Ultrastructural study of closed macular hole- preliminary application of a novel high magnification module combining with OCT.

BACKGROUND: As a novel high magnification module (HMM) combining with OCT (OCT-HMM) is able to detect the microstructure of retina, we apply it to explore the ultrastructure of the macula after closure of the idiopathic macular hole (IMH) by surgery. METHODS: This is an observational case series study in which patients with full-thickness IMHs who had undergone successful macular closure by vitrectomy and internal limiting membrane peeling and healthy subjects were recruited. After comprehensive ophthalmic examinations, the images of macular area were obtained and collected by professional operators using OCT-HMM. Then images were independently analyzed by 4 masked vitreoretinal specialists. RESULTS: A total of 24 IMH eyes and 42 healthy eyes were examined. HMM images were obtained in 10 IMH eyes. Among them, 4 eyes whose macula closed completely with recovery of photoreceptor layer presented a dark arc nasal to the fovea, oriented to the optic, and the notch of arc faced temporally. Six eyes in which the macula closed incompletely with photoreceptor cells loss revealed a dark ring with uneven bright spots inside. The other 14 eyes failed to obtain clear images by OCT-HMM. The contra lateral eyes of the patients and the healthy subjects' eyes succeeded to obtain the HMM images which displayed evenly grey background thickly covered with tiny bright dots that was in similar size and evenly and widely distributed and there no dark arc or ring. OCT B-scan and IR images could be acquired in all of the IMH and healthy eyes. CONCLUSION: The preliminary application of HMM has supplied us a brand-new insight into the microstructure of closed IMH. A dark arc sign could be detected with OCT-HMM in the macula which was functionally closed after surgery that was probably the healing mark on a microstructure photoreceptors level. Its existence and shape indicated that the functional closure followed by a retinal displacement mainly horizontally from temporal side to nasal side but not symmetric centripetally.

Correction: Characterization and Functional Analysis of 4-Coumarate:CoA Ligase Genes in Mulberry.

[This corrects the article DOI: 10.1371/journal.pone.0155814.].

Characterization and Functional Analysis of 4-Coumarate:CoA Ligase Genes in Mul-berry.

A small, multigene family encodes 4-coumarate:CoA ligases (4CLs) that catalyze the ligation of CoA to hydroxycinnamic acids, a branch point directing metabolites to flavonoid or monolignol pathways. In this study, we characterized four 4CL genes from M. notabilis Genome Database, and cloned four Ma4CL genes from M. atropurpurea cv. Jialing No.40. A tissue-specific expression analysis indicated that Ma4CL3 was expressed at higher levels than the other genes, and that Ma4CL3 was strongly expressed in root bark, stem bark, and old leaves. Additionally, the expression pattern of Ma4CL3 was similar to the trend of the total flavonoid content throughout fruit development. A phylogenetic analysis suggested that Mn4CL1, Mn4CL2, and Mn4CL4 belong to class I 4CLs, and Mn4CL3 belongs to class II 4CLs. Ma4CL genes responded differently to a series of stresses. Ma4CL3 expression was higher than that of the other Ma4CL genes following wounding, salicylic acid, and ultraviolet treatments. An in vitro enzyme assay indicated that 4-coumarate acid was the best substrate among cinnamic acid, 4-coumarate acid, and caffeate acid, but no catalytic activity to sinapate acid and ferulate acid. The results of subcellular localization experiments showed that Ma4CL3 localized to the cytomembrane, where it activated transcription. We used different vectors and strategies to fuse Ma4CL3 with stilbene synthase (STS) to construct four Ma4CL-MaSTS co-expression systems to generate resveratrol. The results indicated that only a transcriptional fusion vector, pET-Ma4CL3-T-MaSTS, which utilized a T7 promoter and lac operator for the expression of MaSTS, could synthesize resveratrol.

Mulberry Transcription Factor MnDREB4A Confers Tolerance to Multiple Abiotic Stresses in Transgenic Tobacco.

The dehydration responsive element binding (DREB) transcription factors have been reported to be involved in stress responses. Most studies have focused on DREB genes in subgroups A-1 and A-2 in herbaceous plants, but there have been few reports on the functions of DREBs from the A-3-A-6 subgroups and in woody plants. Moreover, mulberry trees are ecologically and economically important perennial woody plants, but there has been little research on its stress physiology, biochemistry and molecular biology. In this study, a DREB gene from the mulberry tree, designated as MnDREB4A, classified into the A-4 subgroup by our previous study, was selected for further characterization. Our results showed that the MnDREB4A protein was localized to the nucleus where it activated transcription. The promoter of MnDREB4A can direct prominent expression downstream of the ß-glucuronidase (GUS) gene under heat, cold, drought and salt stress, and GUS staining was deepest after 12 h of stress treatment. The MnDREB4A-overexpression transgenic tobacco showed the improved growth phenotype under untreated conditions, such as greener leaves, longer roots, and lower water loss and senescence rates. Overexpression of MnDREB4A in tobacco can significantly enhance tolerance to heat, cold, drought, and salt stresses in transgenic plants. The leaf discs and seedlings of transgenic plants reduced leaf wilting and senescence rates compared to the wild type plants under the different stress conditions. Further investigation showed that transgenic plants also had higher water contents and proline contents, and lower malondialdehyde contents under untreated condition and stress conditions. Our results indicate that the MnDREB4A protein plays an important role in plant stress tolerance.

Characterization and expression profiles of MaACS and MaACO genes from mulberry (Morus alba L.).

1-Aminocyclopropane-1-carboxylic acid synthase (ACS) and 1-aminocyclopropane-1-carboxylic acid oxidase (ACO) are encoded by multigene families and are involved in fruit ripening by catalyzing the production of ethylene throughout the development of fruit. However, there are no reports on ACS or ACO genes in mulberry, partly because of the limited molecular research background. In this study, we have obtained five ACS gene sequences and two ACO gene sequences from Morus Genome Database. Sequence alignment and phylogenetic analysis of MaACO1 and MaACO2 showed that their amino acids are conserved compared with ACO proteins from other species. MaACS1 and MaACS2 are type I, MaACS3 and MaACS4 are type II, and MaACS5 is type III, with different C-terminal sequences. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) expression analysis showed that the transcripts of MaACS genes were strongly expressed in fruit, and more weakly in other tissues. The expression of MaACO1 and MaACO2 showed different patterns in various mulberry tissues. MaACS and MaACO genes demonstrated two patterns throughout the development of mulberry fruit, and both of them were strongly up-regulated by abscisic acid (ABA) and ethephon.

[Suppression subtractive hybridization for the identification of differentially expressed genes in the hippocampus of the offsprings of lead exposed female rats].

OBJECTIVE: To screen and identify differentially expressed genes in the hippocampus of the offsprings of lead exposed female rats in order to provide a theoretical basis for identifying learning and memory deficits related genes. METHODS: RNA was extracted from the hippocampus of young rats with learning and memory deficits due to maternal lead exposure. Suppression subtractive hybridization was used to identify the differentially expressed genes in the hippocampus. RESULTS: An effective subtracted library was constructed which consisted of approximately 200 clones. Sequencing for the library identified 93 clones harboring insertion fragments which included 43 different genes and 4 unknown genes. These genes might be related to learning and memory deficits due to maternal lead exposure. CONCLUSIONS: The up-regulated genes in the hippocampus of young rats from pregnant rats under lead exposure include some housekeeping genes and some proteins involved in cellular protein folding, signal transduction, stress response and DNA methylation. These proteins might be directly related to a significant reduction in learning and memory abilities in the young rats.

A novel mutation in EXT2 gene in a Chinese family with hereditary multiple exostoses.

Hereditary multiple exostoses (HME) is an autosomal-dominant disorder characterized by the presence of bony outgrowths on the long bones. In this report, we describe a Chinese family with HME. Linkage analysis and mutation detection were performed. Linkage with the EXT2 was established in this family. A novel mutation, EXT2 c239-244delG, was identified. Mutation analysis in a family with HME allows for genetic counseling and prenatal diagnosis.