Impaired phrenic nerve axon development and diaphragm neuromuscular junction formation in embryonic Cyfip2-null mice

diaphragms showed spatial heterogeneity: sparse regions exhibited a significant increase in endplate bandwidth, whereas dense regions showed a decreasing trend. Further analysis using synaptophysin and α-bungarotoxin labeling revealed reduced pre- and post-synaptic puncta density and decreased colocalization, despite preserved puncta intensity and volume, indicating impaired synaptic organization. Together, these findings demonstrate that CYFIP2 is required for proper phrenic nerve innervation and NMJ organization during embryonic development. This study extends the functional scope of CYFIP2 to the PNS and establishes the diaphragm as a tractable model for investigating peripheral mechanisms underlying CYFIP2-associated neurodevelopmental disorders.