Effect of NaCl stress on dihaploid tobacco lines tolerant to Potato virus Y

Salinity is an important abiotic factor that limits plant growth and development. The influence of salt stress induced by sodium chloride on plant growth, proline content, level of lipid peroxidation and activities of antioxidative enzymes was studied in F1 hybrid DH10 and four dihaploid lines (207B, 238C, 239K, 244B) of tobacco (Nicotiana tabacum L.). Dihaploids were obtained from anther-derived haploids of hybrid DH10 and were previously proved to be tolerant to Potato virus Y (PVY). In our study, plants were grown in vitro and exposed to NaCl (100 and 200 mM) for 33 days. All dihaploids and hybrid DH10 showed reduced growth after NaCl treatment. They accumulated significant amounts of sodium and proline in response to salt stress as have already been observed in tobacco and other plant species. In tobacco exposed to NaCl the lipid peroxidation level did not increase and activities of superoxide dismutase (SOD), guaiacol peroxidase (POD), ascorbate peroxidase and catalase (CAT) mostly did not change significantly. The exception was line 239K where salt induced higher activities of SOD, CAT and POD. Two (238C and 244B) out of four dihaploids appeared more susceptible to salt stress as they showed weak growth in correlation with high proline and sodium content. Therefore, it seems that salt tolerance is not associated with tolerance to PVY. Variations in malondialdehyde and proline content as well as in enzymes activities observed among tobacco lines imply that dihaploids have different genetic properties which might result in different sensitivity to NaCl.     

Autosomal dominant ataxia: Genetic evidence for locus heterogeneity from a cuban founder-effect population

The locus for autosomal dominant ataxia with a diagnosis of olivo-ponto-cerebellar atrophy at autopsy has been previously assigned to chromosome 6p. However, evidence for two alternative locations has been reported. We have recently described a large potential founder-effect population of such patients in the Holguin province of Cuba. With an estimated 1,000 patients available for analysis, this extensive cluster of families provides a unique opportunity for the definitive localization of the genetic mutation. Linkage analysis between the disease locus in this population and markers within and flanking the HLA region on chromosome 6 were undertaken in 12 families comprising over 100 affected individuals. Despite similarity in the clinical phenotype between those families where the disease locus has been reported to be linked to the HLA locus and the Cuban patients, no evidence of linkage to this region could be demonstrated in the latter. The disease locus was excluded from a 96-cM genetic interval of the short arm of chromosome 6, encompassing the F13A1-HLA-GLO1-MUT/D6S4 loci. These data strongly support the existence of genetic heterogeneity for the disease.     

Chain termini cross-talk in the swapping process of bovine pancreatic ribonuclease

3D domain swapping is the process by which two or more protein molecules exchange part of their structure to form intertwined dimers or higher oligomers. Bovine pancreatic ribonuclease (RNase A) is able to swap the N-terminal α-helix (residues 1-13) and/or the C-terminal β-strand (residues 116-124), thus forming a variety of oligomers, including two different dimers. Cis-trans isomerization of the Asn113-Pro114 peptide group was observed when the protein formed the C-terminal swapped dimer. To study the effect of the substitution of Pro114 on the swapping process of RNase A, we have prepared and characterized the P114A monomeric and dimeric variants of the enzyme. In contrast with previous reports, the crystal structure and NMR data on the monomer reveals a mixed cis-trans conformation for the Asn113-Ala114 peptide group, whereas the X-ray structure of the C-terminal swapped dimer of the variant is very close to that of the corresponding dimer of RNase A. The mutation at the C-terminus affects the capability of the N-terminal α-helix to swap and the stability of both dimeric forms. The present results underscore the importance of the hydration shell in determining the cross-talk between the chain termini in the swapping process of RNase A.     

Atrial natriuretic peptides elevate cyclic GMP levels in primary cultures of rat ependymal cells

The aim of this study was to examine the effect of atrial natriuretic peptides on primary cultures of ependymal cells, as measured by changes in intracellular levels of cyclic GMP. Incubation of ependymal cells with rat atrial natriuretic peptide-(1-28) (rANP) elicited a 30-fold increase in ependymal cGMP content within 1 min and more than a 100-fold increase within 10 min to a plateau value of approximately 30 pmol/mg protein. The C-type natriuretic peptide (CNP) elicited a similar increase in cGMP levels; however the maximal effect was observed within 1 min and the levels subsequently dropped by 90% to a low plateau within 10 min. A comparison of the concentration-response curves for rANP, human ANP-(1-28) (hANP) and CNP showed that rANP, hANP and CNP had similar effects, with regards to elevation of cGMP levels at high concentrations, but with differing EC50 values. These results demonstrate the presence of a heterogenous population of functional ANP receptors in cultured ependymal cells suggesting that ANP may regulate specific ependymal cell activity.     

TRACE ELEMENT ANALYSIS OF PROTEINS DIRECTLY FROM 2D-PAGE: AN EFFICIENT STRATEGY FOR METALLOPROTEOMICS

Inductively coupled plasma–mass spectrometry (ICP-MS) is a sensitive analytical method to detect the total concentrations of elements in biological samples, but it is unable to identify molecules that can bind to metals, and for this reason it is vital to combine this method's use with other biochemical techniques. Therefore, in order to identify elements complexed to specific proteins, a very relevant combination of bidimensional electrophoresis followed by ICP-MS was used. Protein spots from gels were excised and submitted directly to element detection, a method not reported before. This report focused on the use of plasma from people with laryngeal carcinoma. Most elements were below detection level, with only Cr and Pb being observed in all samples. Although the relationship between metals and laryngeal cancer was not conclusive, it is possible to affirm that the methodology utilized here is successful and has the advantage of determining to which proteins the elements bind.     

New data on freshwater psammic Gastrotricha from Brazil

Current knowledge of freshwater gastrotrich fauna from Brazil is underestimated as only two studies are available. The present communication is a taxonomic account of the first-ever survey of freshwater Gastrotricha in Minas Gerais State. Samplings were carried out yielding six species of three Chaetonotidae genera: Aspidiophorus cf. pleustonicus, Ichthydium cf. chaetiferum, Chaetonotus acanthocephalus, Chaetonotus heideri, Chaetonotus cf. succinctus, Chaetonotus sp., and also an undescribed species belonging to the genus Redudasys (incertae sedis): this is the first finding of specimens of Redudasys outside of original type locality. These preliminary observations suggest that the knowledge of the biodiversity of Gastrotricha in the Minas Gerais State, as well as in the whole Brazil, will certainly increase as further investigations are undertaken, and that freshwater Macrodasyida may be more common than previously thought.     

Alternative techniques to study characters of the genitalia in Lepidoptera

The present note aims to describe two alternative methods for observing genitalia in Lepidoptera. The first one provides means to examine both male and female genitalia without spoiling the scales of the abdomen, preserving it attached to the thorax and aesthetically similar to an unexamined specimen. The second one provides ways of observing certain characters on the male genitalia in a non-destructive way, and does not depend on time-consuming removing and dissection of the abdomen. It is expected that the presented techniques will help on morphological studies and on identifying similar species which consistently differ in genitalic armatures.     

The cooperation between hMena overexpression and HER2 signalling in breast cancer

hMena and the epithelial specific isoform hMena(11a) are actin cytoskeleton regulatory proteins belonging to the Ena/VASP family. EGF treatment of breast cancer cell lines upregulates hMena/hMena(11a) expression and phosphorylates hMena(11a), suggesting cross-talk between the ErbB receptor family and hMena/hMena(11a) in breast cancer. The aim of this study was to determine whether the hMena/hMena(11a) overexpression cooperates with HER-2 signalling, thereby affecting the HER2 mitogenic activity in breast cancer. In a cohort of breast cancer tissue samples a significant correlation among hMena, HER2 overexpression, the proliferation index (high Ki67), and phosphorylated MAPK and AKT was found and among the molecular subtypes the highest frequency of hMena overexpressing tumors was found in the HER2 subtype. From a clinical viewpoint, concomitant overexpression of HER2 and hMena identifies a subgroup of breast cancer patients showing the worst prognosis, indicating that hMena overexpression adds prognostic information to HER2 overexpressing tumors. To identify a functional link between HER2 and hMena, we show here that HER2 transfection in MCF7 cells increased hMena/hMena(11a) expression and hMena(11a) phosphorylation. On the other hand, hMena/hMena(11a) knock-down reduced HER3, AKT and p44/42 MAPK phosphorylation and inhibited the EGF and NRG1-dependent HER2 phosphorylation and cell proliferation. Of functional significance, hMena/hMena(11a) knock-down reduced the mitogenic activity of EGF and NRG1. Collectively these data provide new insights into the relevance of hMena and hMena(11a) as downstream effectors of the ErbB receptor family which may represent a novel prognostic indicator in breast cancer progression, helping to stratify patients.