Subcellular proteomics for determining iron-limited remodeling of plastids in the model diatom Thalassiosira pseudonana (Bacillariophyta)

Diatoms are important primary producers in the world's oceans, yet their growth is constrained in large regions by low bioavailable iron (Fe). Low-Fe stress-induced limitation of primary production is due to requirements for Fe in components of essential metabolic pathways including photosynthesis and other chloroplast plastid functions. Studies have shown that under low-Fe stress, diatoms alter plastid-specific processes, including components of electron transport. These physiological changes suggest changes of protein content and in protein abundances within the diatom plastid. While in silico predictions provide putative information on plastid-localized proteins, knowledge of diatom plastid proteins remains limited in comparison to well-studied model photosynthetic organisms. To address this, we employed shotgun proteomics to investigate the proteome of subcellular plastid-enriched fractions from Thalassiosira pseudonana to gain a better understanding of how the plastid proteome is remodeled in response to Fe limitation. Using mass spectrometry-based peptide identification and quantification, we analyzed T. pseudonana grown under Fe-replete and -limiting conditions. Through these analyses, we inferred the relative quantities of each protein, revealing that Fe limitation regulates major metabolic pathways in the plastid, including the Calvin cycle. Additionally, we observed changes in the expression of light-harvesting proteins. In silico localization predictions of proteins identified in this plastid-enriched proteome allowed for an in-depth comparison of theoretical versus observed plastid-localization, providing evidence for the potential of additional protein import pathways into the diatom plastid.     

Antibodies to the vitronectin receptor (integrin alpha V beta 3) inhibit binding and infection of foot-and-mouth disease virus to cultured cells.

The amino acid sequence Arg-Gly-Asp (RGD) is highly conserved on the VP1 proteins of different serotypes and subtypes of foot-and-mouth disease virus (FMDV) and is essential for cell attachment. This sequence is also found in certain extracellular matrix proteins that bind to a family of cell surface receptors called integrins. Within the Picornaviridae family, enterovirus coxsackievirus A9 also has an RGD motif on its VP1 capsid protein and has recently been shown to utilize the vitronectin receptor integrin alpha V beta 3 as a receptor on monkey kidney cells. Competition binding experiments between type A12 FMDV and coxsackievirus A9 using BHK-21 and LLC-MK2 cells revealed shared receptor specificity between these two viruses. Polyclonal anti-serum to the vitronectin receptor and a monoclonal antibody to the alpha V subunit inhibited both FMDV binding and plaque formation, while a monoclonal antibody to the beta 3 subunit inhibited virus binding. In contrast, antibodies to the fibronectin receptor (alpha 5 beta 1) or to the integrin (alpha V beta 5) had no effect on either binding or plaque formation. These data demonstrate that the alpha V beta 3 vitronectin receptor can function as a receptor for FMDV.     

Molecular cloning of mei-41, a gene that influences both somatic and germline chromosome metabolism of Drosophila melanogaster

The mei-41 gene of Drosophila melanogaster plays an essential role in meiosis, in the maintenance of somatic chromosome stability, in postreplication repair and in DNA double-strand break repair. This gene has been cytogenetically localized to polytene chromosome bands 14C4-6 using available chromosomal aberrations. About 60 kb of DNA sequence has been isolated following a bidirectional chromosomal walk that extends over the cytogenetic interval 14C1-6. The breakpoints of chromosomal aberrations identified within that walk establish that the entire mei-41 gene has been cloned. Two independently derived mei-41 mutants have been shown to carry P insertions within a single 2.2 kb fragment of the walk. Since revertants of those mutants have lost the P element sequences, an essential region of the mei-41 gene is present in that fragment. A 10.5 kb genomic fragment that spans the P insertion sites has been found to restore methyl methanesulfonate resistance and female fertility of the mei-41 ^D3 mutants. The results demonstrate that all the sequences required for the proper expression of the mei-41 gene are present on this genomic fragment. This study provides the foundation for molecular analysis of a function that is essential for chromosome stability in both the germline and somatic cells.This Paper is dedicated to the memory of Professor James B. Boyd     

Small mammal populations in relation to hedgerow structure in an arable landscape

The population ecology of small mammals in hedgerows in arable farmland in eastern England is described. Features of hedgerows of importance to individual species are examined. Some 97% of the total 3042 mammals captured were wood mouse Apodemus sylvaticus , yellow-necked mouse Apodemus flavicollis , bank vole Clethrionomys glareolus and common shrew Sorex araneus . Small numbers of harvest mice Micromys minutus , field voles Microtus agrestis , pygmy shrews Sorex minutus and water shrews Neomys fodiens were also caught. Wood mouse, the most numerous species, showed a typical pattern of large numbers in autumn and winter, followed by a simultaneous decline over all hedges in early spring. Population changes were less clear in yellow-necked mouse and bank vole but the yellow-necked mouse was more scarce in the second year of study. Common shrews were most numerous in summer and declined rapidly in autumn. Hedgerow coppicing had a marked effect on yellow-necked mouse numbers but not on wood mouse. In an extensive survey of mammal numbers in relation to hedgerow features, ground cover was found to be the single largest factor influencing size of bank vole populations. Hedgerow condition (lack of gaps) was important to yellow-necked mice, which thrived only in well-established hedgerows. Wood mice appeared little influenced by the characteristics of the hedge. Common shrews were more abundant in hedgerows with adjacent permanent water.     

Germ-Line Effects of a Mutator, Mu2, in Drosophila Melanogaster

A mutator, mu2(a), in Drosophila melanogaster potentiates terminal deficiencies. In the female germ line the y mutant frequency induced by irradiation of mature oocytes with 5 Gy increases approximately twofold in heterozygotes and 20-fold in homozygotes compared with wild type. The recovery of terminal deficiencies is not limited to breaks close to chromosome ends; high frequencies of deficiencies can be recovered with breakpoints located in centric heterochromatin or near the middle of a chromosome arm. Lesions induced by γ-rays are repaired slowly in mu2(a) oocytes, but become ``fixed' as terminal deficiencies upon fertilization. A few lesions induced in wild-type females also produce terminal deficiencies. Mutator males do not exhibit an increase in terminal deletions, regardless of the germ cell stage irradiated. In addition, there is no increase in the mutant frequency when mature sperm are irradiated and fertilize eggs produced by mu2(a) females. The data are consistent with the hypothesis that lesions induced in sperm chromosomes are repaired after fertilization, while lesions induced in oocyte chromosomes are shunted instead to a mechanism that stabilizes broken chromosome ends. We propose that mu2 affects chromosomal structure during oogenesis, thereby modulating DNA repair.