SEMA3B switches axon-axon to axon-glia interactions required for unmyelinated axon envelopment and integrity

During peripheral nerve (PN) development, unmyelinated axons (nmAs) initially form tight fascicles before being separated and enveloped by non-myelinating Schwann cells (nmSCs), which are essential for maintaining nmA integrity. The mechanism underlying this transition from axon-axon to axon-glia interactions remains unclear. Here, we found that inactivating nmSC-derived SEMA3B or its axonal receptor components in mice led to incomplete nmA separation and envelopment by nmSCs, leading to nmA dysfunction and inducing hyperalgesia and allodynia. Conversely, increasing SEMA3B levels in nmSCs accelerated nmA separation and envelopment. SEMA3B transiently promoted nmA defasciculation accompanied by cell adhesion molecule (CAM) endocytosis, thus facilitating subsequent CAM-mediated nmA-mSC association. Restoring SEMA3B expression post-PN injury promoted nmA-nmSC re-association and alleviated hyperalgesia and allodynia. We propose that SEMA3B-induced CAM turnover facilitates the switch from axon-axon to axon-glia interactions, promoting nmA envelopment by nmSCs, which may be exploited to alleviate PN injury-induced pain by accelerating the restoration of nmA integrity. Overall design: We performed single-cell RNA sequencing (scRNA-seq) of sciatic nerves at five critical developmental times, i.e., P0, P7, P14, P28, and P60, encompassing the time window of RB (Remak bundle) assembly,to elucidate the mechaisms driving the transition from axon-axon to axon-glia interaction during RB assembly and markers fro nmscs during this transition.To investigate the presence of potential SEMA3B receptor components on nmAs during RB assembly, we conducted snRNA-seq of dorsal root ganglia (DRGs), comprising primary somatosensory neurons (SNs) projecting mAs and nmAs through the sciatic nerves, from P14 mice