Entry of Botulinum Neurotoxin Subtypes A1 and A2 into Neurons

Botulinum neurotoxins (BoNTs) are the most toxic proteins for humans but also are common therapies for neurological diseases. BoNTs are dichain toxins, comprising an N-terminal catalytic domain (LC) disulfide bond linked to a C-terminal heavy chain (HC) which includes a translocation domain (H_N) and a receptor binding domain (H_C). Recently, the BoNT serotype A (BoNT/A) subtypes A1 and A2 were reported to possess similar potencies but different rates of cellular intoxication and pathology in a mouse model of botulism. The current study measured H_CA1 and H_CA2 entry into rat primary neurons and cultured Neuro2A cells. We found that there were two sequential steps during the association of BoNT/A with neurons. The initial step was ganglioside dependent, while the subsequent step involved association with synaptic vesicles. H_CA1 and H_CA2 entered the same population of synaptic vesicles and entered cells at similar rates. The primary difference was that H_CA2 had a higher degree of receptor occupancy for cells and neurons than HcA1. Thus, H_CA2 and H_CA1 share receptors and entry pathway but differ in their affinity for receptor. The initial interaction of H_CA1 and H_CA2 with neurons may contribute to the unique pathologies of BoNT/A1 and BoNT/A2 in mouse models.