Cellular & molecular biology research最新文献

The cardiac mutant axolotl is an interesting model for studying heart development. The mutant gene results in a failure of heart cells to form organized myofibrils and as a consequence the heart fails to beat. Experiments have shown that mutant hearts can be "rescued" (i.e., turned into normally contracting organs) by the addition of RNA purified from conditioned media produced by normal embryonic anterior endoderm-mesoderm cultures. These corrected hearts form myofibrils of normal morphology. New advances in recombinant DNA technology applied to this system should provide significant insights into the regulatory mechanisms of myofibrillogenesis as well as the inductive processes related to the control of gene expression during embryonic heart development. In a broader biological sense, the use of gene c in axolotls is potentially capable of helping to solve major unanswered questions in modern biology related to the genetic regulation of differentiation in vertebrates.

The BCR gene is implicated in the development of Ph-positive leukemia through its fusion with the nonreceptor tyrosine kinase gene ABL. The normal 160 kDa Bcr protein has several functional domains, and recently one specific role for Bcr was established in the regulation of respiratory burst activity in white blood cells. Bcr expression levels are relatively constant throughout mouse development until adulthood in brain and in hematopoietic tissues, a pattern that is distinctly different from that of the functionally related n-chimerin gene. In the present study, RNA in situ hybridization was used to explore the normal cellular function of Bcr in rodent brain and hematopoietic organs. The data pinpoint the high bcr expression in the brain to the hippocampal pyramidal cell layer and the dentate gyrus, and to the piriform cortex and the olfactory nuclei, reflecting a potentially interesting function for Bcr in these highly specialized brain regions.

The 5 Kd (MW), retinoic acid responsive thymosin beta-10 protein is expressed at relatively high levels in embryonic tissues, and its mRNA is abundant in a variety of tumors and tumor cell lines. Recently this protein (together with other members of the same protein family) was found to be a major intracellular G-actin binding protein. In the present study, plasmid-driven overexpression of thymosin beta-10 gene results in increased susceptibility of permanently transfected fibroblasts to undergo apoptosis. Conversely, knockout of the endogenous gene via overexpression of the antisense mRNA inhibited cell death induced by TNF-alpha and calcium ionophore A23187. Differential expression of thymosin beta-10 influenced cell proliferation, cell morphology, and expression/distribution of the antiapoptotic protein bcl-2. The presence of increased cytoplasmic thymosin beta-10 precipitated significant disruption of phalloidin-stained actin stress fibers while knockout of thymosin expression promoted F-actin assembly. These and other observations suggest that thymosin beta-10 (a) plays a significant and possibly obligatory role in cellular processes controlling apoptosis possibly by acting as an actin-mediated tumor suppressor, (b) perhaps functions as a neoapoptotic influence during embryogenesis, and (c) may mediate some of the pro-apoptotic anticancer actions of retinoids.

HM-1 toxin produced by Hansenula mrakii kills sensitive Saccharomyces cerevisiae. We found that the budding cells and the cells that responded to mating factor were sensitive to HM-1 toxin. These findings indicate that the target sites of HM-1 toxin are developing buds and conjugating tubes. The in vitro activity of beta-1,3-glucan synthase solubilized and partially purified from S. cerevisiae membranes was inhibited by HM-1 toxin at a concentration (around 50 nM, 0.5 micrograms/ml) that coincided well with its minimum inhibitory concentration for the growth of yeast cells. These data indicate that the HM-1 toxin perturbs the synthesis of yeast cell walls by inhibiting the glucan synthesis occurring at a budding site or a conjugating tube, which results in cell lysis.

Using the human T-cell leukemia virus type I (HTLV-I) infected SLB-I T-cell line, we showed in this study that 5-d treatment with the maximal subtoxic 3-methylcholanthrene (3-MC) dose (0.25 microgram/ml), as well as with a 3-MC dose that inhibits 50% of the cell growth (5 micrograms/ml), profoundly increased the level of viral RNA. Exposure to these 3-MC doses for 5 d before transient transfection of HTLV-I LTR-CAT construct into these cells markedly stimulated CAT activity, indicating that 3-MC exerted its effect by a trans-acting mechanism. A similar stimulation was observed when this construct was transfected into 3-MC treated uninfected Jurkat cells, indicating that this trans-acting effect was independent of the viral tax protein. However, although the subtoxic 3-MC dose increased also the capacity of SLB-I cells to transmit the virus to normal peripheral blood lymphocytes in coculture, the toxic dose strongly reduced this capacity. No inhibition by this toxic dose was observed in the viral protein synthesis or processing nor in the final release of the virus from the cells. However, the virions released under the influence of this 3-MC dose were found to contain mainly the uncleaved gag precursor polypeptide and a low level of reverse transcriptase. Thus, the reduced virus transmission capacity of the host cells can be ascribed to this structural defect, which presumably lowered the viral infectivity.

In this review, a rationale is presented for how hypercholesterolemia, hypertension, diabetes mellitus, end-stage renal disease, renal dialysis, and prolonged stress can all lead to atherosclerosis, ischemic heart disease, and stroke. The data indicate that Mg deficiency caused either by poor diet and/or errors in Mg metabolism may be a missing link between diverse cardiovascular risk factors and atherosclerosis. Data from our laboratories and others indicate that reduction in extracellular and intracellular free Mg ions (Mg2+) can induce an entire array of pathophysiological phenomena known to be important in atherogenesis, that is, vasospasm, increased vascular reactivity, elevation in [Ca2+]i, formation of proinflammatory agents, oxygen radicals, platelet aggegation, reduction in cardiac bioenergetics, cardiac failure, oxidation of lipoproteins, gender-related modulation of endothelial-derived relaxing factor/NO, changes in membrane fatty acid saturation, changes in membrane plasmalogens and N-phospholipids (suggesting changes in intracellular phospholipid signals), and probably transcription factors.

Recently, Na+/Pi cotransport activity has been demonstrated in rat liver hepatocytes. Here, we report the isolation of two Na+/Pi cotransporter cDNAs (RNaPi-1a and RNaPi-1b) from a rat liver cDNA library. The two cDNAs have the same coding but different 5'-untranslated regions. The rat cDNAs encode a polypeptide of 465 amino acids, having 62% and 66% identity with the rabbit NaPi-1 and human kidney Na+/Pi cotransporter, respectively. Northern blot analysis showed that a RNaPi-1a--specific probe detected two major transcripts (2.3 and 1.8 kb), whereas a RNaPi-1b--specific probe hybridized with one transcript (1.8 kb) in rat kidney, liver, and hepatocytes in primary culture. Rat liver expressed much higher levels of RNaPi-1a than RNaPi-1b, whereas the converse was true for rat kidney. Low levels of RNaPi-1 mRNAs were also detected in rat heart, brain, and skeletal muscle. These findings indicate that there are at least two isoforms of RNaPi-1 transcripts expressed in liver and kidney and that the levels of expression of the RNaPi-1a and RNaPi-1b may be controlled by tissue-specific factors.

Rhodospirillum rubrum is a spiral anoxygenic photosynthetic bacterium that can exist under either aerobic or anaerobic conditions. The organism thrives in the presence of light or complete darkness and represents one of the oldest species of living organisms, possibly 2-3.5 billion years old. The success of this prokaryotic species may be attributed to the evolution of certain indole compounds that offer protection against life-threatening oxygen radicals produced by an evolutionary harsh environment. Melatonin, N-acetyl-5-methoxytryptamine, is an indolic highly conserved molecule that exists in protists, plants, and animals. This study was undertaken to determine the presence of an immunoreactive melatonin in the kingdom Monera and particularly in the photosynthetic bacterium, R. rubrum, under conditions of prolonged darkness or prolonged light. Immunoreactive melatonin was measured during both the extended day and extended night. Significantly more melatonin was observed during the scotophase than the photophase. This study marks the first demonstration of melatonin in a bacterium. The high level of melatonin observed in bacteria may provide on-site protection of bacterial DNA against free radical attack.

Serine acetyltransferase, a key enzyme in the L-cysteine biosynthetic pathway of sulfate assimilating organisms, catalyzes the formation of O-acetylserine, the immediate precursor of L-cysteine. In higher plants, it is thought that sulfur assimilation occurs primarily in leaf chloroplasts; however, serine acetyltransferase is not localized exclusively in this tissue and organelle. At least three genes for serine acetyltransferase have been identified in the higher plant Arabidopsis thaliana. Reported here is a cDNA corresponding to one of these genes, SAT1, a 1,079 bp clone with an open reading frame predicted to encode a 34-kDa protein that is able to functionally complement a serine acetyltransferase mutant strain of Escherichia coli. The predicted amino acid sequence of SAT1 shows significant homology with bacterial serine acetyltransferases. SAT1, expressed as a recombinant protein, shows serine acetyltransferase enzyme activity and cross-reacts with an antibody against the homologous E. coli enzyme. The first 40 amino acids of the SAT1 polypeptide resembles a plastid transit peptide, but the polypeptide is probably not plastid localized. Genomic DNA blot analysis of A. thaliana showed that SAT1 is a single copy gene and RNA blot analysis revealed that SAT1 is expressed in both leaves and roots.