RESUMO
Homer is a postsynaptic scaffold protein, which has long and short isoforms. The long form of Homer consists of an N-terminal target-binding domain and a C-terminal multimerization domain, linking multiple proteins within a complex. The short form of Homer only has the N-terminal domain and likely acts as a dominant negative regulator. Homer2a, one of the long form isoforms of the Homer family, expresses with a transient peak in the early postnatal stage of mouse cerebellar granule cells (CGCs); however, the functions of Homer2a in CGCs are not fully understood yet. In this study, we investigated the physiological roles of Homer2a in CGCs using recombinant adenovirus vectors. Overexpression of the Homer2a N-terminal domain construct, which was made structurally reminiscent with Homer1a, altered NMDAR1 localization, decreased NMDA currents, and promoted the survival of CGCs. These results suggest that the Homer2a N-terminal domain acts as a dominant negative protein to attenuate NMDAR-mediated excitotoxicity. Moreover, we identified a novel short form N-terminal domain-containing Homer2, named Homer2e, which was induced by apoptotic stimulation such as ischemic brain injury. Our study suggests that the long and short forms of Homer2 are involved in apoptosis of CGCs.
Assuntos
Apoptose , Cerebelo/citologia , Proteínas de Arcabouço Homer/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Isquemia Encefálica/patologia , Proteínas de Arcabouço Homer/química , Proteínas de Arcabouço Homer/genética , Camundongos Endogâmicos ICR , Modelos Biológicos , N-Metilaspartato/metabolismo , Domínios Proteicos , Isoformas de Proteínas/metabolismoRESUMO
The cerebellar cortical circuit of mammals develops via a series of magnificent cellular events in the postnatal stage of development to accomplish the formation of functional circuit architectures. The contribution of genetic factors is thought to be crucial to cerebellar development. Therefore, it is essential to analyze the underlying transcriptome during development to understand the genetic blueprint of the cerebellar cortical circuit. In this review, we introduce the profiling of large numbers of spatiotemporal gene expression data obtained by developmental time-series microarray analyses and in situ hybridization cellular mRNA mapping, and the creation of a neuroinformatics database called the Cerebellar Development Transcriptome Database. Using this database, we have identified thousands of genes that are classified into various functional categories and are expressed coincidently with related cellular developmental stages. We have also suggested the molecular mechanisms of cerebellar development by functional characterization of several identified genes (Cupidin, p130Cas, very-KIND, CAPS2) responsible for distinct cellular events of developing cerebellar granule cells. Taken together, the gene expression profiling during the cerebellar development demonstrates that the development of cerebellar cortical circuit is attributed to the complex but orchestrated transcriptome.
Assuntos
Cerebelo/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento , Animais , Proteínas Reguladoras de Apoptose/fisiologia , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/fisiologia , Proteínas de Transporte/genética , Clonagem Molecular , Proteína Substrato Associada a Crk/genética , Proteína Substrato Associada a Crk/fisiologia , Bases de Dados Genéticas , Exonucleases , Perfilação da Expressão Gênica , Fatores de Troca do Nucleotídeo Guanina/genética , Fatores de Troca do Nucleotídeo Guanina/fisiologia , Proteínas de Arcabouço Homer , Glicoproteínas de Membrana/fisiologia , Camundongos , Proteínas da Mielina/fisiologia , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/fisiologia , Neurônios/fisiologia , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas Tirosina Quinases/fisiologia , Células de Purkinje/fisiologia , Sinapses/genética , Fatores de Transcrição/genéticaRESUMO
MYO18B is a class XVIII myosin, cloned as a tumor suppressor gene candidate. To investigate the mechanisms of MYO18B-dependent tumor suppression, MYO18B-interacting proteins were searched for by a yeast two-hybrid screen. HOMER2, a Homer/Ves1 family protein, was identified as a binding partner of MYO18B. These proteins co-localized in the regions of membrane protrusion and stress fiber, which are known as ones with filamentous actin-rich structures. Expression of HOMER2 enhanced the ability of MYO18B to suppress anchorage-independent growth. These results indicate that HOMER2 and MYO18B cooperate together in tumor suppression.
Assuntos
Proteínas de Transporte/metabolismo , Adesão Celular/fisiologia , Membrana Celular/fisiologia , Proliferação de Células , Miosinas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Proteínas de Arcabouço Homer , Camundongos , Células NIH 3T3 , Ligação ProteicaRESUMO
Microtubule dynamics, one of the key elements in neurite outgrowth, is regulated by various regulatory factors to determine the behavior of the neuronal growth cone and to form the specialized neuronal shape. SCG10 is a neuron-specific stathmin protein with a potent microtubule destabilizing factor and is enriched in the growth cones of the developing neurons. We investigated the functional role of SCG10 in neurite outgrowth using rat hippocampal primary cultured neurons. Genetic manipulation of SCG10 using a short-interfering RNA duplex markedly decreased the SCG10 expression level and significantly suppressed neurite outgrowth. This result was confirmed by immunodepletion experiments. On the other hand, the protein transduction of SCG10 using a polyarginine tag stimulated neurite outgrowth. Such manipulation of the SCG10 expression level affected microtubule morphology within the growth cones. A decrease in the SCG10 level converted the morphology to a more stable state, while an increase converted the morphology to a more dynamic state. However, an excess of SCG10 induced neurite retraction due to an excess of microtubule disassembly. These results suggest that SCG10 serves as an important regulatory factor of growth cone motility by enhancing microtubule dynamics, possibly through increasing the catastrophe frequency.