RESUMEN
The hypothalamus has a vital role in controlling food intake and energy homeostasis; its activity is modulated by neuropeptides and endocrine factors. Mesencephalic astrocyte-derived neurotrophic factor (MANF) is a neurotrophic factor that is also localized in the endoplasmic reticulum (ER) in neurons. Here we show that MANF is highly enriched in distinct nuclei of the mouse hypothalamus, and that MANF expression in the hypothalamus is upregulated in response to fasting. Increasing or decreasing hypothalamic MANF protein levels causes hyperphagia or hypophagia, respectively. Moreover, MANF triggers hypothalamic insulin resistance by enhancing the ER localization and activity of PIP4k2b, a kinase known to regulate insulin signaling. Our findings indicate that MANF influences food intake and body weight by modulating hypothalamic insulin signaling.MANF is a neurotrophic factor that is secreted but also mediates the unfolded protein response acting intracellularly. Here, the authors show that MANF expression in the brain is influenced by nutritional cues, and hypothalamic MANF influences food intake and systemic energy homeostasis.
Asunto(s)
Peso Corporal/fisiología , Ingestión de Alimentos/fisiología , Hipotálamo/metabolismo , Factores de Crecimiento Nervioso/metabolismo , Animales , Western Blotting , Peso Corporal/genética , Ingestión de Alimentos/genética , Estrés del Retículo Endoplásmico/genética , Femenino , Expresión Génica , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Factores de Crecimiento Nervioso/genética , Células PC12 , Interferencia de ARN , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Huntingtin-associated protein 1 (Hap1) is known to be critical for postnatal hypothalamic function and growth. Hap1 forms stigmoid bodies (SBs), unique neuronal cytoplasmic inclusions of unknown function that are enriched in hypothalamic neurons. Here we developed a simple strategy to isolate the SB-enriched fraction from mouse brain. By analyzing Hap1 immunoprecipitants from this fraction, we identified a Hap1-interacting SB component, DDB1 and CUL4 associated factor 7 (Dcaf7)/WD40 repeat 68 (WDR68), whose protein level and nuclear translocation are regulated by Hap1. Moreover, we found that Hap1 bound Dcaf7 competitively in cytoplasm with dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A), a protein implicated in Down syndrome (DS). Depleting Hap1 promoted the DYRK1A-Dcaf7 interaction and increased the DYRK1A protein level. Transgenic DS mice overexpressing DYRK1A showed reduced Hap1-Dcaf7 association in the hypothalamus. Furthermore, the overexpression of DYRK1A in the hypothalamus led to delayed growth in postnatal mice, suggesting that DYRK1A regulates the Hap1-Dcaf7 interaction and postnatal growth and that targeting Hap1 or Dcaf7 could ameliorate growth retardation in DS.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Síndrome de Down/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Transporte Activo de Núcleo Celular , Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Núcleo Celular/metabolismo , Síndrome de Down/genética , Células HEK293 , Humanos , Hipotálamo/metabolismo , Cuerpos de Inclusión/metabolismo , Ratones Noqueados , Ratones Transgénicos , Proteínas del Tejido Nervioso/genética , Neuronas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Tirosina Quinasas/genética , Interferencia de ARN , Quinasas DyrKRESUMEN
Lipidomic analyses of the frontal cortex of Rhesus macaques across three selected age groups (young, sexually-mature, old) revealed that docosahexaenoic acids (DHAs) displayed notable and unique accretions in sexually-mature macaques for all phospholipid classes examined, which were not observable in all remaining polyunsaturated fatty acids (PUFAs) investigated. On the other hand, arachidonic acid (ARA) exhibited sharp attritions in the membrane lipidomes of sexually-mature macaques, a decline which was attenuated only for cardiolipins (CLs). DHA enrichment in phospholipids was lost in old macaques, with accompanying augmentations in very-long-chain sphingomyelins (VLC-SMs). Age-dependent alterations in membrane lipidomes point to a possibly complex temporal interplay between DHA-enriched membrane microdomains and SM-/cholesterol-rich rafts in neural membranes during normative aging. Lipid co-regulation data revealed an increasingly intense degree of co-regulation between membrane lipid classes with age, and suggest that reduction in CLs during normative brain aging may prompt alternative membrane lipid synthetic pathways driven by a compromised energy availability in the aging brain.
Asunto(s)
Envejecimiento/fisiología , Ácido Araquidónico/metabolismo , Ácidos Docosahexaenoicos/metabolismo , Lóbulo Frontal/metabolismo , Microdominios de Membrana/fisiología , Animales , Cardiolipinas/metabolismo , Colesterol/fisiología , Humanos , Macaca mulatta , Masculino , Modelos Animales , Neuronas/metabolismo , Esfingomielinas/metabolismoRESUMEN
Huntington disease (HD) represents a family of neurodegenerative diseases that are caused by misfolded proteins. The misfolded proteins accumulate in the affected brain regions in an age-dependent manner to cause late-onset neurodegeneration. Transgenic mouse models expressing the HD protein, huntingtin, have been widely used to identify therapeutics that may retard disease progression. Here we report that Berberine (BBR), an organic small molecule isolated from plants, has protective effects on transgenic HD (N171-82Q) mice. We found that BBR can reduce the accumulation of mutant huntingtin in cultured cells. More importantly, when given orally, BBR could effectively alleviate motor dysfunction and prolong the survival of transgenic N171-82Q HD mice. We found that BBR could promote the degradation of mutant huntingtin by enhancing autophagic function. Since BBR is an orally-taken drug that has been safely used to treat a number of diseases, our findings suggest that BBR can be tested on different HD animal models and HD patients to further evaluate its therapeutic effects.
Asunto(s)
Berberina/uso terapéutico , Modelos Animales de Enfermedad , Enfermedad de Huntington/tratamiento farmacológico , Animales , Autofagia/efectos de los fármacos , Conducta Animal , Células HEK293 , Humanos , Ratones , Ratones TransgénicosRESUMEN
Defective neurogenesis in the postnatal brain can lead to many neurological and psychiatric disorders, yet the mechanism behind postnatal neurogenesis remains to be investigated. Huntingtin-associated protein 1 (HAP1) participates in intracellular trafficking in neurons, and its absence leads to postnatal death in mice. Here, we used tamoxifen-induced (TM-induced) Cre recombination to deplete HAP1 in mice at different ages. We found that HAP1 reduction selectively affects survival and growth of postnatal mice, but not adults. Neurogenesis, but not gliogenesis, was affected in HAP1-null neurospheres and mouse brain. In the absence of HAP1, postnatal hypothalamic neurons exhibited reduced receptor tropomyosin-related kinase B (TRKB) levels and decreased survival. HAP1 stabilized the association of TRKB with the intracellular sorting protein sortilin, prevented TRKB degradation, and promoted its anterograde transport. Our findings indicate that intracellular sorting of neurotrophin receptors is critical for postnatal neurogenesis and could provide a therapeutic target for defective postnatal neurogenesis.
Asunto(s)
Proteínas del Tejido Nervioso/metabolismo , Neurogénesis , Receptor trkB/metabolismo , Receptores de Factor de Crecimiento Nervioso/metabolismo , Animales , Células Cultivadas , Corteza Cerebral/metabolismo , Regulación del Desarrollo de la Expresión Génica , Células HEK293 , Humanos , Hipotálamo/citología , Hipotálamo/metabolismo , Sistema Límbico/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas del Tejido Nervioso/genética , Células-Madre Neurales/fisiología , Especificidad de Órganos , Estabilidad Proteica , Transporte de Proteínas , Proteolisis , Esferoides Celulares/fisiologíaRESUMEN
OBJECTIVE: To study the leptin resistance mechanism of Xiaoyan Decoction (XD) in lung cancer cachexia (LCC) rats. METHODS: An LCC rat model was established. Totally 40 rats were randomly divided into the normal control group, the LCC model group, the XD group, and the positive control group, 10 in each group. After LCC model was set up, rats in the LCC model group were administered with normal saline, 2 mL each time. Rats in the XD group were administered with XD at the daily dose of 2 mL. Those in the positive control group were administered with Medroxyprogesterone Acetate suspension (20 mg/kg) by gastrogavage at the daily dose of 2 mL. All medication lasted for 14 days. The general condition and tumor growth were observed. Serum levels of leptin and leptin receptor in the hypothalamus were detected using enzyme-linked immunosorbent assay. Contents of neuropeptide Y (NPY) and anorexia for genomic POMC were detected using real-time PCR technique. RESULTS: Serum leptin levels were lower in the LCC model group than in the normal control group with statistical significance (P < 0.05). Compared with the LCC model groups, serum leptin levels significantly increased in the XD group (P < 0.01). Leptin receptor levels in the hypothalamus increased significantly in the LCC model group (P < 0.01). Increased receptor levels in the LCC model group indicated that either XD or Medroxyprogesterone Acetate could effectively reduce levels of leptin receptor with statistical significance (P < 0.01). There was also statistical difference between the XD group and the positive control group (P < 0.05). Contents of NPY was higher in the LCC model group than in the other groups with statistical difference (P < 0.05). There was no statistical difference in NPY between the normal control group and the rest 2 treatment groups (P > 0.05). There was statistical difference in POMC between the normal control group and the LCC model group (P < 0.05). POMC could be decreased in the XD group and the positive control group with statistical significance (P < 0.05), and it was more obviously decreased in the XD group (P < 0.05). CONCLUSIONS: Leptin resistance existed in LCC rats. XD could increase serum leptin levels and reduce leptin receptor levels in the hypothalamus. LCC could be improved by elevating NPY contents in the hypothalamus and reducing POMC contents, promoting the appetite, and increasing food intake from the periphery pathway and the central pathway.
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Caquexia/tratamiento farmacológico , Medicamentos Herbarios Chinos/uso terapéutico , Neoplasias Pulmonares/complicaciones , Animales , Caquexia/etiología , Ingestión de Alimentos , Humanos , Hipotálamo/metabolismo , Leptina/metabolismo , Neuropéptido Y/metabolismo , Distribución Aleatoria , Ratas , Ratas Sprague-DawleyRESUMEN
OBJECTIVE: To observe the clinical efficacy of yiqi jiedu quyu recipe (YJQR) combined endocrine therapy in treating advanced prostate cancer (APC). METHODS: Totally 44 APC patients were randomly assigned to the combination group (23 cases) and the control group (21 cases). All patients received endocrine therapy for treating PC in Guidelines for Diagnosis and Treatment of Urological Diseases in China. After 6 months of treatment serum prostate specific antigen (PSA), free PSA (f-PSA), hemoglobin (Hb), Tcell subsets, natural kill (NK) cells, QLQ-C30 scale, Chinese medical symptoms score, I -PSS score, as well as the incidence of tidal fever were observed. RESULTS: After treatment the PSA, f-PSA, and Hb were improved in the combination group, showing statistical difference when compared with before treatment (P <0.05). Besides, better effects were obtained in the combination group (P <0.05, P <0.01). After treatment CD3, CD4, CD8, CD4/CD8, and NK cell levels were also improved in the combination group, showing statistical difference when compared with before treatment (P <0.05). Besides, better effects were obtained in the combination group (P <0.01). The quality of life (QOL), emotional function, Chinese medical symptoms score, and I -PSS symptom score were improved in the two groups (P <0. 05,P <0. 01). Besides, better effects in the aforesaid indices and the incidence of tidal fever were obtained in the combination group (P <0.05). CONCLUSION: YJQR combined endocrine therapy could obviously improve APC patients' QOL, reduce adverse reactions of Western medical treatment, improve the immune function, and enhance therapeutic effects of endocrine therapy.
Asunto(s)
Medicamentos Herbarios Chinos/uso terapéutico , Fitoterapia , Neoplasias de la Próstata/terapia , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Humanos , Masculino , Persona de Mediana Edad , Neoplasias de la Próstata/patología , Resultado del Tratamiento , Adulto JovenRESUMEN
Mutations in the Abelson helper integration site-1 (AHI1) gene result in N-terminal Ahi1 fragments and cause Joubert syndrome, an autosomal recessive brain malformation disorder associated with delayed development. How AHI1 mutations lead to delayed development remains unclear. Here we report that full-length, but not N-terminal, Ahi1 binds Hap1, a huntingtin-associated protein that is essential for the postnatal survival of mice and that this binding is regulated during neuronal differentiation by nerve growth factor. Nerve growth factor induces dephosphorylation of Hap1A and decreases its association with Ahi1, correlating with increased Hap1A distribution in neurite tips. Consistently, Ahi1 associates with phosphorylated Hap1A in cytosolic, but not in synaptosomal, fractions isolated from mouse brain, suggesting that Ahi1 functions mainly in the soma of neurons. Mass spectrometry analysis of cytosolic Ahi1 immunoprecipitates reveals that Ahi1 also binds Cend1 (cell cycle exit and neuronal differentiation protein 1)/BM88, a neuronal protein that mediates neuronal differentiation and is highly expressed in postnatal mouse brain. Loss of Ahi1 reduces the levels of Cend1 in the hypothalamus of Ahi1 KO mice, which show retarded growth during postnatal days. Overexpressed Ahi1 can stabilize Cend1 in cultured cells. Furthermore, overexpression of Cend1 can rescue the neurite extension defects of hypothalamic neurons from Ahi1 KO mice. Our findings suggest that Cend1 is involved in Ahi1-associated hypothalamic neuronal differentiation in early development, giving us fresh insight into the mechanism behind the delayed development in Joubert syndrome.
Asunto(s)
Diferenciación Celular/fisiología , Regulación de la Expresión Génica/genética , Proteínas de la Membrana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neuronas/fisiología , Proteínas Proto-Oncogénicas/deficiencia , Proteínas Adaptadoras del Transporte Vesicular , Factores de Edad , Animales , Animales Recién Nacidos , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Regulación de la Expresión Génica/efectos de los fármacos , Suspensión Trasera/fisiología , Humanos , Hipotálamo/citología , Hipotálamo/crecimiento & desarrollo , Inmunoprecipitación , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Actividad Motora/efectos de los fármacos , Actividad Motora/genética , Mutación/genética , Factor de Crecimiento Nervioso/farmacología , Neuritas/efectos de los fármacos , Neuritas/fisiología , Neuronas/ultraestructura , Fosforilación/efectos de los fármacos , Ratas , Natación , TransfecciónRESUMEN
It is indicated that there are important molecules interacting with brain nervous systems to regulate feeding and energy balance by influencing the signaling pathways of these systems, but relatively few of the critical players have been identified. In the present study, we provide the evidence for the role of Abelson helper integration site 1 (Ahi1) protein as a mediator of feeding behavior through interaction with serotonin receptor 2C (5-HT(2C)R), known for its critical role in feeding and appetite control. First, we demonstrated the co-localization and interaction between hypothalamic Ahi1 and 5-HT(2C)R. Ahi1 promoted the degradation of 5-HT(2C)R through the lysosomal pathway. Then, we investigated the effects of fasting on the expression of hypothalamic Ahi1 and 5-HT(2C)R. Fasting resulted in an increased Ahi1 expression and a concomitant decreased expression of 5-HT(2C)R. Knockdown of hypothalamic Ahi1 led to a concomitant increased expression of 5-HT(2C)R and a decrease of food intake and body weight. Last, we found that Ahi1 could regulate the expression of neuropeptide Y and proopiomelanocortin. Taken together, our results indicate that Ahi1 mediates feeding behavior by interacting with 5-HT(2C)R to modulate the serotonin signaling pathway.
Asunto(s)
Regulación del Apetito/fisiología , Conducta Alimentaria/fisiología , Hipotálamo/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Proteolisis , Proteínas Proto-Oncogénicas/metabolismo , Receptor de Serotonina 5-HT2C/metabolismo , Proteínas Adaptadoras del Transporte Vesicular , Animales , Humanos , Masculino , Ratones , Neuropéptido Y/biosíntesis , Serotonina/metabolismo , Transducción de Señal/fisiologíaRESUMEN
OBJECTIVE: To study the therapeutic effect and mechanism of thermotherapy combined with thoracic injection of lentinan in treatment of cancerous hydrothorax (CH) in patients with lung cancer. METHODS: Sixty lung cancer patients complicated with CH were randomly assigned to the observation group and the control group, 30 in each. CH was released by closed drainage of the thoracic cavity in all patients. Thermotherapy was given to patients in the observation group after lentinan was thoracically injected, while lentinan was thoracically injected to patients in the control group. RESULTS: The total effective rate of CH improvement was 73.3% (22/30) in the control group and 83.3% (25/30) in the observation group, showing insignificant difference (P>0.05). The stability rate and the weight stability rate by Karnofsky's performance scoring in the observation group were superior to those of the control group, showing significant difference (P<0.05). No hematological reaction, hepatic or renal damage occurred before and after treatment in both groups. Fever, thoracalgia, dyspnea, rash, nausea and vomit appeared in few patients of the two groups, showing insignificant difference (P>0.05). CD3+, CD4+, CD4+/CD8+, and NK obviously increased and CD8+ decreased in the observation group after treatment, showing significant difference from those before treatment (P<0.05). Compared with the control group after treatment, CD3+, CD4+, CD4+/CD8+, and NK obviously increased, CD8+ increased in the observation group (P<0.05). CONCLUSION: Thermotherapy combined with thoracic injection of lentinan showed better effect in treatment of CH in patients with lung cancer. No obvious adverse reaction was seen.
Asunto(s)
Productos Biológicos/uso terapéutico , Hidrotórax/terapia , Hipertermia Inducida , Lentinano/uso terapéutico , Neoplasias Pulmonares/terapia , Adulto , Anciano , Terapia Combinada , Femenino , Humanos , Hidrotórax/etiología , Inyecciones , Masculino , Medicina Tradicional China/métodos , Persona de Mediana EdadRESUMEN
Recent studies suggest that the human Abelson helper integration site-1 (AHI1) gene on chromosome 6 is associated with susceptibility to schizophrenia and autism, two common neuropsychological disorders with depression symptoms. Mouse Ahi1 protein is abundant in the hypothalamus and amygdala, which are important brain regions for controlling emotion. However, the neuronal function of Ahi1 remains unclear. With the Cre-loxP system, we created a mouse model that selectively reduces Ahi1 expression in neuronal cells. Mice with neuronal Ahi1 deficiency show reduced TrkB level in the brain and depressive phenotypes, which can be alleviated by antidepressant drugs or by overexpression of TrkB in the amygdala. Ahi1 deficiency promotes the degradation of endocytic TrkB and reduces TrkB signaling in neuronal cells. Our findings suggest that impaired endocytic sorting and increased degradation of TrkB can induce depression and that this impaired pathway may serve as a previously uncharacterized therapeutic target for depression.
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Amígdala del Cerebelo/metabolismo , Depresión/metabolismo , Hipotálamo/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Transducción de Señal , Proteínas Adaptadoras del Transporte Vesicular , Animales , Depresión/genética , Depresión/terapia , Modelos Animales de Enfermedad , Endocitosis/genética , Humanos , Glicoproteínas de Membrana/genética , Ratones , Ratones Transgénicos , Fenotipo , Proteínas Tirosina Quinasas/genética , Proteínas Proto-Oncogénicas/genéticaRESUMEN
Huntingtin-associated protein-1 (Hap1) is a neuronal protein that associates with huntingtin, the Huntington disease protein. Although Hap1 and huntingtin are known to be involved in intracellular trafficking, whether and how the impairment of Hap1-associated trafficking leads to neurological pathology and symptoms remain to be seen. As Hap1 is enriched in neuronal cells in the brain, addressing this issue is important in defining the role of defective intracellular trafficking in the selective neuropathology associated with Hap1 dysfunction. Here, we find that Hap1 is abundantly expressed in orexin (hypocretin)-producing neurons (orexin neurons), which are distinctly distributed in the hypothalamus and play an important role in the regulation of feeding and behavior. We created conditional Hap1 knock-out mice to selectively deplete Hap1 in orexin neurons via the Cre-loxP system. These mice show process fragmentation of orexin neurons and reductions in food intake, body weight, and locomotor activity. Sucrose density gradient fractionation reveals that loss of Hap1 in the mouse brain also reduces the distribution of trafficking protein complexes and cargo proteins in the fractions that are enriched in synaptosomes. These results suggest that Hap1 is critical for the transport of multiple proteins to the nerve terminals to maintain the integrity of neuronal processes and that selective disruption of the processes of orexin neurons can cause abnormal feeding and locomotor activity.
Asunto(s)
Ingestión de Alimentos/fisiología , Conducta Alimentaria/fisiología , Hipotálamo/metabolismo , Actividad Motora/fisiología , Proteínas del Tejido Nervioso/metabolismo , Neuronas/metabolismo , Neuropéptidos/biosíntesis , Animales , Transporte Biológico/fisiología , Humanos , Proteína Huntingtina , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Péptidos y Proteínas de Señalización Intracelular , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/genética , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Orexinas , Sinaptosomas/metabolismoRESUMEN
OBJECTIVE: To observe the clinical therapeutic effect of Airening Suppository (ARN) on carcinoma fever. METHODS: Adopting randomized and double-blinded method, 54 patients with carcinoma fever were randomly assigned to the observation group and the control group, who were treated respectively with ARN and indomethacin suppository by anal administering, 7 days as one treatment course. The effect on body temperature on patients and side effect occurred were observed 7 days after discontinuation of the medication, and patients' serum tumor necrosis factor alpha (TNF-alpha), interleukin-1beta were determined by RIA, routine test of blood, urine and stool, liver and renal function were tested as well. Body temperature, changes of clinical symptoms, and adverse reaction were observed. RESULTS: The effective rate of fever abating was 86.7% in the observation group and 66.7% in the control group, showing significant difference between them (chi2 = 8.06, P <0.05). After treatment, serum levels of TNF-alpha and IL-1beta in the observation group were much lower than those in the control group (t = 7.477, t = 3.492, P <0.01). Besides, patients' chief clinical symptoms and quality of life were markedly improved in the observation group (t =4.71, P <0.05, chi2 =7.38, P <0.05), all better than those in the control group (P < 0.05). No evident effect on peripheral blood figure, or harmful impact on heart, liver and kidney function was found. CONCLUSION: Airening Suppository has a standing and stable fever abating effect in treating cancerous fever with no obvious adverse reaction, so it is worthy of clinical use and spreading.
Asunto(s)
Medicamentos Herbarios Chinos/uso terapéutico , Fiebre/tratamiento farmacológico , Neoplasias/complicaciones , Adulto , Anciano , Femenino , Fiebre/etiología , Humanos , Masculino , Persona de Mediana Edad , Supositorios , Adulto JovenRESUMEN
Mutant huntingtin can affect vesicular and receptor trafficking via its abnormal protein interactions, suggesting that impairment of intracellular trafficking may contribute to Huntington's disease. There is growing evidence that huntingtin-associated protein-1 (HAP1) also interacts with microtubule-dependent transporters and is involved in intracellular trafficking. However, it remains unclear how the trafficking of HAP1 is regulated and contributes to neuronal function. Here we report that phosphorylation of HAP1 decreases its association with microtubule-dependent transport proteins dynactin p150 and kinesin light chain and reduces its localization in neurite tips. Suppressing HAP1 expression by RNA interference reduces neurite outgrowth and the level of tropomyosin-related kinase A receptor tyrosine kinase (TrkA), a nerve growth factor receptor whose internalization and trafficking are required for neurite outgrowth. HAP1 maintains the normal level of membrane TrkA by preventing the degradation of internalized TrkA. Mutant huntingtin also reduces the association of HAP1 with dynactin p150 and kinesin light chain and thereby decreases the intracellular level of TrkA. These findings suggest that HAP1 trafficking is critical for the stability of TrkA and neurite function, both of which can be attenuated by mutant huntingtin.
Asunto(s)
Proteínas del Tejido Nervioso/metabolismo , Neuritas/fisiología , Neuronas/fisiología , Receptor trkA/metabolismo , Animales , Secuencia de Bases , Cartilla de ADN , Genes Reporteros , Enfermedad de Huntington , Hipotálamo , Microscopía Confocal , Neuritas/ultraestructura , Neuronas/citología , Células PC12 , Feocromocitoma , ARN Interferente Pequeño/genética , Ratas , Ganglio Cervical Superior/fisiologíaRESUMEN
The hypothalamus responds to circulating leptin and insulin in the control of food intake and body weight. A number of neurotransmitters in the hypothalamus, including gamma-aminobutyric acid (GABA), also have key roles in feeding. Huntingtin-associated protein 1 (Hap1) is expressed more abundantly in the hypothalamus than in other brain regions, and lack of Hap1 in mice leads to early postnatal death. Hap1 is also involved in intracellular trafficking of the GABA(A) receptor. Here, we report that fasting upregulates the expression of Hap1 in the rodent hypothalamus, whereas intracerebroventricular administration of insulin downregulates Hap1 by increasing its degradation through ubiquitination. Decreasing the expression of mouse hypothalamic Hap1 by siRNA reduces the level and activity of hypothalamic GABA(A) receptors and causes a decrease in food intake and body weight. These findings provide evidence linking hypothalamic Hap1 to GABA in the stimulation of feeding and suggest that this mechanism is involved in the feeding-inhibitory actions of insulin in the brain.
Asunto(s)
Ingestión de Alimentos , Conducta Alimentaria/fisiología , Hipotálamo/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Receptores de GABA-A/metabolismo , Animales , Peso Corporal , Electrofisiología , Ayuno , Humanos , Hipotálamo/citología , Insulina/metabolismo , Leptina/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas del Tejido Nervioso/genética , Neuronas/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Ratas , Ratas Sprague-Dawley , Ubiquitina/metabolismoRESUMEN
Huntingtin-associated protein-1 (HAP1) was initially identified as an interacting partner of huntingtin, the Huntington disease protein. Unlike huntingtin that is ubiquitously expressed throughout the brain and body, HAP1 is enriched in neurons, suggesting that its dysfunction could contribute to Huntington disease neuropathology. Growing evidence has demonstrated that HAP1 and huntingtin are anterogradely transported in axons and that the abnormal interaction between mutant huntingtin and HAP1 may impair axonal transport. However, the exact role of HAP1 in anterograde transport remains unclear. Here we report that HAP1 interacts with kinesin light chain, a subunit of the kinesin motor complex that drives anterograde transport along microtubules in neuronal processes. The interaction of HAP1 with kinesin light chain is demonstrated via a yeast two-hybrid assay, glutathione S-transferase pull down, and coimmunoprecipitation. Furthermore, HAP1 is colocalized with kinesin in growth cones of neuronal cells. We also demonstrated that knocking down HAP1 via small interfering RNA suppresses neurite outgrowth of PC12 cells. Analysis of live neuronal cells with fluorescence microscopy and fluorescence recovery after photobleaching demonstrates that suppressing the expression of HAP1 or deleting the HAP1 gene inhibits the kinesin-dependent transport of amyloid precursor protein vesicles. These studies provide a molecular basis for the participation of HAP1 in anterograde transport in neuronal cells.
Asunto(s)
Cinesinas/química , Proteínas del Tejido Nervioso/metabolismo , Precursor de Proteína beta-Amiloide/química , Animales , Axones/metabolismo , Transporte Biológico , Western Blotting , Glutatión Transferasa/metabolismo , Hipotálamo/metabolismo , Procesamiento de Imagen Asistido por Computador , Microscopía Fluorescente , Microtúbulos/metabolismo , Modelos Estadísticos , Neuronas/metabolismo , Células PC12 , Unión Proteica , Isoformas de Proteínas , Estructura Terciaria de Proteína , ARN Interferente Pequeño/metabolismo , Ratas , Transfección , Técnicas del Sistema de Dos HíbridosRESUMEN
Huntington's disease (HD) is caused by a polyglutamine expansion in the disease protein huntingtin. The polyglutamine expansion causes huntingtin to interact abnormally with a number of proteins. However, it is unclear whether, and how, huntingtin-associated proteins are involved in the neurodegeneration in HD. Here, we show that huntingtin-associated protein-1 (HAP1), which is involved in intracellular trafficking of epidermal growth factor receptor (EGFR), is highly expressed in the hypothalamus. Mice lacking HAP1 die after birth because of depressed feeding activity. Terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling staining and electron microscopic examination revealed the degeneration in hypothalamic regions that control feeding behavior. Hypothalamic degeneration was also observed in HD transgenic mice that have a significant loss of body weight. Inhibition of HAP1 expression decreases EGFR signaling and cell viability, whereas overexpression of HAP1 enhances this signaling activity and inhibits mutant huntingtin-mediated cytotoxicity. These results suggest that the effect of mutant huntingtin on HAP1 and EGFR signaling may contribute to the hypothalamic neurodegeneration and loss of body weight in HD.