RESUMO
Mycorrhizae are ubiquitous symbioses established between fungi and plant roots. Orchids, in particular, require compatible mycorrhizal fungi for seed germination and protocorm development. Unlike arbuscular mycorrhizal fungi, which have wide host ranges, orchid mycorrhizal fungi are often highly specific to their host orchids. However, the molecular mechanism of orchid mycorrhizal symbiosis is largely unknown compared to that of arbuscular mycorrhizal and rhizobial symbiosis. Here, we report that an endophytic Sebacinales fungus, Serendipita indica, promotes seed germination and the development of protocorms into plantlets in several epiphytic Epidendroideae orchid species (6 species in 2 genera), including Dendrobium catenatum, a critically endangered orchid with high medicinal value. Although plant-pathogen interaction and high meristematic activity can induce the hypoxic response in plants, it has been unclear whether interactions with beneficial fungi, especially mycorrhizal ones, also involve the hypoxic response. By studying the symbiotic relationship between D. catenatum and S. indica, we determined that hypoxia-responsive genes, such as those encoding alcohol dehydrogenase (ADH), are highly induced in symbiotic D. catenatum protocorms. In situ hybridization assay indicated that the ADH gene is predominantly expressed in the basal mycorrhizal region of symbiotic protocorms. Additionally, the ADH inhibitors puerarin and 4-methylpyrazole both decreased S. indica colonization in D. catenatum protocorms. Thus, our study reveals that S. indica is widely compatible with orchids and that ADH and its related hypoxia-responsive pathway are involved in establishing successful symbiotic relationships in germinating orchids.
Assuntos
Basidiomycota , Dendrobium , Micorrizas , Orchidaceae , Simbiose , Dendrobium/genética , Sementes , Micorrizas/fisiologia , Basidiomycota/fisiologia , Orchidaceae/genética , FilogeniaRESUMO
Migration inhibitory factor-related protein 14 (MRP14) is one of calcium-binding proteins, referred as S100A9. The heterodimeric molecule formed by MRP14 with its partner MRP8 (S100A8) is the major fatty acid carrier in neutrophils. The MRP8/14 complex has been also implicated in the intracellular transport of arachidonic acid and its precursors in keratinocytes. We show here the involvement of MRP14 in human esophageal cancer. In an initial study, mRNA differential display-reverse transcription polymerase chain reaction (DD-PCR) was performed with two esophageal carcinomas, one esophageal adenocarcinoma and matched normal adjacent mucosa. DD-PCR with the arbitrary primer OPA3 showed that one cDNA band was highly expressed in normal tissues, but disappeared or substantially decreased in tumor counterparts. It was later identified to be the 3'-end of migration inhibitory factor-related protein 14 (MRP14). Northern blotting, RT-PCR and Western blotting corroborated the down-regulation of MRP14 in 58/64 squamous cell carcinomas and 2/2 adenocarcinomas as compared with adjacent normal epithelia of the esophagus. MRP14 was undetectable in 3/3 esophageal-carcinoma cell lines. Immunochemistry demonstrated that expression of MRP14 was restricted to normal esophageal epithelia. No mutation was found in the genomic DNA of the MRP14 gene by PCR and directed DNA sequencing. Our finding suggested that the reduction of MRP14 expression is a frequent event in Chinese human esophageal cancer.
Assuntos
Calgranulina B/genética , Neoplasias Esofágicas/genética , Regulação Neoplásica da Expressão Gênica , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Adenocarcinoma/epidemiologia , Adenocarcinoma/genética , Adenocarcinoma/patologia , Adulto , Idoso , Northern Blotting , Western Blotting , Calgranulina B/metabolismo , Carcinoma de Células Escamosas/epidemiologia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Núcleo Celular/química , China/epidemiologia , Análise Mutacional de DNA , DNA Complementar/genética , Regulação para Baixo/genética , Epitélio/química , Neoplasias Esofágicas/epidemiologia , Neoplasias Esofágicas/patologia , Feminino , Perfilação da Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , RNA Neoplásico/genética , RNA Neoplásico/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
To better understand the molecular events underlying the development of oesophageal cancer, we have isolated the genes dysregulated in primary oesophageal cancer tissues using a modified differential display polymerase chain reaction (DD-PCR). In the present study, a gene designated C15orf6 was identified. The C15orf6 gene, encompassing 25 kb, is composed of 11 exons with a mRNA of 1948 bp. Database searching showed that C15orf6 was 100% homologous to the Rh type C-glycoprotein (RhCG) with the same open reading frame, but 16 bp longer than RhCG at the 5'-end. The gene was highly expressed in human oesophagus, cervix, oral cavity, skin and kidney, but undetectable in the other 14 adult normal tissues examined. Northern blot, RT-PCR and western blot analysis showed that RhCG/C15orf6 was frequently lost or dramatically reduced in primary oesophageal cancer tissues (30/34) compared with the corresponding normal oesophageal mucosa. Three oesophageal-cancer cell lines tested lacked RhCG/C15orf6 expression. Immunohistochemistry revealed that in normal oesophageal tissues, RhCG/C15orf6 was mainly expressed in the plasma membrane of the epithelial cells. In addition, Rh-associated glycoprotein (RhAG) expression was also commonly silenced in both oesophageal cancer cell lines (2/3) and primary oesophageal cancer tissues (11/13). To our knowledge, this is the first time that RhAG expression has been seen in oesophageal epithelium and extends the functional role of the RhAG protein beyond the erythrocyte. These data suggest that inactivation of RhCG/C15orf6 and RhAG occurs frequently during the development of human oesophageal cancer.