The human chordin gene encodes several differentially expressed spliced variants with distinct BMP opposing activities

During early embryogenesis of both vertebrates and invertebrates, antagonism between bone morphogenetic proteins (BMPs) and several unrelated secreted factors including Chordin (Chd) is a general mechanism by which the dorso-ventral axis is established. High affinity binding of Chd sequesters the BMP ligands in the extracellular space, preventing interactions with their membrane receptors. Another level of regulation consists in processing of vertebrate Chd or its Drosophila counterpart Sog by astacine metalloproteases like Xolloid-BMP-1/Tolloid, respectively, which releases an active BMP. Recently, it was shown that cleavage of Sog by Tolloid could generate novel BMP inhibitory activity and that sog is also capable of stimulation of BMP activity in a tolloid-dependant way. Activity and/or cleavage of Chd/Sog are influenced by other secreted factors like twisted gastrulation. In this study, we have cloned cDNAs of the human chordin gene (CHRD) and characterized alternative splice variants that code for C-truncated forms of the protein. We have found that CHRD is expressed in fetal as well as in adult tissues with relatively high levels in liver, cerebellum and female genital tract, suggesting functions in late embryogenesis and adult physiology. We also show that spliced variants are present with specific patterns in various tissues. When tested in an axis-duplication assay in Xenopus, we find that these variants can antagonize BMP activity. Altogether, these results suggest that, in addition to processing by metalloproteases, alternative splicing (AS) is another mechanism by which sub-products of CHRD can be generated to influence BMP activity in different developmental and physiological situations.     

The LGI1/epitempin gene encodes two protein isoforms differentially expressed in human brain

The leucine-rich, glioma inactivated 1 (LGI1)/Epitempin gene has been linked to two phenotypes as different as gliomagenesis and autosomal dominant lateral temporal epilepsy. Its function and the biochemical features of the encoded protein are unknown. We characterized the LGI1/Epitempin protein product by western blot analysis of mouse and human brain tissues. Two proteins of about 60 and 65 kDa were detected by an anti-LGI1 antibody within the expected molecular mass range. The two proteins appeared to reside in different subcellular compartments, as they were fractionated by differential centrifugation. The specificity of both polypeptides was validated by cell transfection assay and mass spectrometry analysis. Immunoblot analysis of protein distribution in various zones of the human brain revealed variable amounts of both proteins. Notably, these proteins were more abundant in the temporal neocortex than in the hippocampus, the difference in abundance of the 65-kDa product being particularly pronounced. These results suggest that the two protein isoforms encoded by LGI1/Epitempin are differentially expressed in the human brain, and that higher expression levels of these proteins in the lateral temporal cortex may underlie the susceptibility of this brain region to the epileptogenic effects of LGI1/Epitempin mutations.     

The Ultrastructure of Phaseolus vulgaris Chloroplasts

A comparison of bean chloroplasts after being fixed in potassiumpermanganate, osmium, and formaldehyde coupled with negativestaining shows that the general organization of the chloroplastis similar in all cases. However, the mature chloroplasts ofbean vary considerably in the extent and orientation of theinternal membranes—the grana and the interconnecting membranesbetween the grana. The interconnecting membranes are thin, branching,flexuous structures. This is illustrated by serial sectionsand by cross- and face-view sections of osmium- fixed chloroplastsand the best model, which allows for a considerable flexibilityin the orientation of the grana and also describes the thininterconnecting membranes between the grana, is the grana-fretworksystem proposed by Weier (1961). Furthermore, the comparative studies show that the internalregions of grana are separated from the stroma and that thegrana-fretwork systems appear to be a continuous membrane system.This membrane is single along the frets, end compartments, andgranal margins. It is double in the partitions of the granabut separated by a component, possibly a cementing material,which does not stain. It is suggested that this membrane isstructurally similar in the chloroplasts of higher plants, butthat its overall organization may vary from one plant to another.     

Identification of differentially expressed microRNAs by microarray: a possible role for microRNA genes in pituitary adenomas

MicroRNAs (miRNAs) are small non-coding RNAs that control gene expression by targeting mRNA. It has been demonstrated that miRNA expression is altered in many human cancers, suggesting that they may play a role in human neoplasia. To determine whether miRNA expression is altered in pituitary adenomas, we analyzed the entire miRNAome in 32 pituitary adenomas and in 6 normal pituitary samples by microarray and by Real-Time PCR. Here, we show that 30 miRNAs are differentially expressed between normal pituitary and pituitary adenomas. Moreover, 24 miRNAs were identified as a predictive signature of pituitary adenoma and 29 miRNAs were able to predict pituitary adenoma histotype. miRNA expression could differentiate micro- from macro-adenomas and treated from non-treated patient samples. Several of the identified miRNAs are involved in cell proliferation and apoptosis, suggesting that their deregulated expression may be involved in pituitary tumorigenesis. Predictive miRNAs could be potentially useful diagnostic markers, improving the classification of pituitary adenomas.     

A precursor microRNA in a cancer cell nucleus

In line with their broad-based effects, microRNAs (miRNAs), small non-coding RNA molecules ~22 nucleotides long that silence target mRNAs, are thought to act as oncogenes or tumor suppressor genes based on their inhibition of tumor-suppressive and oncogenic mRNAs, respectively. We and others previously showed that global downregulation of miRNAs, a common feature of human tumors, is functionally relevant to oncogenesis as impairment of miRNA biogenesis enhanced transformation in both cancer cells and a K-Ras-driven model of lung cancer. The dysregulation of miRNA biosynthesis in cancer emerges as a cancer-specific mechanism that enhances its tumorigenic capacity. These observations are further supported by the fact that frameshift mutations of TARBP2 occur in sporadic and hereditary carcinomas with microsatellite instability and that DICER1 mutations are associated with familial pleuropulmonary blastoma. Accordingly, it was reported that reduced expression of miRNA-processing factors is associated with poor prognosis in lung cancer and ovarian cancer. Recently we have also demonstrated the presence of Exportin 5 (XPO5) inactivating mutations in tumors with microsatellite instability. This observed genetic defect is responsible for nuclear retention of pre-miRNAs, thereby reducing miRNA processing. The characterized mutant form of the XPO5 protein lacks a C-terminal region that contributes to the formation of the pre-miRNA/XPO5/Ran-GTP ternary complex and the protein itself, as well as pre-miRNAs accumulating in the nucleus of cancer cells. Most importantly, the restoration of XPO5 function reverses the impaired export of pre-miRNAs and has tumor suppressor features. Our data suggest a cancer-specific mechanism to guide the subcellular distribution of miRNA precursors and prevent them from being processed to the active mature miRNA. The control of the miRNA biosynthesis pathway is emerging as an important mechanism in defining the spatiotemporal pattern of miRNA expression in cancer cells.     

The Antiranspirant Effect of Acetylsalcylic Acid on Phaseolus vulgaris

A series of experiments were carried out in order to elucidate whether acetylsalicylic acid (ASA) fed via petioles to beans could affect transpiration rate. In comparison with the water control it was found that a 10^?3M ASA reduces transpiration as much as 43%, a reduction equivalent to 5 × 10^?5M abscisic acid (ABA). Preliminary kinetics on the ABA, ASA and the water control on transpiration rate are presented and the results discussed.     

Distribution of a lectin in tissues of Phaseolus vulgaris

The distribution of lectin in various tissues of Phaseolus vulgaris L. (var. red) has been investigated with the use of a sensitive solid phase enzyme immunoassay. Plants were divided into roots, stems and leaves with a further dissection of the stem into internodes. Isolated tissues, from plants 3-10 weeks old, were screened for their lectin content, and the development of the lectin over this period determined. Stems and roots contained significant amounts of lectin, whereas the leaves only exhibited very low levels. Furthermore, a pronounced and differential increase in the biosynthesis of the lectin was observed in stem sections between 6 and 10 weeks, where the third internode displayed the highest lectin level.     

Comparative bioinformatic analysis of genes expressed in common bean (Phaseolus vulgaris L.) seedlings.

To rapidly and cost-effectively generate gene expression data, we developed an annotated unigene database of common bean (Phaseolus vulgaris L.). In this study, 3 cDNA libraries were constructed from the bean breeding line SEL1308, 1 from young leaf and 2 from seedlings inoculated or not inoculated with the fungal pathogen Colletotrichum lindemuthianum (Sacc. & Magnus) Briosi & Cavara, which causes anthracnose in common bean. To this date, 5255 single-pass sequences have been included in the database after selection based on sequence quality. These ESTs were trimmed and clustered using the computer programs Phred and CAP3 to form a unigene collection of 3126 unique sequences. Within clusters, 318 single nucleotide polymorphisms (SNPs) and 68 insertions-deletions (indels) were found, indicating the presence of paralogous gene families in our database. Each unigene sequence was analyzed for possible function using their similarity to known genes represented in the GenBank database and classified into 14 categories. Only 314 unigenes showed significant similarities to Phaseolus genomic sequences and P. vulgaris ESTs, which indicates that 90% (2818 unigenes) of our database represent newly discovered common bean genes. In addition, 12% (387 unigenes) were shown to be specific to common bean. This study represents a first step towards the discovery of novel genes in beans and a valuable source of molecular markers for expressed gene tagging and mapping.