Structure of transportin SR2, a karyopherin involved in human disease, in complex with Ran

Abstract

Transportin SR2 (TRN-SR2) is a β-type karyopherin responsible for the nuclear import of specific cargoes, including serine/arginine-rich splicing factors. The protein has been implicated in a variety of human diseases, including HIV infection, primary biliary cirrhosis and limb-girdle muscular dystrophy 1F. Towards understanding its molecular mechanism, a 2.9 Å resolution crystal structure of human TRN-SR2 complexed with the small GTPase Ran has been determined. TRN-SR2 is composed of 20 α-helical HEAT repeats forming a solenoid-like fold. The first nine repeats form a `cradle' for the binding of RanGTP, revealing similarities but also differences with respect to the related importin 13 complex.

The HIV-1 Integrase Mutant R263A/K264A Is 2-fold Defective for TRN-SR2 Binding and Viral Nuclear Import

“…Independently, two genome-wide siRNA screens also identified TRN-SR2 as a crucial cofactor for HIV replication (16,17). While this paper was in revision, two papers were published revealing the crystal structure of TRN-SR2 (57,58). The TRN-SR2 mutant, D750R/D751R, was defective for interaction with CPSF6 and proved unable to support HIV-1 infection although reportedly retaining WT level of HIV IN and ASF/SF2 binding (57).…”

N-terminal half of transportin SR2 interacts with HIV integrase

“…Thus AlphaScreen assays involving both the "one-third" constructs of TRN-SR2 and individual inside helices of each HEAT consistently indicate that the binding of IN occurs within the N-terminal half of TRN-SR2. Importantly, we have previously established that the same region of TRN-SR2 is involved in the complex formation with RanGTP (37). Because RanGTP effectively displaces full-length IN from TRN-SR2 (40), we conclude that RanGTP directly competes with HIV IN for binding to TRN-SR2.…”

“…It is interesting to compare this model with the available crystal structures of TRN-SR2 complexes with RanGTP and ASF/SF2 (Fig. 7) (28,37).…”

“…Instead, we have resorted to SAXS experiments that can provide information about the overall structure of the macromolecular complex in a native solution environment (48,49). Thus, we have compared the solution structure of the TRN-SR2⅐CCD-CTD complex and two other TRN-SR2-based protein complexes that have been previously resolved using X-ray crystallography, namely those with RanGTP (37) and with a fragment of ASF/SF2 (28), an important natural cargo of TRN-SR2 (29,50,51). The latter fragment contained residues 106 -230 of ASF/SF2, which corresponds to the second RNA recognition motif (RRM2) and a nuclear localization signal enriched in arginine and serine residues, the so-called RS domain.…”

“…The competitive binding of GTP-charged Ran induces cargo release (34). Previous mutagenesis and structural studies have shown that TRN-SR2 binds RanGTP via the N-terminal region, whereas the cargo attachment typically requires the arginine-rich helix 15b of TRN-SR2 (28,37). The TRN-SR2⅐RanGTP complex eventually returns to the cytoplasm where the RanGTPase-activating protein (RanGAP) together with the Ran-binding protein 1 (RanBP1) lead to the GTP hydrolysis of Ran, which in turn induces its release from TRN-SR2 (39).…”