TY - JOUR
T1 - A synthetic small molecule stalls pre-mRNA splicing by promoting an early-stage U2AF2-RNA complex
AU - Chatrikhi, Rakesh
AU - Feeney, Callen F.
AU - Pulvino, Mary J.
AU - Alachouzos, Georgios
AU - MacRae, Andrew J.
AU - Falls, Zackary
AU - Rai, Sumit
AU - Brennessel, William W.
AU - Jenkins, Jermaine L.
AU - Walter, Matthew J.
AU - Graubert, Timothy A.
AU - Samudrala, Ram
AU - Jurica, Melissa S.
AU - Frontier, Alison J.
AU - Kielkopf, Clara L.
N1 - Funding Information:
We are grateful to Prof. A.V. Smrcka (University of Michigan) for advice designing the screen. This work was supported by grants from the NIH ( R01 GM070503 to C.L.K. and R01 GM122279 to M.S.J.), the National Science Foundation (NSF CHE-1900050 to A.J.F.), UR Ventures (to C.L.K.), and the Edward P. Evans Foundation (to C.L.K., M.J.W., and T.A.G.). Work by R.S. was supported by an NIH Director’s Pioneer award ( DP1 OD006779 ), a Clinical and Translational Sciences ( NCATS ) award ( UL1 TR001412 ), and an NCATS ASPIRE Design Challenge award. A.J.MacR. was supported by an NIH training grant ( T32 GM08646 ). Z.F. was supported by the National Library of Medicine ( T15 LM012495 ). An NSF Major Research Instrumentation grant ( CHE-1725028 ) supported the X-ray diffractometer.
Funding Information:
We are grateful to Prof. A.V. Smrcka (University of Michigan) for advice designing the screen. This work was supported by grants from the NIH (R01 GM070503 to C.L.K. and R01 GM122279 to M.S.J.), the National Science Foundation (NSF CHE-1900050 to A.J.F.), UR Ventures (to C.L.K.), and the Edward P. Evans Foundation (to C.L.K. M.J.W. and T.A.G.). Work by R.S. was supported by an NIH Director's Pioneer award (DP1 OD006779), a Clinical and Translational Sciences (NCATS) award (UL1 TR001412), and an NCATS ASPIRE Design Challenge award. A.J.MacR. was supported by an NIH training grant (T32 GM08646). Z.F. was supported by the National Library of Medicine (T15 LM012495). An NSF Major Research Instrumentation grant (CHE-1725028) supported the X-ray diffractometer. C.L.K. R.C. M.J.P. G.A. A.J.F. and M.S.J. conceived experiments in collaboration with all authors. R.C. screened small-molecule libraries. C.F.F. accomplished in vitro experiments with guidance from J.L.J. M.J.P. and C.L.K. A.J.MacR. and M.S.J. analyzed spliceosome assemblies. Z.F. and R.S. performed computational docking. G.A. W.W.B. and A.J.F. carried out synthesis, purification, and characterization of compounds. S.R. M.J.W. and T.A.G. constructed cell lines. M.J.P. accomplished biological experiments. C.L.K. wrote the paper with contributions from R.C. M.S.J. G.A. A.J.F. M.J.W. and T.A.G. and input from all authors. The authors declare no competing interests. One or more of the authors of this paper received support from a program designed to increase minority representation in science.
Publisher Copyright:
© 2021 Elsevier Ltd
PY - 2021/8/19
Y1 - 2021/8/19
N2 - Dysregulated pre-mRNA splicing is an emerging Achilles heel of cancers and myelodysplasias. To expand the currently limited portfolio of small-molecule drug leads, we screened for chemical modulators of the U2AF complex, which nucleates spliceosome assembly and is mutated in myelodysplasias. A hit compound specifically enhances RNA binding by a U2AF2 subunit. Remarkably, the compound inhibits splicing of representative substrates and stalls spliceosome assembly at the stage of U2AF function. Computational docking, together with structure-guided mutagenesis, indicates that the compound bridges the tandem U2AF2 RNA recognition motifs via hydrophobic and electrostatic moieties. Cells expressing a cancer-associated U2AF1 mutant are preferentially killed by treatment with the compound. Altogether, our results highlight the potential of trapping early spliceosome assembly as an effective pharmacological means to manipulate pre-mRNA splicing. By extension, we suggest that stabilizing assembly intermediates may offer a useful approach for small-molecule inhibition of macromolecular machines.
AB - Dysregulated pre-mRNA splicing is an emerging Achilles heel of cancers and myelodysplasias. To expand the currently limited portfolio of small-molecule drug leads, we screened for chemical modulators of the U2AF complex, which nucleates spliceosome assembly and is mutated in myelodysplasias. A hit compound specifically enhances RNA binding by a U2AF2 subunit. Remarkably, the compound inhibits splicing of representative substrates and stalls spliceosome assembly at the stage of U2AF function. Computational docking, together with structure-guided mutagenesis, indicates that the compound bridges the tandem U2AF2 RNA recognition motifs via hydrophobic and electrostatic moieties. Cells expressing a cancer-associated U2AF1 mutant are preferentially killed by treatment with the compound. Altogether, our results highlight the potential of trapping early spliceosome assembly as an effective pharmacological means to manipulate pre-mRNA splicing. By extension, we suggest that stabilizing assembly intermediates may offer a useful approach for small-molecule inhibition of macromolecular machines.
KW - S34F mutant
KW - U2AF
KW - U2AF
KW - U2AF1
KW - myelodysplastic syndrome
KW - ribonucleoprotein targeting
KW - spliceosome inhibition
KW - splicing factor mutation
KW - therapeutic strategy
UR - http://www.scopus.com/inward/record.url?scp=85112540054&partnerID=8YFLogxK
U2 - 10.1016/j.chembiol.2021.02.007
DO - 10.1016/j.chembiol.2021.02.007
M3 - Article
C2 - 33689684
AN - SCOPUS:85112540054
SN - 2451-9456
VL - 28
SP - 1145-1157.e6
JO - Cell Chemical Biology
JF - Cell Chemical Biology
IS - 8
ER -