RESUMEN
We analyzed the expression profile and cellular localization of the maize (Zea mays) Rpd3-type histone deacetylases genes ZmRpd3/101, ZmRpd3/102, and ZmRpd3/108 (indicated as ZmHDA101, ZmHDA102, and ZmHDA108 in the Plant Chromatin Database). This study shows that maize Rpd3 transcripts are present in all the organs and cellular domains analyzed, but we found that their amounts change during development, accumulating in the inner region of the endosperm, in vascular zones of the nucellus, in the tapetum, and in the tetrads. A similar expression profile and nucleus-cytoplasmic localization was observed for ZmRpd3 proteins. Glutathione S-transferase pull-down assays show that ZmRpd3 proteins can interact with the maize retinoblastoma-related (ZmRBR1) protein, an important regulator of cell cycle progression, and with the maize retinoblastoma-associated protein (ZmRbAp1). However, the three ZmRpd3 proteins do not mutually compete in the binding. These results suggest a general role of ZmRpd3 genes in the plant cell cycle and development. These observations also provide indications on possible mechanisms regulating their transcription and protein accumulation. Similarities in the gene expression profiles and protein interactions may indicate that functional redundancy among members of the ZmRpd3 gene family exists. However, a degree of functional divergence is also supported by our findings.
Asunto(s)
Regulación de la Expresión Génica de las Plantas/genética , Histona Desacetilasas/genética , Zea mays/enzimología , Secuencia de Aminoácidos , Secuencia de Bases , Cartilla de ADN , ADN Complementario/genética , Perfilación de la Expresión Génica/métodos , Regulación Enzimológica de la Expresión Génica/genética , Datos de Secuencia Molecular , Filogenia , Reacción en Cadena de la Polimerasa , ARN de Planta/genética , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Transcripción Genética , Zea mays/clasificación , Zea mays/genética , Zea mays/crecimiento & desarrolloRESUMEN
A maize histone deacetylase gene was identified as a homolog of yeast Hda1. The predicted protein corresponds to a previously purified maize deacetylase that is active as a protein monomer with a molecular weight of 48,000 and is expressed in all tissues of germinating embryos. Hda1 is synthesized as an enzymatically inactive protein with an apparent molecular weight of 84,000 that is processed to the active 48-kD form by proteolytic removal of the C-terminal part, presumably via a 65-kD intermediate. The enzymatically inactive 84-kD protein also is part of a 300-kD protein complex of unknown function. The proteolytic cleavage of ZmHda1 is regulated during maize embryo germination in vivo. Expression of the recombinant full-length protein and the 48-kD form confirmed that only the smaller enzyme form is active as a histone deacetylase. In line with this finding, we show that the 48-kD protein is able to repress transcription efficiently in a reporter gene assay, whereas the full-length protein, including the C-terminal part, lacks full repression activity. This report on the processing of Hda1-p84 to enzymatically active Hda1-p48 demonstrates that proteolytic cleavage is a mechanism to regulate the function of Rpd3/Hda1-type histone deacetylases.
Asunto(s)
Histona Desacetilasas/metabolismo , Zea mays/enzimología , Secuencia de Aminoácidos , Animales , Baculoviridae/genética , Secuencia de Bases , Línea Celular , Clonación Molecular , ADN de Plantas/genética , Endopeptidasas/metabolismo , Histona Desacetilasas/química , Histona Desacetilasas/genética , Datos de Secuencia Molecular , Peso Molecular , Procesamiento Proteico-Postraduccional , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Homología de Secuencia de Aminoácido , Transcripción Genética , Zea mays/embriología , Zea mays/genéticaRESUMEN
In mammalian cells the product of the human retinoblastoma tumour suppressor gene (pRb) can recruit Rpd3-like histone deacetylases to repress transcription. In this study, we investigated whether this mechanism might also be relevant in plants and found both conserved and distinct features. The expression profiles of the Zea mays Rpd3-type histone deacetylase (ZmRpd3I) and the retinoblastoma-related (ZmRBR1) homologues were analysed during endosperm development. GST pull-down and immunoprecipitation experiments showed a physical interaction between ZmRBRI and ZmRpd3I. Because ZmRpd3I lacks a LXCXE motif, conserved in several pRb-interacting proteins, we have mapped the amino acid domains involved in the ZmRBR1/ZmRpd3I interaction. Furthermore, we observed that ZmRbAp1, a maize member of the MSI/RbAp family, facilitated this protein interaction. Co-transformations of tobacco protoplasts with plasmids expressing ZmRBRI and ZmRpd3I showed that the two proteins cooperate in repressing gene transcription. Our findings represent the first indication that in plants a regulator of important biological processes, ZmRBRI, can recruit a histone deacetylase, ZmRpd3I, to control gene transcription.
