Rab9A is required for delivery of cargo from recycling endosomes to melanosomes

<jats:title>Summary</jats:title><jats:p>Melanosomes are a type of lysosome‐related organelle that is commonly defective in Hermansky–Pudlak syndrome. Biogenesis of melanosomes is regulated by <jats:styled-content style="fixed-case">BLOC</jats:styled-content>‐1, ‐2, ‐3, or <jats:styled-content style="fixed-case">AP</jats:styled-content>‐1, ‐3 complexes, which mediate cargo transport from recycling endosomes to melanosomes. Although several Rab <jats:styled-content style="fixed-case">GTP</jats:styled-content>ases have been shown to regulate these trafficking steps, the precise role of Rab9A remains unknown. Here, we found that a cohort of Rab9A associates with the melanosomes and its knockdown in melanocytes results in hypopigmented melanosomes due to mistargeting of melanosomal proteins to lysosomes. In addition, the Rab9A‐depletion phenotype resembles Rab38/32‐inactivated or <jats:styled-content style="fixed-case">BLOC</jats:styled-content>‐3‐deficient melanocytes, suggesting that Rab9A works in line with <jats:styled-content style="fixed-case">BLOC</jats:styled-content>‐3 and Rab38/32 during melanosome cargo transport. Furthermore, silencing of Rab9A, Rab38/32 or its effector <jats:styled-content style="fixed-case">VARP</jats:styled-content>, or <jats:styled-content style="fixed-case">BLOC</jats:styled-content>‐3‐deficiency in melanocytes decreased the length of <jats:styled-content style="fixed-case">STX</jats:styled-content>13‐positive recycling endosomal tubules and targeted the <jats:styled-content style="fixed-case">SNARE</jats:styled-content> to lysosomes. This result indicates a defect in directing recycling endosomal tubules to melanosomes. Thus, Rab9A and its co‐regulatory <jats:styled-content style="fixed-case">GTP</jats:styled-content>ases control <jats:styled-content style="fixed-case">STX</jats:styled-content>13‐mediated cargo delivery to maturing melanosomes.</jats:p>