RESUMEN
Aberrant activation of mTOR signaling in acute myeloid leukemia (AML) results in a survival advantage that promotes the malignant phenotype. To improve our understanding of factors that contribute to mammalian target of rapamycin (mTOR) signaling activation and identify novel therapeutic targets, we searched for unique interactors of mTOR complexes through proteomics analyses. We identify cyclin dependent kinase 9 (CDK9) as a novel binding partner of the mTOR complex scaffold protein, mLST8. Our studies demonstrate that CDK9 is present in distinct mTOR-like (CTOR) complexes in the cytoplasm and nucleus. In the nucleus, CDK9 binds to RAPTOR and mLST8, forming CTORC1, to promote transcription of genes important for leukemogenesis. In the cytoplasm, CDK9 binds to RICTOR, SIN1, and mLST8, forming CTORC2, and controls messenger RNA (mRNA) translation through phosphorylation of LARP1 and rpS6. Pharmacological targeting of CTORC complexes results in suppression of growth of primitive human AML progenitors in vitro and elicits strong antileukemic responses in AML xenografts in vivo.
Asunto(s)
Carcinogénesis/efectos de los fármacos , Quinasa 9 Dependiente de la Ciclina/antagonistas & inhibidores , Leucemia Mieloide Aguda/tratamiento farmacológico , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Diana Mecanicista del Complejo 2 de la Rapamicina/metabolismo , ARN Mensajero/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Animales , Antimetabolitos Antineoplásicos/farmacología , Apoptosis , Biomarcadores de Tumor/metabolismo , Carcinogénesis/metabolismo , Carcinogénesis/patología , Proliferación Celular , Quinasa 9 Dependiente de la Ciclina/genética , Quinasa 9 Dependiente de la Ciclina/metabolismo , Citarabina/farmacología , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Ratones , Ratones Desnudos , Fosforilación , Biosíntesis de Proteínas , Proteoma/análisis , ARN Mensajero/efectos de los fármacos , ARN Mensajero/genética , Transducción de Señal , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Primary myelofibrosis (PMF) is a clonal hematologic malignancy characterized by BM fibrosis, extramedullary hematopoiesis, circulating CD34+ cells, splenomegaly, and a propensity to evolve to acute myeloid leukemia. Moreover, the spleen and BM of patients harbor atypical, clustered megakaryocytes, which contribute to the disease by secreting profibrotic cytokines. Here, we have revealed that megakaryocytes in PMF show impaired maturation that is associated with reduced GATA1 protein. In investigating the cause of GATA1 downregulation, our gene-expression study revealed the presence of the RPS14-deficient gene signature, which is associated with defective ribosomal protein function and linked to the erythroid lineage in 5q deletion myelodysplastic syndrome. Surprisingly, reduced GATA1 expression and impaired differentiation were limited to megakaryocytes, consistent with a proproliferative effect of a GATA1 deficiency on this lineage. Importantly, expression of GATA1 effectively rescued maturation of PMF megakaryocytes. Together, these results suggest that ribosomal deficiency contributes to impaired megakaryopoiesis in myeloproliferative neoplasms.
Asunto(s)
Regulación hacia Abajo , Factor de Transcripción GATA1/biosíntesis , Megacariocitos/metabolismo , Mielofibrosis Primaria/metabolismo , Trombopoyesis , Animales , Deleción Cromosómica , Cromosomas Humanos Par 5/genética , Cromosomas Humanos Par 5/metabolismo , Factor de Transcripción GATA1/genética , Humanos , Megacariocitos/patología , Ratones , Mielofibrosis Primaria/genética , Mielofibrosis Primaria/patología , Proteínas Ribosómicas/biosíntesis , Proteínas Ribosómicas/genéticaRESUMEN
Glioblastoma multiforme remains the deadliest malignant brain tumor, with glioma stem cells (GSC) contributing to treatment resistance and tumor recurrence. We have identified MAPK-interacting kinases (MNK) as potential targets for the GSC population in glioblastoma multiforme. Isoform-level subtyping using The Cancer Genome Atlas revealed that both MNK genes (MKNK1 and MKNK2) are upregulated in mesenchymal glioblastoma multiforme as compared with other subtypes. Expression of MKNK1 is associated with increased glioma grade and correlated with the mesenchymal GSC marker, CD44, and coexpression of MKNK1 and CD44 predicts poor survival in glioblastoma multiforme. In established and patient-derived cell lines, pharmacologic MNK inhibition using LY2801653 (merestinib) inhibited phosphorylation of the eukaryotic translation initiation factor 4E, a crucial effector for MNK-induced mRNA translation in cancer cells and a marker of transformation. Importantly, merestinib inhibited growth of GSCs grown as neurospheres as determined by extreme limiting dilution analysis. When the effects of merestinib were assessed in vivo using an intracranial xenograft mouse model, improved overall survival was observed in merestinib-treated mice. Taken together, these data provide strong preclinical evidence that pharmacologic MNK inhibition targets mesenchymal glioblastoma multiforme and its GSC population. IMPLICATIONS: These findings raise the possibility of MNK inhibition as a viable therapeutic approach to target the mesenchymal subtype of glioblastoma multiforme. Mol Cancer Res; 14(10); 984-93. ©2016 AACR.
Asunto(s)
Antineoplásicos/administración & dosificación , Neoplasias Encefálicas/tratamiento farmacológico , Glioblastoma/tratamiento farmacológico , Indazoles/administración & dosificación , Péptidos y Proteínas de Señalización Intracelular/genética , Células Madre Neoplásicas/enzimología , Niacinamida/análogos & derivados , Proteínas Serina-Treonina Quinasas/genética , Animales , Antineoplásicos/farmacología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular , Glioblastoma/genética , Glioblastoma/patología , Humanos , Receptores de Hialuranos , Indazoles/farmacología , Ratones , Clasificación del Tumor , Niacinamida/administración & dosificación , Niacinamida/farmacología , Fosforilación/efectos de los fármacos , Análisis de Supervivencia , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
We provide evidence for a unique pathway engaged by the type II IFN receptor, involving mTORC2/AKT-mediated downstream regulation of mTORC1 and effectors. These events are required for formation of the eukaryotic translation initiation factor 4F complex (eIF4F) and initiation of mRNA translation of type II interferon-stimulated genes. Our studies establish that Rictor is essential for the generation of type II IFN-dependent antiviral and antiproliferative responses and that it controls the generation of type II IFN-suppressive effects on normal and malignant hematopoiesis. Together, our findings establish a central role for mTORC2 in IFNγ signaling and type II IFN responses.
Asunto(s)
Proteínas Portadoras/metabolismo , Factor 4F Eucariótico de Iniciación/metabolismo , Interferón gamma/metabolismo , Complejos Multiproteicos/metabolismo , Receptores de Interferón/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismo , Animales , Antivirales/química , Antivirales/metabolismo , Quimiocina CXCL10/metabolismo , Fibroblastos/metabolismo , Regulación de la Expresión Génica , Hematopoyesis , Células Madre Hematopoyéticas/citología , Humanos , Subunidad gamma del Factor 3 de Genes Estimulados por el Interferón/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina , Diana Mecanicista del Complejo 2 de la Rapamicina , Ratones , Ratones Noqueados , Fosforilación , Polirribosomas/metabolismo , Biosíntesis de Proteínas , Proteína Asociada al mTOR Insensible a la Rapamicina , Células U937RESUMEN
We investigated the efficacy of targeting the PIM kinase pathway in Philadelphia chromosome-positive (Ph+) leukemias. We provide evidence that inhibition of PIM, with the pan-PIM inhibitor SGI-1776, results in suppression of classic PIM effectors and also elements of the mTOR pathway, suggesting interplay between PIM and mTOR signals. Our data demonstrate that PIM inhibition enhances the effects of imatinib mesylate on Ph+ leukemia cells. We also found that PIM inhibition results in suppression of leukemic cell proliferation and induction of apoptosis of Ph+ leukemia cells, including those resistant to imatinib mesylate. Importantly, inhibition of PIM results in enhanced suppression of primary leukemic progenitors from patients with CML. Altogether these findings suggest that pharmacological PIM targeting may provide a unique therapeutic approach for the treatment of Ph+ leukemias.
Asunto(s)
Proteínas de Fusión bcr-abl/genética , Genes abl/genética , Imidazoles/farmacología , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-bcr/genética , Piridazinas/farmacología , Relación Dosis-Respuesta a Droga , Humanos , Mesilato de Imatinib/farmacología , Células K562 , Mutación , Cromosoma Filadelfia , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Células Tumorales CultivadasRESUMEN
We provide evidence that human SLFN5, an interferon (IFN)-inducible member of the Schlafen (SLFN) family of proteins, exhibits key roles in controlling motility and invasiveness of renal cell carcinoma (RCC) cells. Our studies define the mechanism by which this occurs, demonstrating that SLFN5 negatively controls expression of the matrix metalloproteinase 1 gene (MMP-1), MMP-13, and several other genes involved in the control of malignant cell motility. Importantly, our data establish that SLFN5 expression correlates with a better overall survival in a large cohort of patients with RCC. The inverse relationship between SLFN5 expression and RCC aggressiveness raises the possibility of developing unique therapeutic approaches in the treatment of RCC, by modulating SLFN5 expression.
Asunto(s)
Carcinoma de Células Renales/patología , Proteínas de Ciclo Celular/genética , Neoplasias Renales/patología , Metaloproteinasa 13 de la Matriz/biosíntesis , Metaloproteinasa 1 de la Matriz/biosíntesis , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/mortalidad , Proteínas de Ciclo Celular/biosíntesis , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular , Regulación Neoplásica de la Expresión Génica , Humanos , Interferón-alfa/farmacología , Neoplasias Renales/genética , Neoplasias Renales/mortalidad , Invasividad Neoplásica/genética , Interferencia de ARN , ARN Mensajero/biosíntesis , ARN Interferente PequeñoRESUMEN
We provide evidence that S6 kinase 1 (S6K1) Aly/REF-like target (SKAR) is engaged in IFN-α signaling and plays a key role in the generation of IFN responses. Our data demonstrate that IFN-α induces phosphorylation of SKAR, which is mediated by either the p90 ribosomal protein S6 kinase (RSK) or p70 S6 kinase (S6K1), in a cell type-specific manner. This type I IFN-inducible phosphorylation of SKAR results in enhanced interaction with the eukaryotic initiation factor (eIF)4G and recruitment of activated RSK1 to 5' cap mRNA. Our studies also establish that SKAR is present in cap-binding CBP80 immune complexes and that this interaction is mediated by eIF4G. We demonstrate that inducible protein expression of key IFN-α-regulated protein products such as ISG15 and p21(WAF1/CIP1) requires SKAR activity. Importantly, our studies define a requirement for SKAR in the generation of IFN-α-dependent inhibitory effects on malignant hematopoietic progenitors from patients with chronic myeloid leukemia or myeloproliferative neoplasms. Taken altogether, these findings establish critical and essential roles for SKAR in the regulation of mRNA translation of IFN-sensitive genes and induction of IFN-α biological responses.
Asunto(s)
Interferón-alfa/metabolismo , Proteínas Nucleares/metabolismo , Proteínas de Unión al ARN/metabolismo , Transducción de Señal , Animales , Antineoplásicos/farmacología , Línea Celular Tumoral , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Citocinas/metabolismo , Guanosina/análogos & derivados , Guanosina/metabolismo , Humanos , Ratones , Complejo Proteico Nuclear de Unión a la Caperuza/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Quinasas S6 Ribosómicas 90-kDa/metabolismo , Transducción de Señal/efectos de los fármacos , Ubiquitinas/metabolismoRESUMEN
Interferons (IFNs) are released by cells on exposure to various stimuli, including viruses, double-stranded RNA, and other cytokines and various polypeptides. These IFNs play important physiological and pathophysiological roles in humans. Many clinical studies have established activity for these cytokines in the treatment of several malignancies, viral syndromes, and autoimmune disorders. In this review, the regulatory effects of type I and II IFN receptors on the translation-initiation process mediated by mechanistic target of rapamycin (mTOR) and mitogen-activated protein kinase (MAPK) pathways and the known mechanisms of control of mRNA translation of IFN-stimulated genes are summarized and discussed.
Asunto(s)
Enfermedades Autoinmunes/inmunología , Interferón Tipo I/metabolismo , Interferón gamma/metabolismo , Neoplasias/inmunología , ARN Mensajero/metabolismo , Receptor de Interferón alfa y beta/metabolismo , Virosis/inmunología , Animales , Regulación de la Expresión Génica , Humanos , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Biosíntesis de Proteínas , ARN Mensajero/genética , Receptores de Interferón/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
We provide evidence that type I IFN-induced STAT activation is diminished in cells with targeted disruption of the Rictor gene, whose protein product is a key element of mTOR complex 2. Our studies show that transient or stable knockdown of Rictor or Sin1 results in defects in activation of elements of the STAT pathway and reduced STAT-DNA binding complexes. This leads to decreased expression of several IFN-inducible genes that mediate important biological functions. Our studies also demonstrate that Rictor and Sin1 play essential roles in the generation of the suppressive effects of IFNα on malignant erythroid precursors from patients with myeloproliferative neoplasms. Altogether, these findings provide evidence for critical functions for Rictor/Sin1 complexes in type I IFN signaling and the generation of type I IFN antineoplastic responses.
Asunto(s)
Antineoplásicos/farmacología , Proteínas Portadoras/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Interferón Tipo I/farmacología , Transcripción Genética/efectos de los fármacos , Animales , Proteínas Portadoras/genética , Células Cultivadas , Fibroblastos/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Células Madre Hematopoyéticas/efectos de los fármacos , Células Madre Hematopoyéticas/patología , Humanos , Ratones , Fosforilación , Policitemia Vera/metabolismo , Policitemia Vera/patología , Proteína Asociada al mTOR Insensible a la Rapamicina , Transducción de SeñalRESUMEN
Chronic myeloid leukemia (CML) and Ph+ acute lymphoblastic leukemia (ALL) are characterized by the presence of the BCR-ABL oncoprotein, which leads to activation of a plethora of pro-mitogenic and pro-survival pathways, including the mTOR signaling cascade. We provide evidence that in BCR-ABL expressing cells, treatment with tyrosine kinase inhibitors (TKIs) results in upregulation of mRNA levels and protein expression of sestrin3 (SESN3), a unique cellular inhibitor of mTOR complex 1 (mTORC1). Such upregulation appears to be mediated by regulatory effects on mTOR, as catalytic inhibition of the mTOR kinase also induces SESN3. Catalytic mTOR inhibition also results in upregulation of SESN3 expression in cells harboring the TKI-insensitive T315I-BCR-ABL mutant, which is resistant to imatinib mesylate. Overexpression of SESN3 results in inhibitory effects on different Ph+ leukemic cell lines including KT-1-derived leukemic precursors, indicating that SESN3 mediates anti-leukemic responses in Ph+ cells. Altogether, our findings suggest the existence of a novel mechanism for the generation of antileukemic responses in CML cells, involving upregulation of SESN3 expression.
Asunto(s)
Proteínas de Fusión bcr-abl/biosíntesis , Regulación Leucémica de la Expresión Génica , Proteínas de Choque Térmico/biosíntesis , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Animales , Línea Celular Tumoral , Proteínas de Fusión bcr-abl/genética , Proteínas de Choque Térmico/genética , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Complejos Multiproteicos/antagonistas & inhibidores , Complejos Multiproteicos/genética , Complejos Multiproteicos/metabolismo , Serina-Treonina Quinasas TOR/antagonistas & inhibidores , Serina-Treonina Quinasas TOR/genética , Serina-Treonina Quinasas TOR/metabolismo , Regulación hacia Arriba/genéticaRESUMEN
There is emerging evidence that the IFN-inducible family of Slfn genes and proteins play important roles in cell cycle progression and control of cellular proliferation, but the precise functional roles of different Slfn members in the regulation of tumorigenesis remain unclear. In the present study, we undertook a systematic analysis on the expression and functional relevance of different mouse Slfn genes in malignant melanoma and renal cell carcinoma cells. Our studies demonstrate that several mouse Slfn genes are up-regulated in response to IFN treatment of mouse melanoma and renal cell carcinoma cells, including Slfn1, Slfn2, Slfn4, Slfn5, and Slfn8. Our data show that Slfn2 and Slfn3 play essential roles in the control of mouse malignant melanoma cell proliferation and/or anchorage-independent growth, suggesting key and non-overlapping roles for these genes in the control of malignant melanoma tumorigenesis. In renal cell carcinoma cells, in addition to Slfn2 and Slfn3, Slfn5 also exhibits important antineoplastic effects. Altogether, our findings indicate important functions for distinct mouse Slfn genes in the control of tumorigenesis and provide evidence for differential involvement of distinct members of this gene family in controlling tumorigenesis. They also raise the potential of future therapeutic approaches involving modulation of expression of members of this family of genes in malignant melanoma and renal cell carcinoma.
Asunto(s)
Carcinoma de Células Renales/metabolismo , Proteínas de Ciclo Celular/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias Renales/metabolismo , Melanoma/metabolismo , Proteínas de Neoplasias/metabolismo , Animales , Antivirales/farmacología , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/patología , Proteínas de Ciclo Celular/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Interferón gamma/farmacología , Neoplasias Renales/genética , Neoplasias Renales/patología , Melanoma/genética , Melanoma/patología , Ratones , Proteínas de Neoplasias/genética , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genéticaRESUMEN
Alternative splicing (AS) is an efficient mechanism that involves the generation of transcriptome and protein diversity from a single gene. Defects in pre-messenger RNA (mRNA) splicing are an important cause of numerous diseases, including cancer. AS of pre-mRNA as a target for cancer therapy has not been well studied. We have reported previously that a splicing factor, polypyrimidine tract-binding protein (PTB), is overexpressed in ovarian tumors compared with matched normal controls, and knockdown of PTB expression by short-hairpin RNA impairs ovarian tumor cell growth, colony formation, and invasiveness. Given the complexity of PTB's molecular functions, a chemical method for controlling PTB activity might provide a therapeutic and experimental tool. However, no commercially available PTB inhibitors have yet been described. To expand our ability to find novel inhibitors, we developed a robust, fluorometric, cell-based high-throughput screening assay in 96-well plates that reports on the splicing activity of PTB. In an attempt to use the cells for large-scale chemical screens to identify PTB modulators, we established cell lines stably expressing the reporter gene. Our results suggest that this high-throughput assay could be used to identify small-molecule modulators of PTB activity. Based on these findings and the role that upregulated PTB has on cell proliferation and malignant properties of tumors, targeting PTB for inhibition with small molecules offers a promising strategy for cancer therapy.
Asunto(s)
Empalme Alternativo , Ensayos Analíticos de Alto Rendimiento/métodos , Bibliotecas de Moléculas Pequeñas , Empalme Alternativo/efectos de los fármacos , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Regulación de la Expresión Génica/efectos de los fármacos , Genes Reporteros , Humanos , Proteínas del Tejido Nervioso/genética , Proteína de Unión al Tracto de Polipirimidina/genética , Proteína de Unión al Tracto de Polipirimidina/metabolismo , Precursores del ARN/genética , Reproducibilidad de los ResultadosRESUMEN
Nuclear factor (NF)-YB, a subunit of the transcription factor nuclear factor Y (NF-Y) complex, binds and activates CCAAT-containing promoters. Our previous work suggested that NF-YB may be a mediator of topoisomerase IIα (Top2α), working through the Top2α promoter. DNA topoisomerase II (Top2) is an essential nuclear enzyme and the primary target for several clinically important anticancer drugs. Our teniposide-resistant human lymphoblastic leukemia CEM cells (CEM/VM-1-5) express reduced Top2α protein compared with parental CEM cells. To study the regulation of Top2α during the development of drug resistance, we found that NF-YB protein expression is increased in CEM/VM-1-5 cells compared with parental CEM cells. This further suggests that increased NF-YB may be a negative regulator of Top2α in CEM/VM-1-5 cells. We asked what causes the up-regulation of NF-YB in CEM/VM-1-5 cells. We found by microRNA profiling that hsa-miR-485-3p is lower in CEM/VM-1-5 cells compared with CEM cells. MicroRNA target prediction programs revealed that the 3'-untranslated region (3'-UTR) of NF-YB harbors a putative hsa-miR-485-3p binding site. We thus hypothesized that hsa-miR-485-3p mediates drug responsiveness by decreasing NF-YB expression, which in turn negatively regulates Top2α expression. To test this, we overexpressed miR-485-3p in CEM/VM-1-5 cells and found that this led to reduced expression of NF-YB, a corresponding up-regulation of Top2α, and increased sensitivity to the Top2 inhibitors. Results in CEM cells were replicated in drug-sensitive and -resistant human rhabdomyosarcoma Rh30 cells, suggesting that our findings represent a general phenomenon. Ours is the first study to show that miR-485-3p mediates Top2α down-regulation in part by altered regulation of NF-YB.
Asunto(s)
Antígenos de Neoplasias/biosíntesis , Factor de Unión a CCAAT/metabolismo , ADN-Topoisomerasas de Tipo II/biosíntesis , Proteínas de Unión al ADN/biosíntesis , Etopósido/toxicidad , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , MicroARNs/fisiología , Antígenos de Neoplasias/genética , Antineoplásicos/toxicidad , Línea Celular Tumoral , ADN-Topoisomerasas de Tipo II/genética , Proteínas de Unión al ADN/antagonistas & inhibidores , Proteínas de Unión al ADN/genética , Regulación hacia Abajo/efectos de los fármacos , Resistencia a Antineoplásicos , Regulación Enzimológica de la Expresión Génica/fisiología , Humanos , Proteínas de Unión a Poli-ADP-Ribosa , Tenipósido/toxicidad , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Through inhibitory G protein-coupled melatonin receptors, melatonin regulates intracellular signaling systems and also the transcriptional activity of certain genes. Clock genes are proposed as regulatory factors in forming dopamine-related behaviors and mood and melatonin has the ability to regulate these processes. Melatonin-mediated changes in clock gene expression have been reported in brain regions, including the striatum, that are crucial for the development of dopaminergic behaviors and mood. However, it is not known whether melatonin receptors present in striatum mediate these effects. Therefore, we investigated the role of the melatonin/melatonin receptor system on clock gene expression using a model of primary neuronal cultures prepared from striatum. We found that melatonin at the receptor affinity range (i.e., nm) affects the expression of the clock genes mPer1, mClock, mBmal1 and mNPAS2 (neuronal PAS domain protein 2) differentially in a pertussis toxin-sensitive manner: a decrease in Per1 and Clock, an increase in NPAS2 and no change in Bmal1 expression. Furthermore, mutating MT1 melatonin receptor (i.e., MT1 knockouts, MT1(-/-)) reversed melatonin-induced changes, indicating the involvement of MT1 receptor in the regulatory action of melatonin on neuronal clock gene expression. Therefore, by controlling clock gene expression we propose melatonin receptors (i.e., MT1) as novel therapeutic targets for the pathobiologies of dopamine-related behaviors and mood.