RESUMEN
PTPRK is a receptor tyrosine phosphatase that is linked to the regulation of growth factor signalling and tumour suppression. It is stabilized at the plasma membrane by trans homophilic interactions upon cell-cell contact. PTPRK regulates cell-cell adhesion but is also reported to regulate numerous cancer-associated signalling pathways. However, the signalling mechanism of PTPRK remains to be determined. Here, we find that PTPRK regulates cell adhesion signalling, suppresses invasion and promotes collective, directed migration in colorectal cancer cells. In vivo, PTPRK supports recovery from inflammation-induced colitis. In addition, we confirm that PTPRK functions as a tumour suppressor in the mouse colon and in colorectal cancer xenografts. PTPRK regulates growth factor and adhesion signalling, and suppresses epithelial to mesenchymal transition (EMT). Contrary to the prevailing notion that PTPRK directly dephosphorylates EGFR, we find that PTPRK regulation of both EGFR and EMT is independent of its catalytic function. This suggests that additional adaptor and scaffold functions are important features of PTPRK signalling.
Asunto(s)
Transición Epitelial-Mesenquimal , Humanos , Animales , Ratones , Receptores ErbB/metabolismo , Receptores ErbB/genética , Transducción de Señal , Adhesión Celular/genética , Movimiento Celular , Línea Celular Tumoral , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Colitis/patología , Colitis/metabolismo , Colitis/genética , Colitis/inducido químicamente , Proteínas Tirosina Fosfatasas Clase 2 Similares a Receptores/metabolismo , Proteínas Tirosina Fosfatasas Clase 2 Similares a Receptores/genética , Intestinos/patologíaRESUMEN
Bach2 codes for a transcriptional regulator exerting major influences on T cell-mediated immune regulation. Effector CTLs derived from in vitro activation of murine CD8+ T cells showed increased proliferative and cytolytic capacity in the absence of BACH2. Before activation, BACH2-deficient splenic CD8+ T cells had a higher abundance of memory and reduced abundance of naïve cells compared to wild-type. CTLs derived from central memory T cells were more potently cytotoxic than those derived from naïve T cells, but even within separated subsets, BACH2-deficiency conferred a cytotoxic advantage. Immunofluorescence and electron microscopy revealed larger granules in BACH2-deficient compared to wild-type CTLs, and proteomic analysis showed an increase in granule content, including perforin and granzymes. Thus, the enhanced cytotoxicity observed in effector CTLs lacking BACH2 arises not only from differences in their initial differentiation state but also inherent production of enlarged cytolytic granules. These results demonstrate how a single gene deletion can produce a CTL super-killer.
Asunto(s)
Linfocitos T CD8-positivos , Citotoxicidad Inmunológica , Ratones , Animales , Eliminación de Gen , Proteómica , Linfocitos T Citotóxicos , Perforina , Granzimas/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genéticaRESUMEN
T cell receptor activation of naïve CD8+ T lymphocytes initiates their maturation into effector cytotoxic T lymphocytes (CTLs), which can kill cancer and virally infected cells. Although CTLs show an increased reliance on glycolysis upon acquisition of effector function, we found an essential requirement for mitochondria in target cellkilling. Acute mitochondrial depletion in USP30 (ubiquitin carboxyl-terminal hydrolase 30)deficient CTLs markedly diminished killing capacity, although motility, signaling, and secretion were all intact. Unexpectedly, the mitochondrial requirement was linked to mitochondrial translation, inhibition of which impaired CTL killing. Impaired mitochondrial translation triggered attenuated cytosolic translation, precluded replenishment of secreted killing effectors, and reduced the capacity of CTLs to carry out sustained killing. Thus, mitochondria emerge as a previously unappreciated homeostatic regulator of protein translation required for serial CTL killing.
Asunto(s)
Citotoxicidad Inmunológica/inmunología , Mitocondrias/enzimología , Proteínas Mitocondriales/metabolismo , Linfocitos T Citotóxicos/inmunología , Tioléster Hidrolasas/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Movimiento Celular/genética , Células Cultivadas , Citotoxicidad Inmunológica/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias/genética , Proteínas Mitocondriales/genética , Biosíntesis de Proteínas , Linfocitos T Citotóxicos/enzimología , Tioléster Hidrolasas/genéticaRESUMEN
By developing a high-density murine immunophenotyping platform compatible with high-throughput genetic screening, we have established profound contributions of genetics and structure to immune variation (http://www.immunophenotype.org). Specifically, high-throughput phenotyping of 530 unique mouse gene knockouts identified 140 monogenic 'hits', of which most had no previous immunologic association. Furthermore, hits were collectively enriched in genes for which humans show poor tolerance to loss of function. The immunophenotyping platform also exposed dense correlation networks linking immune parameters with each other and with specific physiologic traits. Such linkages limit freedom of movement for individual immune parameters, thereby imposing genetically regulated 'immunologic structures', the integrity of which was associated with immunocompetence. Hence, we provide an expanded genetic resource and structural perspective for understanding and monitoring immune variation in health and disease.
Asunto(s)
Infecciones por Enterobacteriaceae/inmunología , Variación Genética/genética , Ensayos Analíticos de Alto Rendimiento/métodos , Inmunofenotipificación/métodos , Infecciones por Salmonella/inmunología , Animales , Citrobacter/inmunología , Infecciones por Enterobacteriaceae/microbiología , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Modelos Animales , Salmonella/inmunología , Infecciones por Salmonella/microbiologíaRESUMEN
Mutations in the human NBEAL2 gene cause gray platelet syndrome (GPS), a bleeding diathesis characterized by a lack of α granules in platelets. The functions of the NBEAL2 protein have not been explored outside platelet biology, but there are reports of increased frequency of infection and abnormal neutrophil morphology in patients with GPS. We therefore investigated the role of NBEAL2 in immunity by analyzing the phenotype of Nbeal2-deficient mice. We found profound abnormalities in the Nbeal2-deficient immune system, particularly in the function of neutrophils and NK cells. Phenotyping of Nbeal2-deficient neutrophils showed a severe reduction in granule contents across all granule subsets. Despite this, Nbeal2-deficient neutrophils had an enhanced phagocyte respiratory burst relative to Nbeal2-expressing neutrophils. This respiratory burst was associated with increased expression of cytosolic components of the NADPH oxidase complex. Nbeal2-deficient NK cells were also dysfunctional and showed reduced degranulation. These abnormalities were associated with increased susceptibility to both bacterial (Staphylococcus aureus) and viral (murine CMV) infection in vivo. These results define an essential role for NBEAL2 in mammalian immunity.