Parylation dependent nuclear export of HMGB1 via exportin 1 contributes to morphine tolerance

Nucleocytoplasmic transport has been implicated in chronic pain, particularly through importin-mediated nuclear import. However, the role of nuclear export in opioid-induced adaptations remains unclear. Exportin 1 (XPO1), a key nuclear export protein, has not been studied in the context of morphine tolerance. This study explores the contribution of XPO1 and its interaction with pro-inflammatory mediators in the development of opioid tolerance and hyperalgesia.

A rat model of chronic morphine administration was used to assess changes in spinal XPO1 expression and activity. Phosphorylation status of XPO1 and upstream kinases were evaluated by immunoblotting. Proteomic analysis of cerebrospinal fluid (CSF) was performed to identify secreted factors associated with morphine exposure. Co-immunoprecipitation and in vitro assays were used to examine the interaction between XPO1 and high mobility group box 1 (HMGB1). The role of PARP1-mediated poly(ADP-ribosyl)ation (PARylation) in regulating HMGB1 nuclear export was also investigated. Behavioral assays were used to assess the impact of pharmacological inhibition of XPO1 and PARP1 on morphine tolerance and nociceptive hypersensitivity.

Chronic morphine exposure led to significant upregulation of XPO1 in spinal neurons, accompanied by phosphorylation at serine 1010 by serine/threonine kinase 38 (STK38), which enhanced its nuclear export function. CSF proteomics revealed elevated levels of HMGB1, a pro-inflammatory mediator. XPO1 inhibition suppressed HMGB1 secretion. Mechanistically, PARP1-mediated PARylation of HMGB1 was essential for its interaction with XPO1 and subsequent nuclear export. Combined low-dose inhibition of XPO1 and PARP1 reversed established morphine tolerance and alleviated mechanical hypersensitivity. However, intrathecal administration of recombinant HMGB1 abolished these effects, reinstating morphine tolerance.

These findings reveal a novel mechanism by which STK38-driven phosphorylation of XPO1 and PARP1-mediated modification of HMGB1 coordinate nuclear export and extracellular signaling in the context of opioid tolerance. Dual inhibition of XPO1 and PARP1 represents a promising therapeutic strategy to suppress neuroinflammation and enhance opioid analgesic efficacy.