Cold acclimation and cold-regulated gene expression in ABA mutants of Arabidopsis thaliana

We have examined the cold-induced enhancement of freezing tolerance and expression of cold-regulated (cor) genes in Arabidopsis thaliana (L.) Heynh (Landsberg erecta) and abscisic acid (ABA)-deficient (aba) and ABA-insensitive (abi) mutants derived from it. The results indicate that the abi mutations had no apparent effect on freezing tolerance, while the aba mutations did: cold-acclimated aba mutants were markedly impaired in freezing tolerance compared to wild-type plants. In addition, it was observed that non-frozen leaves from both control and cold-treated aba mutant plants were more ion-leaky than those from corresponding wild-type plants. These data are consistent with previous observations indicating that ABA levels can affect freezing tolerance. Whether ABA has a direct role in the enhancement of freezing tolerance that occurs during cold acclimation, however, is uncertain. Several studies have suggested that ABA might mediate certain changes in gene expression that occur during cold acclimation. Our data indicate that the ABA-induced expression of three ABA-regulated Arabidopsis cor genes was unaffected in the abi2, abi3, and aba-1 mutants, but was dramatically impaired in the abi1 mutant. Cold-regulated expression of all three cor genes, however, was nearly the same in wild-type and abi1 mutant plants. These data suggest that the cold-regulated and ABA-regulated expression of the three cor genes may be mediated through independent control mechanisms.     

Immunochemical characterisation of parathyroid hormone-related protein from tumor and non-tumor cells

The molecular forms of parathyroid hormone-related protein (PTHRP) in conditioned media from the BEN human lung cancer cell line, rat parathyroid cells (PT-r) and human keratinocytes were studied by gel-filtraton chromatography with assay of PTHRP by immunoassays and bioassay. Immunoreactivity (1–86 and 1–34) and bioactivity (1–34) in conditioned media eluted as a coincident major peak (approx. molecular mass 19–22 kDa) and there was evidence of amino-terminal species in the molecular mass range 10–16 kDa in BEN and keratinocyte media. Western blotting of PTHRP affinity purified by monoclonal antibodies directed at regions 1–34 or 37–67, identified a major species in all cell cytosols and media with an apparent molecular mass of 24–25 kDa, consistently slighty larger than recombinant PTHRP(1–141) (mobility of 21 kDa) which may represent an intact or native form of PTHRP. Additional amino-terminal species were identified in medium from keratinocytes (16 and 7 kDa), BEN cells (18 and 14 kDa) and PT-R cells (17 kDa), suggesting that processing occurs at the C-terminus and within the mid-region to form a range of amino-terminal fragments.     

Pneumocystis carinii shows DNA homology with the ustomycetous red yeast fungi

Pneumocystis carinii causes life-threatening pneumonia in T-lymphocyte-immunodeficient subjects in transplant and oncology units or with acquired immune deficiency syndrome (AIDS). Recent DNA homology studies show P. carinii to be a fungus. To investigate the biology and epidemiology of this parasite further, we elected to determine for it a more precise taxonomic assignment within the fungal kingdom. We screened a wide range of organisms representing the major orders of fungi using DNA amplification and subsequently sequenced a portion of the mitochondrial gene encoding the large subunit ribosomal RNA. Our data show that the opportunistic pulmonary pathogen P. carinii is closely related to the ustomycetous red yeast fungi, a group which includes organisms that are extensively distributed throughout the environment and which release many widely dispersed airborne spores.     

Identification and characterization of an {alpha}-mannosidase from Trypanosoma cruzi

In this report we describe the first purification and characterizationof the acid -mannosidase from the human parasite Trypanosomacruzi. The purified enzyme exhibited a native mol. wt of 240000 Da and is apparently composed of four identical subunitsof mol. wt 58 000 Da. Each of the four subunits contains oneN-linked high-mannose-type oligosaccharide. The -mannosidaseexhibited a pH optimum of 3.5 and a pI of 5.9. This low pH optimumand the ability of swainsonine to inhibit its activity suggestthat the -mannosidase is a lysosomal enzyme. Antibodies againstthe T.cruzi enzyme did not react with mammalian lysosomal -mannosidaseand, conversely, antibody against a rat lysosomal -mannosidasedid not react with the T.cruzi enzyme. Thus, the T.cruzi enzymeappears to be distinct from its mammalian counterpart. -mannosidase lysosomal enzyme Trypanosoma cruzi     

The adenine nucleotide translocator of higher plants is synthesized as a large precursor that is processed upon import into mitochondria.

Two maize genes and cDNAs encoding the mitochondrial adenine nucleotide translocator (ANT), a nuclear-encoded inner mitochondrial membrane carrier protein, have previously been isolated in this laboratory. Sequence analysis revealed the existence of much longer open reading frames than the corresponding fungal and mammalian ANT genes. Potato ANT cDNAs have subsequently been isolated and sequenced and alignment of the deduced plant amino acid sequences with the equivalent fungal and mammalian polypeptides indicated that the plant proteins contain N-terminal extensions. When the plant cDNA clones are expressed in vitro they direct the synthesis of precursor proteins that are specifically processed at the N-terminus upon import into isolated mitochondria. N-terminal amino acid sequence data obtained from the native proteins purified from both maize and potato mitochondria has allowed identification of the putative processing sites. Further import analysis has shown that two distinct regions of the maize precursor protein contain targeting information, the 97 amino acids at the N-terminus and the 267 C-terminal amino acids. This is the first report that provides experimental evidence that the adenine nucleotide translocator of higher plants is synthesized as a large precursor protein that is specifically cleaved upon import into mitochondria. Import of ANT into higher plant mitochondria therefore appears to be different to the corresponding process in fungal and mammalian systems where targeting of ANT to mitochondria is mediated by internal signals and there is no N-terminal processing.     

Functional levels of mitochondrial anion transport proteins in non-insulin-dependent diabetes mellitus

The effect of non-insulin-dependent diabetes mellitus (i.e., NIDDM; type 2 diabetes) on the levels of functional mitochondrial anion transport proteins has been determined utilizing a chemically-induced neonatal model of NIDDM. We hypothesized that moderate insulin deficiency exacerbated by the insulin resistance, which is characteristic of NIDDM, would cause changes in mitochondrial anion transporter function that were similar to those we have previously shown to occur in insulin-dependent diabetes mellitus (i.e., IDDM; type 1 diabetes) (Arch. Biochem. Biophys. 280: 181–191, 1990). Our experimental approach consisted of the extraction of the pyruvate, dicarboxylate and citrate transport proteins from the mitochondrial inner membrane with Triton X-114 using rat liver mitoplasts (prepared from diabetic and control animals) as the starting material, followed by the functional reconstitution of each transporter in a proteoliposomal system. This strategy permitted the quantification of the functional levels of these three transporters in the absence of the complications that arise when such measurements are carried out with intact mitochondria (or mitoplasts). We found that experimental NIDDM did not cause significant changes in the extractable and reconstitutable specific (and total) transport activities of the pyruvate, dicarboxylate, and citrate transporters. These results are in marked contrast to our previous findings obtained using rats with IDDM and negated our hypothesis. The present results, in combination with our earlier findings, allow us to conclude that insulin plays an important role in the regulation of mitochondrial anion transporter function. Accordingly, in this model of NIDDM, where the level of insulin is not profoundly deficient, transporter function is unaltered, whereas in IDDM, where a profound insulinopenia exists, transporter function is altered. Furthermore, the present studies suggest that in the neonatal model of NIDDM the three mitochondrial transporters investigated are neither affected by, nor are they the sites of the well documented hepatic post-receptor insulin resistance which is characteristic of this disease.     

Pastoral dairy marketing and household wealth interactions and their implications for calves and humans in Ethiopia

Surveys of pastoral households in a semi-nomadic Borana community during 1987–1988 were used to test the hypothesis that poorer families living closest to a market town would be most affected by the enhanced opportunity to sell dairy products, which would intensify competition between people and calves for milk and have negative implications for calf management. These poorer families indeed reported the highest rates of milk offtake per cow, and the milk increment was probably sold to purchase more grain for human consumption at the expense of milk intake for the calf. Consequently, this strategy may increase the susceptibility of malnourished calves to disease, especially those from lower-producing dams. Benefits of improved human energy intake from grain and retention of livestock capital must be weighed against risks of calf death and possible malnutrition of people from milk restriction when assessing dairy marketing trade-offs that are most acute for the poor. Opportunity to sell dairy products at favorable terms of trade helps the poorest people survive, and their risks could be mitigated by policies that facilitate grain marketing in the rangelands and interventions that improve calf feeding management, diversify human diets, and create alternative opportunities for women to generate income. The households postulated to be most at risk were identified from a complex, but logical, interaction among factors of distance to market, household wealth, and the quality of milking cows held. This indicates that targeting such needy groups for development assistance may require a more detailed and interdisciplinary analysis of production systems than is commonly practiced.