UNC-45A Is a Nonmuscle Myosin IIA Chaperone Required for NK Cell Cytotoxicity via Control of Lytic Granule Secretion

Abstract

NK cell’s killing is a tightly regulated process under the control of specific cytoskeletal proteins. This includes Wiskott-Aldrich-Syndrome protein, Wiskott-Aldrich-Syndrome protein-interacting protein, cofilin, Munc13-4, and nonmuscle myosin IIA (NMIIA). These proteins play a key role in controlling NK-mediated cytotoxicity either via regulating the attachment of lytic granules to the actin-based cytoskeleton or via promoting the cytoskeletal reorganization that is requisite for lytic granule release. UNC-45A is a highly conserved member of the UNC-45/CRO1/She4p family of proteins that act as chaperones for both conventional and nonconventional myosin. Although we and others have shown that in lower organisms and in mammalian cells NMIIA-associated functions, such as cytokinesis, cell motility, and organelle trafficking, are dependent upon the presence of UNC-45A, its role in NK-mediated functions is largely unknown. In this article, we describe UNC-45A as a key regulator of NK-mediated cell toxicity. Specifically we show that, in human NK cells, UNC-45A localize at the NK cell immunological synapse of activated NK cells and is part of the multiprotein complex formed during NK cell activation. Furthermore, we show that UNC-45A is disposable for NK cell immunological synapse formation and lytic granules reorientation but crucial for lytic granule exocytosis. Lastly, loss of UNC-45A leads to reduced NMIIA binding to actin, suggesting that UNC-45A is a crucial component in regulating human NK cell cytoskeletal dynamics via promoting the formation of actomyosin complexes.

Loss-of-Function Mutations in UNC45A Cause a Syndrome Associating Cholestasis, Diarrhea, Impaired Hearing, and Bone Fragility

“…UNC45A appears to be ubiquitously expressed and has been postulated to be involved in cytoskeletal functions, such as cell division or exocytosis. 27,28 These data are consistent with the multi-organ defects observed in our affected individuals, as well as the structural pathology in the gut of our zebrafish mutants. However, other features of the protein are not reflected in the phenotype seen in humans.…”

UNC-45A is required for neurite extension via controlling NMII activation

“…More recently, we showed that UNC-45A promotes the NMII-associated cytoskeletal reorganization required during cell secretion (Iizuka et al ., 2015). Thus we sought to investigate whether UNC-45A plays a role in neuronal development.…”

UNC-45A is preferentially expressed in epithelial cells and binds to and co-localizes with interphase MTs

“…This includes studies from our laboratories showing that UNC-45A co-localizes with NMII in mammalian cells including cancer cells, NK cells, and neurons. 17,19,20 This also includes a study from Dr. Lappalainen's group showing that the UCS domain of UNC-45A co-localizes with stress fibers in the U2OS cell line where it promotes myosin folding and stress fibers assembly. 6 More recently we and others have shown that UNC-45A has independent functions in actomyosin and MT systems.…”

“…Third, fixation methods other than cold methanol (which we have used here and previously to show that UNC-45A is a MAP 5 ) may not be suitable for MT localization or co-localization studies because their either do not preserve MTs well, or interfere with antigen binding. With regards to this point, our work on co-localization of UNC-45A with NMII in NK cells and neurons 17,19,20 as well work from Drs. Lappalainen and Chadli's groups were all done using formaldehyde fixation because that is the preferred method for studying the actomyosin system.…”