A new role for FBP21 as regulator of Brr2 helicase activity

Henning, Lars; Santos, K.F.; Sticht, Jana; Jehle, Stefanie; Lee, Chung-Tien; Wittwer, Malte; Urlaub, Henning; Stelzl, Ulrich; Wahl, Markus C.; Freund, Christian

doi:10.1093/nar/gkx535

Cited by 28 publications

(65 citation statements)

References 61 publications

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“…2D), and via its interaction with BRR2 NC , it may potentially modulate BRR2 activity. Intriguingly, an α-helical element of FBP21, which was implicated in regulating BRR2 activity (Henning et al 2017), is positioned between BRR2 NC and U4/U6 proteins that are bound to U4/ U6 stem I (Fig. 2D).…”

Section: Structure Of the Precatalytic B Complexmentioning

confidence: 99%

Structural Insights into Nuclear pre-mRNA Splicing in Higher Eukaryotes

Kastner

Will

Stark

et al. 2019

Cold Spring Harb Perspect Biol

179

164

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The spliceosome is a highly complex, dynamic ribonucleoprotein molecular machine that undergoes numerous structural and compositional rearrangements that lead to the formation of its active site. Recent advances in cyroelectron microscopy (cryo-EM) have provided a plethora of near-atomic structural information about the inner workings of the spliceosome. Aided by previous biochemical, structural, and functional studies, cryo-EM has confirmed or provided a structural basis for most of the prevailing models of spliceosome function, but at the same time allowed novel insights into splicing catalysis and the intriguing dynamics of the spliceosome. The mechanism of pre-mRNA splicing is highly conserved between humans and yeast, but the compositional dynamics and ribonucleoprotein (RNP) remodeling of the human spliceosome are more complex. Here, we summarize recent advances in our understanding of the molecular architecture of the human spliceosome, highlighting differences between the human and yeast splicing machineries. 7 Insights into the function of PPIases and IBC proteins during splicing 8 Analysis of the conformational dynamics of hB act 9 Structure of human C and C * complexes 10 Insights into the mechanism of action of spliceosomal helicases 11 Large-scale movements of spliceosomal components 12 Future directions References

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Section: Structure Of the Precatalytic B Complexmentioning

confidence: 99%

Structural Insights into Nuclear pre-mRNA Splicing in Higher Eukaryotes

Kastner

Will

Stark

et al. 2019

Cold Spring Harb Perspect Biol

179

164

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“…Recently, the human spliceosomal protein FBP21 was found to be a novel Brr2 regulator that also exerts its effects by interacting via an IDR at the Brr2 C-terminal Sec63 unit (54). Notably, a short, positively charged peptide in the C-terminal region of FBP21 (sequence FKKRR) strongly contributed to Brr2 binding but was insufficient to influence Brr2 helicase activity (54). Similar positively charged sequences are contained in the Brr2-binding regions 1 (3-IKKRNKIR-10) and 2 (53-IKFKKVPKR-61) of Ntr2, consistent with the idea that they also comprise anchoring points on Brr2 C-Sec63 .…”

Section: Brr2 Regulation From a Distancementioning

confidence: 99%

Intrinsically Disordered Protein Ntr2 Modulates the Spliceosomal RNA Helicase Brr2

Wollenhaupt

Henning

Sticht

et al. 2018

Biophysical Journal

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Precursor messenger RNA splicing is mediated by the spliceosome, a large and dynamic molecular machine composed of five small nuclear RNAs and numerous proteins. Many spliceosomal proteins are predicted to be intrinsically disordered or contain large disordered regions, but experimental validation of these predictions is scarce, and the precise functions of these proteins are often unclear. Here, we show via circular dichroism spectroscopy, dynamic light scattering, and NMR spectroscopy that the yeast spliceosomal disassembly factor Ntr2 is largely intrinsically disordered. Peptide SPOT analyses, analytical size-exclusion chromatography, and surface plasmon resonance measurements revealed that Ntr2 uses an N-terminal region to bind the C-terminal helicase unit of the Brr2 RNA helicase, an enzyme involved in spliceosome activation and implicated in splicing catalysis and spliceosome disassembly. NMR analyses suggested that Ntr2 does not adopt a tertiary structure and likely remains disordered upon complex formation. RNA binding and unwinding studies showed that Ntr2 downregulates Brr2 helicase activity in vitro by modulating the fraction of helicase molecules productively bound to the RNA substrate. Our data clarify the nature of a physical link between Brr2 and Ntr2, and point to the possibility of a functional Ntr2-Brr2 interplay during splicing.

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“…However, the CC most likely does not simply represent a passive landing pad for other proteins, as some of the interacting proteins also influence the activity of Brr2. For example, the intrinsically disordered proteins Ntr2 (in yeast) and FBP21 (in human) bind to the CC of Brr2 and tune down its unwinding activity in vitro (32,33). In the respective studies, it was speculated that their binding to the CC might influence the communication between the cassettes.…”

Section: Implications For Brr2 Regulation By Protein Partnersmentioning

confidence: 99%

“…In addition, a Cterminal Jab1 domain of the spliceosomal master regulator, Prp8, can either inhibit or activate Brr2, depending on whether or not a C-terminal tail of the domain is inserted into the helicase's RNA-binding tunnel (29)(30)(31). Other proteins binding to Brr2 can also modulate its activities in vitro (32,33).…”

Section: Introductionmentioning

confidence: 99%

“…Only the N-terminal cassette (NC) of Brr2 is an active NTPase/RNA helicase, while the C-terminal cassette (CC) is considered a pseudo-enzyme (13,34). Nevertheless, the CC is essential for yeast viability (35), can still bind but does not hydrolyze ATP (34) and serves as an interaction platform for a number of other splicing factors (36,37), some of which have been shown to modulate Brr2 activity by presently unknown mechanisms (32,33). Indeed, in human (h) Brr2, the CC itself can activate the NC helicase (34).…”

Section: Introductionmentioning

confidence: 99%

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Positive and negative intra-molecular modulation in a dual-cassette RNA helicase

Vester

Kuropka

et al. 2019

Preprint

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RNA helicase Brr2 is required for the activation of the spliceosome prior to the first catalytic step of splicing. Brr2 represents a distinct subgroup of Ski2-like nucleic acid helicases whose members comprise tandem helicase cassettes. Only the Nterminal cassette of Brr2 is an active ATPase and can unwind substrate RNAs. The C-terminal cassette represents a pseudo-enzyme that can stimulate RNA-related activities of the N-terminal cassette. However, the molecular mechanisms, by which the C-terminal cassette modulates the activities of the N-terminal unit remain elusive. Here, we show that N-and C-terminal cassettes adopt vastly different relative orientations in a crystal structure of Brr2 in complex with an activating domain of the spliceosomal Prp8 protein as compared to the crystal structure of isolated Brr2. Likewise, the cassettes occupy different relative positions and engage in different intercassette contacts during different stages of splicing. Engineered disulfide bridges that lock the cassettes in two different relative orientations have opposite effects on RNA-related activities of the N-terminal cassette compared to the unrestrained protein.Moreover, different relative positioning of the cassettes strongly influences ATP hydrolysis by the N-terminal cassette. Our results demonstrate that the inactive C-terminal cassette of Brr2 can exert both positive and negative influence on the active N-terminal helicase unit from a distance. Medicine,

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A new role for FBP21 as regulator of Brr2 helicase activity

Cited by 28 publications

References 61 publications

Structural Insights into Nuclear pre-mRNA Splicing in Higher Eukaryotes

Structural Insights into Nuclear pre-mRNA Splicing in Higher Eukaryotes

Intrinsically Disordered Protein Ntr2 Modulates the Spliceosomal RNA Helicase Brr2

Positive and negative intra-molecular modulation in a dual-cassette RNA helicase

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