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A targeted siRNA screen identifies regulators of Cdc42 activity at the natural killer cell immunological synapse.
- Source :
-
Science signaling [Sci Signal] 2011 Nov 29; Vol. 4 (201), pp. ra81. Date of Electronic Publication: 2011 Nov 29. - Publication Year :
- 2011
-
Abstract
- Natural killer (NK) cells kill tumor cells and virally infected cells, and an effective NK cell response requires processes, such as motility, recognition, and directional secretion, that rely on cytoskeletal rearrangement. The Rho guanosine triphosphatase (GTPase) Cdc42 coordinates cytoskeletal reorganization downstream of many receptors. The Rho-related GTPase from plants 1 (ROP1) exhibits oscillatory activation behavior at the apical plasma membrane of growing pollen tubes; however, a similar oscillation in Rho GTPase activity has so far not been demonstrated in mammalian cells. We hypothesized that oscillations in Cdc42 activity might occur within NK cells as they interact with target cells. Through fluorescence lifetime imaging of a Cdc42 biosensor, we observed that in live NK cells forming immunological synapses with target cells, Cdc42 activity oscillated after exhibiting an initial increase. We used protein-protein interaction networks and structural databases to identify candidate proteins that controlled Cdc42 activity, leading to the design of a targeted short interfering RNA screen. The guanine nucleotide exchange factors RhoGEF6 and RhoGEF7 were necessary for Cdc42 activation within the NK cell immunological synapse. In addition, the kinase Akt and the p85α subunit of phosphoinositide 3-kinase (PI3K) were required for Cdc42 activation, the periodicity of the oscillation in Cdc42 activity, and the subsequent polarization of cytotoxic vesicles toward target cells. Given that PI3Ks are targets of tumor therapies, our findings suggest the need to monitor innate immune function during the course of targeted therapy against these enzymes.
- Subjects :
- Biological Clocks genetics
Biological Clocks immunology
Cell Line, Transformed
Cell- and Tissue-Based Therapy methods
Class Ia Phosphatidylinositol 3-Kinase genetics
Class Ia Phosphatidylinositol 3-Kinase immunology
Class Ia Phosphatidylinositol 3-Kinase metabolism
Cytoskeleton genetics
Cytoskeleton immunology
Cytoskeleton metabolism
Enzyme Activation genetics
Enzyme Activation immunology
Guanine Nucleotide Exchange Factors genetics
Guanine Nucleotide Exchange Factors immunology
Guanine Nucleotide Exchange Factors metabolism
Humans
Immunity, Cellular genetics
Immunity, Innate genetics
Immunological Synapses enzymology
Immunological Synapses genetics
Killer Cells, Natural enzymology
Neoplasms enzymology
Neoplasms genetics
Neoplasms immunology
Neoplasms therapy
Proto-Oncogene Proteins c-akt
Rho Guanine Nucleotide Exchange Factors
cdc42 GTP-Binding Protein genetics
cdc42 GTP-Binding Protein metabolism
Immunological Synapses immunology
Killer Cells, Natural immunology
RNA, Small Interfering
cdc42 GTP-Binding Protein immunology
Subjects
Details
- Language :
- English
- ISSN :
- 1937-9145
- Volume :
- 4
- Issue :
- 201
- Database :
- MEDLINE
- Journal :
- Science signaling
- Publication Type :
- Academic Journal
- Accession number :
- 22126964
- Full Text :
- https://doi.org/10.1126/scisignal.2001729