CHERP

CHERP is a nuclear spliceosomal protein that regulates alternative pre-mRNA splicing as part of the U2 snRNP machinery (PMID:23148228, PMID:35269695). It localizes to the nucleus and nuclear speckles, associates with the U2 snRNA spliceosomal complex, and binds a phosphorylated form of RNA polymerase II (PMID:23148228, PMID:24078636). CHERP forms a tight, mutually stabilizing complex with SR140 (U2SURP) and SPF45 (RBM17) that represses short alternative exons flanked by suboptimal 3' splice sites and controls intron retention, acting on targets including FOXM1, SPDL1, UPF3A, and IP3R1 pre-mRNAs (PMID:24078636, PMID:30332651, PMID:30977118, PMID:34544891). Through these splicing events, CHERP is required for cell-cycle progression: its depletion drives nuclear accumulation of poly(A)+ RNA, G2/M arrest, DNA damage, and impaired proliferation, with the growth defect partially rescued by restoring UPF3A or by mimicking the FOXM1/SPDL1 splicing outcomes (PMID:30977118, PMID:34544891, PMID:35269695). An earlier model in which CHERP acted as a direct ER/cytoplasmic regulator of IP3R and RyR1 Ca2+ channels (PMID:10794731, PMID:21454501) was re-evaluated, with its effects on Ca2+ signaling and growth reinterpreted as indirect consequences of altered spliceosomal function (PMID:23148228). CHERP additionally binds ALG-2 in a Ca2+-dependent manner, providing a candidate link between Ca2+ signaling and splicing regulation (PMID:24078636).

1. 2000 Medium

   Established CHERP as a functional protein affecting Ca2+ homeostasis and proliferation, the founding observation that defined the gene's phenotypic footprint.

   Evidence Antisense knockdown in HEL cells with Ca2+ mobilization, DNA synthesis assays, and IP3R co-localization by immunofluorescence

   PMID:10794731

   Open questions at the time

   • Co-localization does not demonstrate direct molecular interaction with IP3R
   • Cannot distinguish direct Ca2+ channel regulation from indirect effects
   • Mechanism of the growth arrest unresolved
2. 2003 Medium

   Extended the Ca2+/proliferation phenotype to a second cell type and placed CHERP upstream of or parallel to ER Ca2+ release rather than altering IP3R levels.

   Evidence Antisense knockdown in Jurkat cells with fura-2 imaging, NFAT translocation, and cyclin D1 Western blotting

   PMID:12656674

   Open questions at the time

   • Did not identify the molecular target through which CHERP affects Ca2+ release
   • Indirect versus direct mechanism not resolved
3. 2011 Medium

   Reported a physical and functional link to the RyR1 Ca2+ channel, reinforcing the ER/SR-centric model of CHERP function.

   Evidence Co-purification of CHERP with a soluble cytosolic RyR1 fragment, SR co-localization, and siRNA + Ca2+ release assay in HEK-293 cells

   PMID:21454501

   Open questions at the time

   • Co-purification with an overexpressed fragment may not reflect endogenous direct binding
   • Did not exclude indirect contribution via splicing later proposed for CHERP
4. 2012 Medium

   Reassigned CHERP from a cytoplasmic Ca2+ channel regulator to a nuclear spliceosome-associated protein, redirecting the entire mechanistic interpretation of the earlier growth phenotypes.

   Evidence Subcellular fractionation, nuclear immunofluorescence, and co-IP with U2 snRNP components plus proliferation assays

   PMID:23148228

   Open questions at the time

   • Did not define which splicing targets account for the growth defect
   • Did not formally exclude a residual direct Ca2+ channel role
5. 2013 High

   Provided direct molecular evidence for nuclear splicing function and linked CHERP back to Ca2+ signaling via ALG-2, showing it binds pre-mRNA and regulates IP3R1 splicing.

   Evidence Co-IP with phospho-RNA Pol II and ALG-2, nuclear speckle imaging, live-cell recruitment imaging, RNA-IP, and siRNA + RT-PCR splicing assay in HT1080 cells

   PMID:24078636

   Open questions at the time

   • Functional consequence of Ca2+-dependent ALG-2 recruitment on splicing output not established
   • Genome-wide target spectrum not yet defined
6. 2018 High

   Defined a mutually stabilizing module of CHERP with RBM17 and U2SURP and showed it controls splicing of RNA-processing factors, establishing CHERP as part of a coordinated splicing regulatory complex.

   Evidence Reciprocal co-IP, siRNA knockdown, and RNA-seq splicing analysis across mouse and human cells with Western blotting

   PMID:30332651

   Open questions at the time

   • Stoichiometry and architecture of the CHERP-RBM17-U2SURP complex not resolved
   • Direct versus indirect splicing targets not fully separated
7. 2019 Medium

   Identified UPF3A exon 4 as a specific regulated target of the CHERP-SR140 complex and tied the proliferation defect causally to splicing output via rescue.

   Evidence Co-IP, siRNA knockdown, RT-PCR splicing assay, UPF3A overexpression rescue, and an in vivo mouse tumor model

   PMID:30977118

   Open questions at the time

   • Rescue was only partial, implying additional downstream targets
   • Mechanism linking the splicing defect to DNA breaks not detailed
8. 2021 High

   Defined the catalytic logic of the SPF45-SR140-CHERP complex as repression of short exons at suboptimal 3' splice sites and connected it to specific cell-cycle gene splicing controlling G2/M progression.

   Evidence Co-IP, siRNA knockdown of all three factors, RNA-seq, minigene assays, cell cycle analysis, and splicing-isoform epistasis in HeLa cells

   PMID:34544891

   Open questions at the time

   • Structural basis for recognition of suboptimal 3' splice sites unknown
   • Individual contribution of CHERP within the trimeric complex not isolated
9. 2022 Medium

   Demonstrated that CHERP is required for nuclear mRNA export competence and controls intron retention through U2 snRNP interaction, broadening its role beyond exon repression.

   Evidence siRNA knockdown, RNA-FISH for poly(A)+ RNA, RNA-seq intron-retention analysis, FACS cell cycle, and live-cell imaging in U2OS cells

   PMID:35269695

   Open questions at the time

   • Whether poly(A)+ RNA accumulation is a direct splicing consequence or a secondary effect not resolved
   • Sequence determinants of intron retention correlative rather than mechanistic

• How Ca2+/ALG-2 signaling functionally tunes CHERP-dependent splicing, and the structural basis by which the CHERP-SR140-SPF45 complex recognizes suboptimal splice sites, remain unresolved.
• No structural model of the CHERP-containing U2 snRNP subcomplex
• Functional output of Ca2+-dependent ALG-2 recruitment to CHERP speckles undefined
• Complete catalogue of physiologically critical splicing targets not established