Dissecting the Dual Activity of Cellular Retinoic Acid Binding Protein 2 (CRABP2)

Abstract

Cellular retinoic acid‐binding protein 2 (CRABP2) displays two distinct functions. The classical function of this protein is to directly deliver retinoic acid (RA) to retinoic acid receptors (RARs), thereby enhancing the transcriptional activation of RARs and upregulating expression of target genes, including some involved in suppression of tumor growth. We have recently elucidated a second function of CRABP2, which is exerted in the absence of RA. We found that apo‐CRABP2 cooperates with the RNA‐binding and stabilizing protein HuR to upregulate and stabilize mRNAs. CRABP2 directly interacts with HuR and enhances its affinity for target mRNAs, including some involved in regulating oncogenic properties. Notably, while apo‐CRABP2 is cytosolic and binds to HuR, the complex dissociates upon binding of RA. Holo‐CRABP2 then translocates to the nucleus where it delivers RA to RAR. CRABP2 then returns to the cytosol and re‐assocaites with HuR, showing that CRABP2 is able to perform both of its functions in parallel. The goal of this work is to determine the structural charcteristics of CRABP2 that mediate it's interaction with HuR.The RA‐responsiveness of the interaction between CRABP2 and HuR suggests that the residues on CRABP2 that stabilize these interactions can “sense” ligand‐binding, i.e. that they undergo conformational changes upon ligation of CRABP2. Analysis of the reported three dimensional X‐ray crystal structures of holo‐ and apo‐CRABP2 revealed two surface residues, Arg60 and Lys107, whose side chains are placed at different conformations in the absence and presence of the ligand. We have found that Arg60 is necessary for the RA‐responsiveness of the CRABP2‐HuR complex and mutation of this residue prevents CRABP2 from cooperating with RAR. Additionally, we have found that Lys107 is necessary for CRABP2 to cooperate with HuR, but mutation of this residue does not interfere with the ability of CRABP2 to cooperate with RAR. As CRABP2 displays tumor suppressive activities, current work is examining which of CRABP2's functions is responsible for the tumor supressive activity of the protein.Support or Funding InformationThis work was supported by NIH grant R01‐CA166955. A.V. was partially supported by NIH grant R25‐CA148052.