title: Mpl traffics to the cell surface through conventional and unconventional routes.
abstract: Myeloproliferative neoplasms (MPNs) are often characterized by JAK2 or calreticulin (CALR) mutations, indicating aberrant trafficking in pathogenesis. This study focuses on Mpl trafficking and Jak2 association using two model systems: human erythroleukemia cells (HEL; JAK2V617F) and K562 myeloid leukemia cells (JAK2WT). Consistent with a putative chaperone role for Jak2, Mpl and Jak2 associate on both intracellular and plasma membranes (shown by proximity ligation assay) and siRNA-mediated knockdown of Jak2 led to Mpl trapping in the endoplasmic reticulum (ER). Even in Jak2 sufficient cells, Mpl accumulates in punctate structures that partially colocalize with ER-tracker, the ER exit site marker (ERES) Sec31a, the autophagy marker LC3 and LAMP1. Mpl was fused to miniSOG, a genetically encoded tag for correlated light and electron microscopy. Results suggest that a fraction of Mpl is taken up into autophagic structures from the ER and routed to autolyososomes. Surface biotinylation shows that both immature and mature Mpl reach the cell surface; in K562 cells Mpl is also released in exosomes. Both forms rapidly internalize upon ligand addition, while recovery is primarily attributed to immature Mpl. Mpl appears to reach the plasma membrane via both conventional ER-Golgi and autolysosome secretory pathways, as well as recycling.
authors.affiliation: Department of Pathology & Cancer Center, University of New Mexico Health Sciences Center, Albuquerque, NM, 87131, USA; INSERM UMR892/CNRS UMR6299, Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Institut de Recherche Thérapeutique-Université de Nantes, Nantes, 44000, France.
authors.forename: Cédric
authors.initials: C
authors.lastname: Cleyrat
authors.forename: Anza
authors.initials: A
authors.lastname: Darehshouri
authors.forename: Mara P
authors.initials: MP
authors.lastname: Steinkamp
authors.forename: Mathias
authors.initials: M
authors.lastname: Vilaine
authors.forename: Daniela
authors.initials: D
authors.lastname: Boassa
authors.forename: Mark H
authors.initials: MH
authors.lastname: Ellisman
authors.forename: Sylvie
authors.initials: S
authors.lastname: Hermouet
authors.forename: Bridget S
authors.initials: BS
authors.lastname: Wilson
mesh_terms: D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D055354:Exosomes / Q000378:metabolism; D006801:Humans; D053614:Janus Kinase 2 / Q000378:metabolism; D020014:K562 Cells; D051907:Lysosomal Membrane Proteins / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D021381:Protein Transport / Q000502:physiology; D053628:Receptors, Thrombopoietin / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000378:metabolism; D052118:Lysosomal-Associated Membrane Protein 1
chemical_list: D053614:Janus Kinase 2; D051907:Lysosomal Membrane Proteins; D008869:Microtubule-Associated Proteins; D053628:Receptors, Thrombopoietin; D033921:Vesicular Transport Proteins; C489116:LAMP1 protein, human; D052118:Lysosomal-Associated Membrane Protein 1; C477012:MAP1LC3A protein, human; C507455:SEC31A protein, human; C082218:MPL protein, human; C507924:JAK2 protein, human
keywords: JAK2; MPNs; Mpl; autophagy; miniSOG