS RNA-CstF77 complex
Electron Microscopy Data Bank ID: EMD-20861.

https://www.ebi.ac.uk/emdb/EMD-20861

Article and author information

Author details

Lena Maria Muckenfuss

Department of Biochemistry, University of Zurich, Zurich, Switzerland

Competing interests
The authors declare that no competing interests exist.
Anabel Carmen Migenda Herranz

Department of Biochemistry, University of Zurich, Zurich, Switzerland

Competing interests
The authors declare that no competing interests exist.
Franziska M Boneberg

Department of Biochemistry, University of Zurich, Zurich, Switzerland

Competing interests
The authors declare that no competing interests exist.
Marcello Clerici

Department of Biochemistry, University of Zurich, Zurich, Switzerland

Competing interests
The authors declare that no competing interests exist.
Martin Jinek

Department of Biochemistry, University of Zurich, Zurich, Switzerland

For correspondence
jinek@bioc.uzh.ch

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-7601-210X

Funding

Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (NCCR RNA and Disease)

Martin Jinek

Boehringer Ingelheim Fonds

Lena Maria Muckenfuss

Howard Hughes Medical Institute (55008735)

Martin Jinek

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.