Plant gene responses to frequency-specific sound signals

We identified a set of sound-responsive genes in plants using a sound-treated subtractive library and demonstrated sound regulation through mRNA expression analyses. Under both light and dark conditions, sound up-regulated expression of rbcS and ald. These are also light-responsive genes and these results suggest that sound could represent an alternative to light as a gene regulator. Ald mRNA expression increased significantly with treatment at 125 and 250 Hz, whereas levels decreased significantly with treatment at 50 Hz, indicating a frequency-specific response. To investigate whether the ald promoter responds to sound, we generated transgenic rice plants harboring a chimeric gene comprising a fusion of the ald promoter and GUS reporter. In three independent transgenic lines treated with 50 or 250 Hz for 4 h, GUS mRNA expression was up-regulated at 250 Hz, but down-regulated at 50 Hz. Thus, the sound-responsive mRNA expression pattern observed for the ald promoter correlated closely with that of ald, suggesting that the 1,506 bp ald promoter is sound-responsive. Therefore, we propose that in transgenic plants, specific frequencies of sound treatment could be used to regulate the expression of any gene fused to the ald promoter.     

Impact of the TCR Signal on Regulatory T Cell Homeostasis,Function, and Trafficking

Signaling through the T cell antigen receptor (TCR) is important for the homeostasis of naïve and memory CD4⁺ T cells. The significance of TCR signaling in regulatory T (Treg) cells has not been systematically addressed. Using an Ox40-cre allele that is prominently expressed in Treg cells, and a conditional null allele of the gene encoding p56^Lck, we have examined the importance of TCR signaling in Treg cells. Inactivation of p56^Lck resulted in abnormal Treg homeostasis characterized by impaired turnover, preferential redistribution to the lymph nodes, loss of suppressive function, and striking changes in gene expression. Abnormal Treg cell homeostasis and function did not reflect the involvement of p56^Lck in CD4 function because these effects were not observed when CD4 expression was inactivated by Ox40-cre.The results make clear multiple aspects of Treg cell homeostasis and phenotype that are dependent on a sustained capacity to signal through the TCR.     

Genetic polymorphism, mapping, and characterization of isocitrate dehydrogenase in Anopheles quadrimaculatus

The isocitrate dehydrogenase-2 (Idh-2) locus of Anopheles quadrimaculatus was analyzed genetically and the enzyme was characterized physiochemically. Three-point testcrosses involving chromosome 3 markers showed that in female hybrids the gene sequence and the map distances are: nonstripe (st)--6.8--Idh-2--43.5--short antenna (Sa). Reduced recombination frequencies were observed in male hybrids (st--3.4--Idh-2--25.5--Sa). Idh-2 activity gradually increases during development and reaches a peak intensity in adults. Maximum enzyme activity of Idh-2 was obtained at pH 7.5. One-minute heat treatment at 50 degrees C caused about 50 percent reduction of IDH-2. Ethylene diamine tetraacetic acid (EDTA 5 mM) and p-chloromercuribenzoate (pCMB 10(-5)M) caused complete loss of activity of IDH-2, but pretreatment of the enzyme in situ with mercaptoethanol protected the activity of allozymes from inhibition by pCMB treatment.     

Specific antigens inhibit plasmacytoma cells bearing phosphorylcholine-binding myeloma proteins.

The influence of a synthetic adjuvant active glycopeptide, N-acetylmuramyl-l-alanyl-d-isoglutamine (MDP), and of some of its analogs on the in vitro immune response to sheep red blood cells was studied using Mishell and Dutton in vitro stimulation system. When MDP and adjuvant active analogs were incubated with normal spleen cells, increased cell recovery was observed after 3 or 4 days of culture, showing a good correlation between the adjuvant activity in vivo and the enhancement of cell viability in vitro. The analogs which were found to have an adjuvant activity in vivo were equally effective in stimulating in vitro both the background hemolytic PFC and the immune response to sheep red blood cells. However, those which were inactive in vivo were effective in vitro but only at high concentration levels.     

Multiple forms of alkaline phosphatase in mouse preimplantation embryos

Mouse embryos were extracted with 0.5% Triton X-100 and subjected to cellulose acetate electrophoresis. In fertilized eggs, two forms of alkaline phosphatase (ALP), a slow-moving form and a fast-moving form, were observed. As cleavage proceeded, the fast-moving form disappeared, and the slow-moving form, the mobility of which was similar to that of the slow-moving form of the kidney, became gradually dominant up to the blastocyst stage (named 'embryonic' form). With blastulation, another fast-moving form showing a similar mobility to the lung ALP began to appear in blastocysts and showed a transient dominance in hatched blastocysts. After implantation, both the embryonic form and the fast-moving form gradually faded, and were eventually replaced by the new form, which may be named 'fetal form' in Day 7 embryos. These results clearly demonstrated that ALP activity does exist in embryos at all stages of preimplantation development. Moreover, the changes in multiple forms of ALP correlated with embryonic development may suggest that these multiple forms may have differential roles in the process of early development.     

Effect pf protease inhibitors on DNA degradation in gamma-irradiated rat thymocytes

Protease inhibitors, antipain and pepstatin, decrease the postirradiation degradation of DNA in gamma-irradiated rat thymocytes. It is suggested that a protease-dependent process of induction of DNA breaks takes place in irradiated cells.     

Malonyl-CoA decarboxylase from Mycobacterium tuberculosis and Pseudomonas fluorescens.

Malonyl-CoA decarboxylase was partially purified (nearly 1000-fold) from Mycobacterium tuberculosis H37Ra by ammonium sulfate precipitation, gel filtration with Sepharose 6B, and chromatography on DEAE Sephacel, carboxymethyl-Sephadex, and NADP-agarose. Polyacrylamide gel electrophoresis showed a major band (60–70%), which contained the enzymatic activity, and a minor band which had no decarboxylase activity. The molecular weight of the enzyme was 44,000, and the PI and pH optimum were 6.7 and 5.5, respectively. The enzyme showed a typical Michaelis-Menten substrate saturation, with an apparent K_m and V of 0.2 mm and 3.85 μmol/min/mg, respectively. It catalyzed decarboxylation of methylmalonyl-CoA only at 5% of the rate observed with malonyl-CoA, whereas malonic acid and succinyl-CoA were not decarboxylated. Antibodies prepared against malonyl-CoA decarboxylase from the uropygial glands of goose and rat liver mitochondria did not inhibit the bacterial enzyme. Avidin did not inhibit the enzyme suggesting that biotin was not involved in the reaction. Thiol-directed reagents inhibited the enzyme as did CoA, acetyl-CoA, propionyl-CoA, methylmalonyl-CoA, and succinyl-CoA. Malonyl-CoA decarboxylase was also partially purified from malonate-grown Pseudomonas fluorescens. The molecular weight of this enzyme was 56,000 and the pH optimum and apparent K_m were 5.5 and 1 mm, respectively. Unlike the mycobacterial enzyme, this enzyme was insensitive to p-hydroxymercuribenzoate, acetyl-CoA, and propionyl-CoA, and it was less sensitive to inhibition by succinyl-CoA and CoA than the mycobacterial enzyme. The size and properties of the two bacterial enzymes suggest that these are quite unlike the mammalian and avian enzymes and that they constitute a different class of malonyl-CoA decarboxylases.     

Protein disulphide isomerase is required for signal peptide peptidase-mediated protein degradation

The human cytomegalovirus glycoprotein US2 induces dislocation of MHC class I heavy chains from the endoplasmic reticulum (ER) into the cytosol and targets them for proteasomal degradation. Signal peptide peptidase (SPP) has been shown to be integral for US2-induced dislocation of MHC class I heavy chains although its mechanism of action remains poorly understood. Here, we show that knockdown of protein disulphide isomerase (PDI) by RNA-mediated interference inhibited the degradation of MHC class I molecules catalysed by US2 but not by its functional homolog US11. Overexpression of the substrate-binding mutant of PDI, but not the catalytically inactive mutant, dominant-negatively inhibited US2-mediated dislocation of MHC class I molecules by preventing their release from US2. Furthermore, PDI associated with SPP independently of US2 and knockdown of PDI inhibited SPP-mediated degradation of CD3δ but not Derlin-1-dependent degradation of CFTR DeltaF508. Together, our data suggest that PDI is a component of the SPP-mediated ER-associated degradation machinery.