COS1, a two-component histidine kinase that is involved in hyphal development in the opportunistic pathogen Candida albicans

Two-component histidine kinases recently have been found in eukaryotic organisms including fungi, slime molds, and plants. We describe the identification of a gene, COS1, from the opportunistic pathogen Candida albicans by using a PCR-based screening strategy. The sequence of COS1 indicates that it encodes a homolog of the histidine kinase Nik-1 from the filamentous fungus Neurospora crassa. COS1 is also identical to a gene called CaNIK1 identified in C. albicans by low stringency hybridization using CaSLN1 as a probe [Nagahashi, S., Mio, T., Yamada-Okabe, T., Arisawa, M., Bussey, H. & Yamada-Okabe, H. (1998) Microbiol. 44, 425–432]. We assess the function of COS1/CaNIK1 by constructing a diploid deletion mutant. Mutants lacking both copies of COS1 appear normal when grown as yeast cells; however, they exhibit defective hyphal formation when placed on solid agar media, either in response to nutrient deprivation or serum. In constrast to the Δnik-1 mutant, the Δcos1/Δcos1 mutant does not demonstrate deleterious effects when grown in media of high osmolarity; however both Δnik-1 and Δcos1/Δcos1 mutants show defective hyphal formation. Thus, as predicted for Nik-1, Cos1p may be involved in some aspect of hyphal morphogenesis and may play a role in virulence properties of the organism.

Evidence for balancing selection operating at the het-c heterokaryon incompatibility locus in a group of filamentous fungi

In filamentous fungi, het loci (for heterokaryon incompatibility) are believed to regulate self/nonself-recognition during vegetative growth. As filamentous fungi grow, hyphal fusion occurs within an individual colony to form a network. Hyphal fusion can occur also between different individuals to form a heterokaryon, in which genetically distinct nuclei occupy a common cytoplasm. However, heterokaryotic cells are viable only if the individuals involved have identical alleles at all het loci. One het locus, het-c, has been characterized at the molecular level in Neurospora crassa and encodes a glycine-rich protein. In an effort to understand the role of this locus in filamentous fungi, we chose to study its evolution by analyzing het-c sequence variability in species within Neurospora and related genera. We determined that the het-c locus was polymorphic in a field population of N. crassa with close to equal frequency of each of the three allelic types. Different species and even genera within the Sordariaceae shared het-c polymorphisms, indicating that these polymorphisms originated in an ancestral species. Finally, an analysis of the het-c specificity region shows a high occurrence of nonsynonymous substitution. The persistence of allelic lineages, the nearly equal allelic distribution within populations, and the high frequency of nonsynonymous substitutions in the het-c specificity region suggest that balancing selection has operated to maintain allelic diversity at het-c. Het-c shares this particular evolutionary characteristic of departing from neutrality with other self/nonself-recognition systems such as major histocompatibility complex loci in mammals and the S (self-incompatibility) locus in angiosperms.

Structural Requirements of Tom40 for Assembly into Preexisting TOM Complexes of Mitochondria

Tom40 is the major subunit of the translocase of the outer mitochondrial membrane (the TOM complex). To study the assembly pathway of Tom40, we have followed the integration of the protein into the TOM complex in vitro and in vivo using wild-type and altered versions of the Neurospora crassa Tom40 protein. Upon import into isolated mitochondria, Tom40 precursor proteins lacking the first 20 or the first 40 amino acid residues were assembled as the wild-type protein. In contrast, a Tom40 precursor lacking residues 41 to 60, which contains a highly conserved region of the protein, was arrested at an intermediate stage of assembly. We constructed mutant versions of Tom40 affecting this region and transformed the genes into a sheltered heterokaryon containing a tom40 null nucleus. Homokaryotic strains expressing the mutant Tom40 proteins had growth rate defects and were deficient in their ability to form conidia. Analysis of the TOM complex in these strains by blue native gel electrophoresis revealed alterations in electrophoretic mobility and a tendency to lose Tom40 subunits from the complex. Thus, both in vitro and in vivo studies implicate residues 41 to 60 as containing a sequence required for proper assembly/stability of Tom40 into the TOM complex. Finally, we found that TOM complexes in the mitochondrial outer membrane were capable of exchanging subunits in vitro. A model is proposed for the integration of Tom40 subunits into the TOM complex.

Phosphate transporters from the higher plant Arabidopsis thaliana.

Two cDNAs (AtPT1 and AtPT2) encoding plant phosphate transporters have been isolated from a library prepared with mRNA extracted from phosphate-starved Arabidopsis thaliana roots, The encoded polypeptides are 78% identical to each other and show high degree of amino acid sequence similarity with high-affinity phosphate transporters of Saccharomyces cerevisiae, Neurospora crassa, and the mycorrhizal fungus Glomus versiforme. The AtPT1 and AtPT2 polypeptides are integral membrane proteins predicted to contain 12 membrane-spanning domains separated into two groups of six by a large charged hydrophilic region. Upon expression, both AtPT1 and AtPT2 were able to complement the pho84 mutant phenotype of yeast strain NS219 lacking the high-affinity phosphate transport activity. AtPT1 and AtPT2 are representatives of two distinct, small gene families in A. thaliana. The transcripts of both genes are expressed in roots and are not detectable in leaves. The steady-state level of their mRNAs increases in response to phosphate starvation.

Uso de modelagem de nicho fundamental na avaliação do padrão de distribuição geográfica de espécies vegetais

Atualmente vêm sendo desenvolvidas e utilizadas várias técnicas de modelagem de distribuição geográfica de espécies com os mais variados objetivos. Algumas dessas técnicas envolvem modelagem baseada em análise ambiental, nas quais os algoritmos procuram por condições ambientais semelhantes àquelas onde as espécies foram encontradas, resultando em áreas potenciais onde as condições ambientais seriam propícias ao desenvolvimento dessas espécies. O presente estudo trata do uso da modelagem preditiva de distribuição geográfica, através da utilização de algoritmo genético e algoritmo de distância, de espécies como ferramenta para a conservação de espécies vegetais, em três situações distintas: modelagem da distribuição do bioma cerrado no estado de São Paulo; previsão da ocorrência de espécies arbóreas visando à restauração da cobertura vegetal na bacia do Médio Paranapanema e modelagem da distribuição de espécies ameaçadas de extinção (Byrsonima subterranea). A metodologia empregada e os resultados obtidos foram considerados satisfatórios para a geração de modelos de distribuição geográfica de espécies vegetais, baseados em dados abióticos, para as regiões de estudo. A eficácia do modelo em predizer a ocorrência de espécies do cerrado é maior se forem utilizados apenas pontos de amostragem com fisionomias de cerrado, excluindo-se áreas de transição. Para minimizar problemas decorrentes da falta de convergência do algoritmo utilizado GARP (Genetic Algorithm for Rule Set Production), foram gerados 100 modelos para cada espécie modelada. O uso de modelagem pode auxiliar no entendimento dos padrões de distribuição de um bioma ou ecossistema em uma análise regional e local.

Heterogeneidade espacial e variabilidade temporal de dois reservatórios com diferentes graus de trofia, no Estado de São Paulo

O entendimento da comunidade fitoplanctônica em sistemas instáveis, como por exemplo reservatórios, necessita conhecimento de escalas de variabilidade. Com base nisso, um estudo sobre a heterogeneidade espacial e variabilidade temporal de dois reservatórios com diferentes graus de trofia, no Estado de São Paulo foi realizado em 20 estações no reservatório de Salto Grande e em 19 no reservatório do Lobo, em 3 dias consecutivos, em quatro períodos: outubro de 1999, janeiro, abril e junho e julho de 2000. Para tanto foram determinadas as concentrações de nutrientes totais e dissolvidos, material em suspensão, carbono inorgânico, clorofila a, biomassa, densidade, composição e produtividade primária da comunidade fitoplanctônica e os perfis de oxigênio dissolvido, temperatura, pH e condutividade. Os dois reservatórios tiveram estruturas espaciais semelhantes com a formação de três zonas distintas. A zona de rio, misturada, com menor penetração de luz e maior concentração de nutrientes, a zona de transição, e a zona lacustre, mais estratificada, com maior penetração de luz e menor concentração de nutrientes. Apesar dessa compartimentalização a heterogeneidade espacial no reservatório de Salto Grande foi maior que no reservatório do Lobo, sobretudo em função do gradiente longitudinal de nutrientes e luz. A variabilidade diária (3 dias) nos dois reservatórios não foi significativa na determinação da comunidade fitoplanctônica. A escala de variabilidade sazonal, nos dois reservatórios, foi determinada, principalmente pela variação nos padrões de estratificação e mistura sendo, assim, determinante na composição da comunidade fitoplanctônica. Essa influência foi mais evidente no reservatório do Lobo. A variação temporal e heterogeneidade espacial das mais abundantes espécies e grupos taxonômicos da comunidade fitoplanctônica, (Microcystis aeruginosa, Anabaena crassa e Anabaena circinalis em Salto Grande e Aphanocapsa delicatissima, Coelastrum reticulatum e Aulacoseira granulata no Lobo) nos dois reservatórios foram determinados pelos complexos processos de estratificação e mistura e da disponibilidade de luz. Os resultados obtidos são importantes para o entendimento da variabilidade ambiental de reservatórios tropicais e no planejamento de amostragens que visem o gerenciamento desses sistemas.

Omics for Investigating Chitosan as an Antifungal and Gene Modulator

Chitosan is a biopolymer with a wide range of applications. The use of chitosan in clinical medicine to control infections by fungal pathogens such as Candida spp. is one of its most promising applications in view of the reduced number of antifungals available. Chitosan increases intracellular oxidative stress, then permeabilizes the plasma membrane of sensitive filamentous fungus Neurospora crassa and yeast. Transcriptomics reveals plasma membrane homeostasis and oxidative metabolism genes as key players in the response of fungi to chitosan. A lipase and a monosaccharide transporter, both inner plasma membrane proteins, and a glutathione transferase are main chitosan targets in N. crassa. Biocontrol fungi such as Pochonia chlamydosporia have a low content of polyunsaturated free fatty acids in their plasma membranes and are resistant to chitosan. Genome sequencing of P. chlamydosporia reveals a wide gene machinery to degrade and assimilate chitosan. Chitosan increases P. chlamydosporia sporulation and enhances parasitism of plant parasitic nematodes by the fungus. Omics studies allow understanding the mode of action of chitosan and help its development as an antifungal and gene modulator.

Paleomagnetic and stable isotope measurements and distribution of benthic foraminifera in sediment core PS1388-3 from the Antarctic continental margin in the eastern Weddell Sea

A stable isotope record from the eastern Weddell Sea from 69°S is presented. For the first time, a 250,000-yr record from the Southern Ocean can be correlated in detail to the global isotope stratigraphy. Together with magnetostratigraphic, sedimentological and micropalaeontological data, the stratigraphic control of this record can be extended back to 910,000 yrs B.P. A time scale is constructed by linear interpolation between confirmed stratigraphic data points. The benthic d18O record (Epistominella exigua) reflects global continental ice volume changes during the Brunhes and late Matuyama chrons, whereas the planktonic isotopic record (Neogloboquadrina pachyderma) may be influenced by a meltwater lid caused by the nearby Antarctic ice shelf and icebergs. The worldwide climatic improvement during deglaciations is documented in the eastern Weddell Sea by an increase in production of siliceous plankton followed, with a time lag of approximately 10,000 yrs, by planktonic foraminifera production. Peak values in the difference between planktonic and benthic d13C records, which are 0.5 per mil higher during warm climatic periods than during times with expanded continental ice sheets, also suggest increased surface productivity during interglacials in the Southern Ocean.

Distribution of foraminifera, pre-Pliocene foraminifera and macrofossil debris in sediment core CRP-2 (Table 1)

The stratigraphic distribution, assemblage content, paleoecology and age of foraminifera recovered in fourteen of sixteen samples from the 5.63 m thick CRP-2 (Lithostratigraphic Unit 2.2) are discussed. LSU 2.2 comprises four discrete lithologic beds. The upward sequence is informally referred to as the lower sand bed, diamicton bed, middle sand bed, and upper sand bed and it is surmised that these four units are closely related in time. The lower sand bed (~1.5m), which overlies lower Miocene sediments and from which it is separated by the Ross Sea Unconformity, contains traces of recycled Miocene diatoms but is otherwise barren of biogenic material. The diamicton bed (~2.42 m) contains 21 species of benthic foraminifera, with assemblages consistently dominated by Cassidulinoides porrectus, Ammoelphidiella antarctica, Rosalina cf. globularis, Cibicides refulgens, and Ehrenbergina glabra. The overlying middle sand bed (~1.9 m) contains 13 species. with C. porrectus and E. glabra dominant and A. antarctica less common than in the underlying diamicton bed. The upper sand bed (~0.46 m) contains four species and very few tests. The diamicton bed and middle sand bed assemblages are considered to be near in situ thanatocoenoses; and sediments interpreted as marine in origin but influenced by hyposaline waters and nearby ice. Planktic taxa are absent, perhaps indicating the presence of tidewater glaciers, sea ice and/or hyposaline surface waters. The small assemblage in the upper sand bed is more problematic and may be recycled. On the basis of foraminifera in the diamicton and middle sand beds. LSU 2.2 is assigned to the Pliocene. The overlying diamicton in LSU 2.1 contains abundant Quaternary foraminifera.

Down-core distribution of live and dead benthic foraminifera in deep sea sediments from the Weddell Sea and the Californian continental borderland

Five short cores sub-sampled from box cores from three sites in the eastern Weddell Sea off Antarctica and in the eastern Pacific off southern California, covering a range in water depth from 500 to 2000 m, were analysed for the down-core distribution of live (stained with Rose Bengal) and dead benthic foraminifera. In the California continental borderland, Planulina ariminensis, Rosalina columbiensis and Trochammina spp. live attached to agglutinated polychaetes tubes that rise above the sedimentwater interface. Bolivina spissa lives exclusively in or on the uppermost sediment. Stained specimens of Chilostomella ovoidea are found down to 6 cm within the sediment and specimens of Globobulimina pacifica down to a maximum of 8 cm. Delta13C values of live G. pacifica decrease with increasing depth from the sediment surface down to 7 cm core depth, indicating that this infaunal species utilizes13C-depleted carbon from pore waters. In the dead, predominantly calcareous benthic forminiferal assemblage, selective dissolution of small delicate tests in the upper sediment column causes a continuous variation in species proportions. In the eastern Weddell Sea, the calcareous Bulimina aculeata lives in a carbonate corrosive environment exclusively in or on the uppermost sediment. The arenaceous Cribrostomoides subglobosum, Recurvoides contortus and some Reophax species are frequently found within the top 4 cm of the sediment, whereas stained specimens of Haplophragmoides bradyi, Glomospira charoides and Cribrostomoides wiesneri occur in maximum abundance below the uppermost 1.5 cm. Species proportions in the dead, predominantly arenaceous, benthic foraminiferal assemblage change in three distinct steps. The first change is caused by calcite dissolution at the sediment-water interface, the second coincides with the lower boundary of intense bioturbation, and the third results from the geochemical shift from oxidizing to reducing conditions below a compacted ash layer.