Anisotropic sensor and memory device with a ferromagnetic tunnel barrier as the only magnetic element.

Multiple spin functionalities are probed on Pt/La2Co0.8Mn1.2O6/Nb:SrTiO3, a device composed by a ferromagnetic insulating barrier sandwiched between non-magnetic electrodes. Uniquely, La2Co0.8Mn1.2O6 thin films present strong perpendicular magnetic anisotropy of magnetocrystalline origin, property of major interest for spintronics. The junction has an estimated spin-filtering efficiency of 99.7% and tunneling anisotropic magnetoresistance (TAMR) values up to 30% at low temperatures. This remarkable angular dependence of the magnetoresistance is associated with the magnetic anisotropy whose origin lies in the large spin-orbit interaction of Co2+ which is additionally tuned by the strain of the crystal lattice. Furthermore, we found that the junction can operate as an electrically readable magnetic memory device. The findings of this work demonstrate that a single ferromagnetic insulating barrier with strong magnetocrystalline anisotropy is sufficient for realizing sensor and memory functionalities in a tunneling device based on TAMR.

The Pzh1 protein phosphatase and the Spm1 protein kinase are involved in the regulation of the plasma membrane H+-ATPase in fission yeast.

We have previously shown that the mutation of the Schizosaccharomyces pombe PPZ-like protein phosphatase encoded by the gene pzh1+ results in increased tolerance to sodium and in hypersensitivity to potassium ions. A similar phenotype has also been reported for deletants in the spm1/pmk1 gene, encoding a mitogen-activated protein (MAP) kinase. We have found that the sodium tolerance phenotype of pzh1 deletants is stronger than that of spm1 mutants, and both effects are additive. Therefore, most probably both gene products mediate different pathways on sodium tolerance. In our hands, mutation of the kinase does not alter the tolerance to potassium, but it yields cells more tolerant to magnesium ions. While in budding yeast the mutations are synthetically lethal, fission yeast cells lacking both the phosphatase and the kinase genes are viable. Interestingly, their ability to export H+ to the medium is greatly impaired (although not that of pzh1 or spm1 single mutants). We have observed that, although the amount of the H+-ATPase in the plasma membrane is not altered, the activity of the enzyme is lower than normal and cannot be induced by glucose. These observations suggest that the activity of the H+-ATPase in fission yeast might be regulated by phospho-dephosphorylation mechanisms that might involve the pzh1+ and spm1+ gene products.

The presence of enhancers adjacent to the Ac promoter increases the abundance of transposase mRNA and alters the timing of Ds excision in Arabidopsis.

Two copies of domain B of the CaMV 35S promoter were inserted ca. 300 bp upstream of the transcriptional start site of the Ac transposase gene. Four independent Arabidopsis transformants containing this fusion (35SenhAc::TPase) were made and the abundance of transposase mRNA in each of them was determined. The presence of the enhancers increased the abundance of the transposase mRNA by about 12-fold compared to that found in plants containing an Ac promoter fusion to the transposase gene (Ac::TPase). Hybrid plants carrying 35SenhAc::TPase and a Ds element inserted in a streptomycin phosphotransferase (SPT) gene were constructed and the frequency with which Ds excision occurred in the developing cotyledons was measured. Moreover, the number of progeny of these hybrid plants which inherited an SPT gene activated by Ds excision was studied in individual F2 families. Those derived from 35SenhAc::TPase often contained higher proportions of streptomycin-resistant (strepR) F2 progeny than those derived from Ac::TPase. These high frequencies of strepR seedlings were comparable to those previously detected after activation of Ds by a CaMV 35S promoter fusion to transposase (35S::TPase), but occurred in fewer families. The higher frequency with which this occurred in families derived from 35SenhAc::TPase compared to Ac::TPase suggests that the presence of enhancers adjacent to the native Ac promoter can influence transposase gene expression, and in this case often results in earlier excision of Ds during plant development.

A unitary basis for different Hairy-wing mutations of Drosophila melanogaster.

Hairy-wing (Hw) mutations are caused by modifications of the achaete-scute complex (AS-C) which promote development of extra sensory organs on the cuticle of Drosophila melanogaster. We show that the extreme Hw49c allele contains an inversion with a breakpoint within the AS-C, while the weak Hw685 allele is associated with a terminal deletion of the X chromosome which removes the achaete region of the AS-C. In both cases, foreign DNA in contact with the breakpoints presumably enhances expression of AS-C genes. Overexpression of achaete (T5) or scute alpha (T4) genes was previously found in Hw mutants associated with insertions of transposable elements (Hw1, HwBS and HwUa, Campuzano et al., 1986). In situ hybridizations to Hw49c and Hw1 larval sections show that the overexpression causes an abnormally generalized distribution of T4 and/or T5 transcripts in imaginal discs. Such distribution correlates with development of extra sensory organs in ectopic positions. We also show that in Hw685 a moderate overexpression of the T4 gene largely replaces the absence of the T5 gene in the development of the notum chaetae pattern. We propose that overexpression of T4/T5 genes in normal or ectopic positions is at the basis of the Hw effect.

Isolation and structural characterization of a cDNA encoding Arabidopsis thaliana 3-hydroxy-3-methylglutaryl coenzyme A reductase.

The enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase (EC 1.1.1.34) catalyses the synthesis of mevalonate, the specific precursor of all isoprenoid compounds present in plants. We have characterized two overlapping cDNA clones that encompass the entire transcription unit of an HMG-CoA reductase gene from Arabidopsis thaliana. The transcription product has an upstream non-coding sequence of 70 nucleotides preceding an open reading frame of 1776 bases and a 3' untranslated region in which two alternative polyadenylation sites have been found. The analysis of the nucleotide sequence reveals that the cDNA encodes a polypeptide of 592 residues with a molecular mass of 63,605 Da. The hydropathy profile of the protein indicates the presence of two highly hydrophobic domains near the N-terminus. A sequence of 407 amino acids corresponding to the C-terminal part of the protein (residues 172-579), which presumably contains the catalytic site, shows a high level of similarity to the region containing the catalytic site of the hamster, human, yeast and Drosophila enzymes. The N-terminal domain contains two putative membrane-spanning regions, in contrast to the enzyme from other organisms which has seven trans-membrane regions. A. thaliana contains two different HMG-CoA reductase genes (HMG1 and HMG2), as estimated by gene cloning and Southern blot analysis. Northern blot analysis reveals a single transcript of 2.4 kb in leaves and seedlings, which presumably corresponds to the expression of the HMG1 gene.

Regulation of salt tolerance in fission yeast by a protein-phosphatase-Z-like Ser/Thr protein phosphatase.

In the yeast Saccharomyces cerevisiae, Na+ efflux is mediated by the Ena1 ATPase, and the expression of the ENA1 gene is regulated by the Ppz1 and Ppz2 Ser/Thr protein phosphatases. On the contrary, in the fission yeast Schizosaccharomyces pombe, effective output of Na+ is attributed to the H+/Na+ antiporter encoded by the sod2 gene. We have isolated a S. pombe gene (pzh1) that encodes a 515-amino-acid protein that is 78% identical, from residue 193 to the COOH terminus, to the PPZ1 and PPZ2 gene products. Bacterially expressed Pzh1p shows enzymatic characteristics virtually identical to those of recombinant Ppz1p. When expressed in high-copy number from the PPZ1 promoter, the pzh1 ORF rescues the caffeine-induced lytic defect and slightly decreases the high salt tolerance of S. cerevisiae ppz1delta mutants. Disruption of pzh1 yields viable S. pombe cells and has virtually no effect on tolerance to caffeine or osmotic stress, but it renders the cells highly tolerant to Na+ and Li+, and hypersensitive to K+. Although lack of pzh1 results in a 2-3-fold increase in sod2 mRNA, the pzh1 mutation significantly increases salt tolerance in the absence of the sod2 gene, suggesting that the phosphatase also regulates a Sod2-independent mechanism. Therefore, the finding of a PPZ-like protein phosphatase involved in the regulation of salt tolerance in fission yeast reveals unexpected aspects of cation homeostasis in this organism.

Arabidopsis thaliana contains two differentially expressed 3-hydroxy-3-methylglutaryl-CoA reductase genes, which encode microsomal forms of the enzyme.

The enzyme 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR; EC 1.1.1.34) catalyzes the first rate-limiting step in plant isoprenoid biosynthesis. Arabidopsis thaliana contains two genes, HMG1 and HMG2, that encode HMGR. We have cloned these two genes and analyzed their structure and expression. HMG1 and HMG2 consist of four exons and three small introns that interrupt the coding sequence at equivalent positions. The two genes share sequence similarity in the coding regions but not in the 5'- or 3'-flanking regions. HMG1 mRNA is detected in all tissues, whereas the presence of HMG2 mRNA is restricted to young seedlings, roots, and inflorescences. The similarity between the two encoded proteins (HMGR1 and HMGR2) is restricted to the regions corresponding to the membrane and the catalytic domains. Arabidopsis HMGR2 represents a divergent form of the enzyme that has no counterpart among plant HMGRs characterized so far. By using a coupled in vitro transcription-translation assay, we show that both HMGR1 and HMGR2 are cotranslationally inserted into endoplasmic reticulum-derived microsomal membranes. Our results suggest that the endoplasmic reticulum is the only cell compartment for the targeting of HMGR in Arabidopsis and support the hypothesis that in higher plants the formation of mevalonate occurs solely in the cytosol.

Excess function hairy-wing mutations caused by gypsy and copia insertions within structural genes of the achaete-scute locus of Drosophila.

Hairy-wing (Hw) mutations cause the differentiation of extra chaetes on the cuticle of Drosophila. They are associated with modifications of the achaete-scute complex that consist, in the mutants studied, of insertions of the transposable elements gypsy (Hw1, HwBS) or copia (HwUa). gypsy and copia are inserted in achaete and scute transcribed regions, respectively. Transcription of the insertion-split genes starts at the normal site but terminates within the transposable element sequences. The RNA truncated within gypsy is 5-20 times more abundant than its homolog in wild-type flies. The abundance is reduced in Hw1 revertants and Hw1 stocks carrying su(Hw) mutations. These and other data suggest that the excess function phenotypes of Hw mutations are generated by an increase in achaete or scute transcripts.

Sequence analysis of a 13.4 kbp fragment from the left arm of chromosome XV reveals a malate dehydrogenase gene, a putative Ser/Thr protein kinase, the ribosomal L25 gene and four new open reading frames.

A 13421 bp fragment located near the left telomere of chromosome XV (cosmid pEOA461) has been sequenced. Seven non-overlapping open reading frames (ORFs) encoding polypeptides longer than 100 residues have been found (AOB859, AOC184, AOE375, AOX142i, AOE423, AOA476 and AOE433). An additional ORF (AOE131) is found within AOA476. Three of them (AOC184, AOA476 and AOE433) show no remarkable identity with proteins deposited in the data banks. ORF AOB859 is quite similar to a hypothetical yeast protein of similar size located in chromosome VI, particularly within the C-terminal half. AOE375 encodes a new member of the glycogen synthase kinase-3 subfamily of Ser/Thr protein kinases. AOX142i is the gene encoding the previously described ribosomal protein L25. AOE423 codes for a protein virtually identical to the MDH2 malate dehydrogenase isozyme. However, our DNA sequence shows a single one-base insertion upstream of the reported initiating codon. This would produce a larger ORF by extending 46 residues the N-terminus of the protein. The existence of this insertion has been confirmed in three different yeast strains, including FY1679.

Elevated levels of Activator transposase mRNA are associated with high frequencies of Dissociation excision in Arabidopsis.

The Activator (Ac) element of maize is active at a low frequency in Arabidopsis. To determine whether this is due to poor expression of the Ac transposase gene, we obtained and studied 19 Arabidopsis transformants containing fusions of the octopine synthase (ocs), nopaline synthase (nos), cauliflower mosaic virus (CaMV) 35S, or Ac promoters to the transposase open reading frame. These transformants were examined both for their ability to drive excision of a Dissociation (Ds) element from a streptomycin resistance gene and for the abundance of the transposase mRNA. Most transformants containing the CaMV 35S fusion have high levels of transposase transcript and drive high frequencies of somatic and germinal excision. These results demonstrated that Arabidopsis contains all of the host functions required for high frequency excision of Ds. Moreover, transposase mRNA abundance varied about 1000-fold among our transformants; this variation enabled us to demonstrate that for the Ac, ocs, and CaMV 35S fusion, raising the mRNA level is closely correlated with increasing excision frequency. We discuss our data in relation to the behavior of Ac in Arabidopsis, maize, and tobacco.

Nanoscale multiphase separation at La(2/3)Ca(1/3)MnO3/SrTiO3 interfaces.

55Mn nuclear magnetic resonance experiments are reported on a series of fully strained epitaxial La(2/3)Ca(1/3)MnO3 thin films on SrTiO3. We have found evidence of multiple phase segregation into ferromagnetic metallic and nonmetallic regions as well as regions that are nonferromagnetic and insulating. These insulating regions are mainly located close to interfaces and may have a significant impact on the performance of spin-tunnel devices. As a result of phase segregation, the ferromagnetic coupling within the metallic regions is depressed. This accounts for the reduction of the Curie temperature and conductivity in nanometric thin films.

The Schizosaccharomyces pombe Pzh1 protein phosphatase regulates Na+ ion influx in a Trk1-independent fashion.

We have previously shown that fission yeast encodes a PPZ-like phosphatase, designated Pzhl, which is an important determinant of cation homeostasis. pzh1 delta mutants display increased tolerance to Na+ ions, but they are hypersensitive to KC1 [Balcells, L., Gómez, N., Casamayor, A., Clotet, J. & Ariño, J. (1997) Eur. J. Biochem. 250, 476-483]. We have immunodetected Pzh1 in yeast extracts and found that this phosphatase is largely associated with particulate fractions. Cells defective in Pzh1 do not show altered efflux of Na+ or Li+ ions, but they accumulate these cations more slowly than wild-type cells. K+ ion content of pzh1 delta cells is about twice that of wild-type cells, and this can be explained by decreased efflux of K+. Therefore, Pzh1 may regulate both Na+ influx and K+ efflux in fission yeast. To test the possible relationship between K+ uptake, Na+ tolerance and Pzh1 function, we deleted the trk1+ gene, which encodes a putative high-affinity transporter of K+ ions. trkl delta mutants grew well even at relatively low concentrations of KCl and did not show significantly altered content or influx of K+ ions. However, they showed a Na(+)-sensitive phenotype which was greatly intensified by deletion of the sod2+ gene (which encodes the major determinant for efflux of Na+ ions), and clearly ameliorated by deletion of the pzh1 phosphatase, as well as by moderate concentrations of KCl in the medium. These results suggest that Trk1 does not mediate the effect of Pzh1 on NaCl tolerance and that fission yeast contains efficient systems, other than Trk1, for uptake of K+ ions.

Sequence analysis of a 12 801 bp fragment of the left arm of yeast chromosome XV containing a putative 6-phosphofructo-2-kinase gene, a gene for a possible glycophospholipid-anchored surface protein and six other open reading frames.

The DNA sequence of a 12,801 bp fragment located near the left telomere of chromosome XV has been determined. Sequence analysis reveals eight open reading frames (ORFs) encoding polypeptides larger than 100 residues. ORFs AOE129 and AOAA121 are in opposite strands and they overlap at their 3' ends. AOE397 has similarity with phosphofructokinase genes from other organisms and may code for a second 6-phosphofructo-2-kinase of Saccharomyces cerevisiae. Sequence of AOA471 shows significant similarity with yeast genes coding for glycophospholipid-containing proteins. AOD1341 would code for a 1341 amino acids long protein with a predicted ATP/GTP-binding site and a transmembrane domain.

Cloning and characterization of the Arabidopsis thaliana SQS1 gene encoding squalene synthase--involvement of the C-terminal region of the enzyme in the channeling of squalene through the sterol pathway.

Squalene synthase (SQS) catalyzes the first committed step of the sterol biosynthetic pathway. A full-length Arabidopsis thaliana SQS cDNA has been isolated by combining library screening and PCR-based approaches. Arabidopsis SQS is encoded by a small gene family of two genes (SQS1 and SQS2) which are organized in a tandem array. SQS1 and SQS2 have an identical organization with regard to intron positions and exon sizes and encode SQS isoforms showing a high level of sequence conservation (79% identity and 88% similarity). The isolated cDNA has been assigned to the SQS1 gene product, SQS1. RNA blot analysis has shown that the 1.6-kb SQS1 mRNA is detected in all plant tissues analyzed (inflorescenses, leaves, stems and roots) although the transcript is especially abundant in roots. Arabidopsis SQS1 isoform is unable to complement the SQS-defective Saccharomyces cerevisiae strain 5302, although SQS activity was detected in the microsomal fraction of the transformed yeast strain. However, a chimeric SQS resulting from the replacement of the 66 C-terminal residues of the Arabidopsis enzyme by the 111 C-terminal residues of the Schizosaccharomyces pombe enzyme was able to confer ergosterol prototrophy to strain 5302. Labeling studies using [3H]farnesyl-P2 and microsomal fractions obtained from yeast strains expressing either Arabidopsis SQS1 or chimeric Arabidopsis/S. pombe SQS derivatives indicated that the C-terminal region of the enzyme is involved in the channeling of squalene through the yeast sterol pathway.