title: Alternative splicing of the neurofibromatosis type 1 pre-mRNA is regulated by the muscleblind-like proteins and the CUG-BP and ELAV-like factors.
abstract: BACKGROUND
  Alternative splicing is often subjected to complex regulatory control that involves many protein factors and cis-acting RNA sequence elements. One major challenge is to identify all of the protein players and define how they control alternative expression of a particular exon in a combinatorial manner. The Muscleblind-like (MBNL) and CUG-BP and ELAV-Like family (CELF) proteins are splicing regulatory proteins, which function as antagonists in the regulation of several alternative exons. Currently only a limited number of common targets of MBNL and CELF are known that are antagonistically regulated by these two groups of proteins.
  
  
  RESULTS
  Recently, we identified neurofibromatosis type 1 (NF1) exon 23a as a novel target of negative regulation by CELF proteins. Here we report that MBNL family members are positive regulators of this exon. Overexpression of MBNL proteins promote exon 23a inclusion in a low MBNL-expressing cell line, and simultaneous siRNA-mediated knockdown of MBNL1 and MBNL2 family members in a high MBNL-expressing cell line promotes exon 23a skipping. Importantly, these two groups of proteins antagonize each other in regulating inclusion of exon 23a. Furthermore, we analyzed the binding sites of these proteins in the intronic sequences upstream of exon 23a by UV cross-linking assays. We show that in vitro, in addition to the previously identified preferred binding sequence UGCUGU, the MBNL proteins need the neighboring sequences for optimal binding.
  
  
  CONCLUSIONS
  This study along with our previous work that demonstrated roles for Hu, CELF, and TIA-1 and TIAR proteins in the regulation of NF1 exon 23a establish that this exon is under tight, complex control.
authors.affiliation: Department of Genetics, Case Western Reserve University, Cleveland, OH, 44106, USA.
authors.forename: Victoria A
authors.initials: VA
authors.lastname: Fleming
authors.forename: Cuiyu
authors.initials: C
authors.lastname: Geng
authors.forename: Andrea N
authors.initials: AN
authors.lastname: Ladd
authors.forename: Hua
authors.initials: H
authors.lastname: Lou
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D051743:CCAAT-Enhancer-Binding Protein-delta / Q000378:metabolism*; D000067879:CELF1 Protein; D002460:Cell Line; D005091:Exons; D006367:HeLa Cells; D006801:Humans; D009456:Neurofibromatosis 1 / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051381:Rats
chemical_list: C497252:CEBPD protein, human; D000067879:CELF1 Protein; C102843:CELF1 protein, human; C493663:MBNL1 protein, human; C499444:MBNL2 protein, human; D012322:RNA Precursors; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D051743:CCAAT-Enhancer-Binding Protein-delta