PT - JOURNAL ARTICLE AU - Luke S Ferro AU - Lisa Eshun-Wilson AU - Mert Gölcük AU - Jonathan Fernandes AU - Teun Huijben AU - Eva Gerber AU - Amanda Jack AU - Katelyn Costa AU - Mert Gür AU - Qianglin Fang AU - Eva Nogales AU - Ahmet Yildiz TI - The mechanism of motor inhibition by microtubule-associated proteins AID - 10.1101/2020.10.22.351346 DP - 2020 Jan 01 TA - bioRxiv PG - 2020.10.22.351346 4099 - http://biorxiv.org/content/early/2020/10/23/2020.10.22.351346.1.short 4100 - http://biorxiv.org/content/early/2020/10/23/2020.10.22.351346.1.full AB - Microtubule (MT)-associated proteins (MAPs) regulate intracellular transport by selectively recruiting or excluding kinesin and dynein motors from MTs. We used single-molecule and cryo-electron imaging to determine the mechanism of MAP-motor interactions in vitro. Unexpectedly, we found that the regulatory role of a MAP cannot be predicted based on whether it overlaps with the motor binding site or forms liquid condensates on the MT. Although the MT binding domain (MTBD) of MAP7 overlaps with the kinesin-1 binding site, tethering of kinesin-1 by the MAP7 projection domain supersedes this inhibition and results in biphasic regulation of kinesin-1 motility. Conversely, the MTBD of tau inhibits dynein motility without overlapping with the dynein binding site or by forming tau islands on the MT. Our results indicate that MAPs sort intracellular cargos moving in both directions, as neither dynein nor kinesin can walk on a MAP-coated MT without favorably interacting with that MAP.HIGHLIGHTSMAP7 binds to a novel site and can coexist with tau on the MT.Kinesin-1 motility is biphasically regulated by MAP7 accumulation on the microtubule.MT decoration of MAPs inhibits motors even when they do not block the motor binding site.Motors need to interact with a MAP to walk on MAP-decorated MTsCompeting Interest StatementThe authors have declared no competing interest.