Propiverine, a frequently-prescribed
pharmaceutical for the treatment of symptoms associated with
overactive bladder syndrome, provoked massive intranuclear and cytosolic
protein inclusions in rat proximal tubule epithelium, primarily consisting of the
peroxisomal targeting signal 1 (PTS1) containing
protein d-amino acid oxidase (DAAO). As this type of nephropathy was also observed for other drugs, the aim was to determine whether
propiverine interferes with trafficking and/or import of peroxisomal
proteins. To elucidate this, DAAO- and
propiverine-specific interaction partners from human HEK293 and rat WKPT cell lines and rat kidney and liver homogenate were determined using co-immunoprecipitation with subsequent nano-ESI-LC-MS/MS analyses. Corroboration of the role of DAAO- and/or
propiverine-specific interaction partners in the
drug-induced DAAO accumulation was sought via specific immunofluorescence staining of rat kidney sections from control and
propiverine-treated rats. Above analyses demonstrated the interaction of
propiverine with several
protein classes, foremost peroxisomal
proteins (DAAO, MFE2, HAOX2) and
proteins of the
protein quality control system, i.e. chaperones (HSP70 and DnaJ co-chaperones),
proteases and proteasomal
proteins (regulatory subunits of the
26S proteasome; Rpn1/2). The immunofluorescence analysis revealed mislocalization of many PTS1-proteins (DAAO, CAT, MFE2, ACOX1, EHHADH) in rat renal sections, strongly suggesting that
propiverine primarily binds to PTS1
proteins resulting in the formation of PTS1 but not PTS2 or peroxisomal
membrane protein (PMP) accumulations. Moreover, chaperones involved in peroxisomal trafficking (HSC70, DnaJB1) and peroxisomal biogenesis factor
proteins (PEX3, PEX5, PEX7), also presented with distinct mislocalization patterns. Concomitantly, an increased number of peroxisomes was observed, suggestive of a compensatory mechanism for the presumably suboptimally functioning peroxisomes. Overall, the data presented suggested that
propiverine interacts exclusively with DAAO or with a selected number of PTS1
proteins. The consequence of this interaction is the abrogated trafficking and peroxisomal import of PTS1
proteins concomitant with their nuclear and cytosolic accumulation due to inhibited degradation and imbalanced protein homeostasis.