A team from the University of Tokyo and Juntendo University has identified a previously unknown domain in the motor protein kinesin-2 that acts like a molecular “hook” to recognise and attach cargo in neurons.

Using high-resolution cryo-electron microscopy, they mapped the “HAC” (Hook-like Adaptor and Cargo-binding) domain, which enables the motor protein to pick up the right cellular cargo and deliver it with precision. This discovery sheds light on how neurons maintain efficient transport and has implications for understanding neurodegenerative diseases and other intracellular transport disorders.

Using cryo-electron microscopy and molecular dynamics simulations, the team reconstructed the structure of the heterotrimeric kinesin-2 complex (KIF3A/KIF3B/KAP3) bound to the cargo protein adenomatous polyposis coli (APC). Their analysis revealed a previously unknown structural motif in the tail region of KIF3A and KIF3B, termed the hook-like adaptor and cargo-binding (HAC) domain. This domain acts as a molecular “hook,” enabling the motor protein to assemble adaptors and recognize cargo with high specificity.

“Our study has uncovered a previously unknown ‘hook-like’ structural element, the HAC domain, in the tail of the motor protein kinesin-2,” Prof. Hirokawa said. “This domain acts as a molecular ‘connector’ that allows the motor to correctly recognize and transport its cargo inside cells.”

The HAC domain consists of a helix–β-hairpin–helix motif that forms a scaffold for the adaptor protein KAP3 and the cargo protein APC. The researchers identified four distinct binding interfaces between KIF3 and KAP3, with KIF3A playing the dominant role in cargo recognition. The structure also resembles cargo-binding architectures seen in other motor proteins, such as dynein and kinesin-1, suggesting a shared framework for cargo recognition.

“This discovery builds on decades of research from our laboratory, which first identified and characterised the complete family of mammalian kinesin motor proteins in the 1980s and 1990s and later revealed how these molecular ‘vehicles’ move along the cytoskeletal ‘highways’ of the cell,” Prof. Hirokawa said. “While we have long understood how these motors travel, the remaining mystery was how they know what to carry. Our new findings provide the first atomic-level insight into this ‘logistics code’ of cellular transport.”

The team validated the structural model using cross-linking mass spectrometry, biochemistry, and neuronal cell biology, demonstrating that the HAC domain binds specifically to the ARM repeat region of APC, a tumor suppressor protein involved in neuronal RNA transport. KIF3A contributed the majority of binding energy, while KIF3B played a supporting structural role.

Defects in intracellular transport have been linked to neurodegenerative diseases, neurodevelopmental disorders, and ciliopathies. Understanding motor protein cargo recognition offers a molecular basis for developing new diagnostic tools and therapeutic approaches. The study also highlights potential applications in drug discovery targeting motor-cargo interactions and the design of artificial transport systems.