Two new scorpion toxins that target voltage-gated Ca2+ and Na+ channels

This report describes the isolation, primary structure determination, and functional characterization of two similar toxins from the scorpion Parabuthus granulatus named kurtoxin-like I and II (KLI and KLII, respectively). KLII from P. granulatus is identical to kurtoxin from Parabuthus transvaalicus (a 63 amino-acid long toxin) whereas KLI is a new peptide containing 62 amino acid residues closely packed by four disulfide bridges with a molecular mass of 7244. Functional assays showed that both toxins, KLI and kurtoxin from P. granulatus, potently inhibit native voltage-gated T-type Ca(2+) channel activity in mouse male germ cells. In addition, KLI was shown to significantly affect the gating mechanisms of recombinant Na(+) channels and weakly block alpha(1)3.3Ca(V) channels expressed in Xenopus oocytes. KLI and kurtoxin from P. granulatus represent new probes to study the role of ion channels in germ cells, as well as in cardiac and neural tissue.     

Symplastic continuity between companion cells and the translocation stream: long-distance transport is controlled by retention and retrieval mechanisms in the phloem

Substantial symplastic continuity appears to exist between companion cells (CCs) and sieve elements of the phloem, which suggests that small solutes within the CC are subject to indiscriminate long-distance transport via the translocation stream. To test this hypothesis, the distributions of exotic and endogenous solutes synthesized in the CCs of minor veins were studied. Octopine, a charged molecule derived from arginine and pyruvate, was efficiently transported through the phloem but was also transferred in substantial amounts to the apoplast, and presumably other non-phloem compartments. The disaccharide galactinol also accumulated in non-phloem compartments, but long-distance transport was limited. Conversely, sucrose, raffinose, and especially stachyose demonstrated reduced accumulation and efficient transport out of the leaf. We conclude that small metabolites in the cytosol of CCs do enter the translocation stream indiscriminately but are also subject to distributive forces, such as nonselective and carrier-mediated membrane transport and symplastic dispersal, that may effectively clear a compound from the phloem or retain it for long-distance transport. A model is proposed in which the transport of oligosaccharides is an adaptive strategy to improve photoassimilate retention, and consequently translocation efficiency, in the phloem.     

Maleimide-oligo(ethylene glycol) derivatives of camptothecin as albumin-binding prodrugs: synthesis and antitumor efficacy

In situ binding of thiol-reactive prodrugs to the cysteine-34 position of circulating albumin is a new approach in drug delivery. Therefore, five maleimide-bearing derivatives of the anticancer drug camptothecin (CPT) were developed as albumin-binding prodrugs. These compounds were synthesized by reacting heterobifunctional cross-linkers based on oligo(ethylene glycols) [3-6 (O-CH(2)-CH(2)) units] bearing a maleimide group on one end and a carboxylic acid group on the other with camptothecin 20-O-glycinate. Incorporating oligo(ethylene glycol) chains into the prodrugs enhanced their water-solubility when compared to the parent compound (up to 27-fold). HPLC studies showed that the prodrugs react almost quantitatively with the cysteine-34 position of endogenous albumin within a few minutes after incubation of the CPT derivatives with human blood plasma. The therapeutic potential of two of the prodrugs was assessed in nude mice bearing a colon xenograft (HT-29). Both albumin-binding derivatives of camptothecin were well-tolerated and showed enhanced antitumor efficacy when compared to CPT.     

Activin induces x-zone apoptosis that inhibits luteinizing hormone-dependent adrenocortical tumor formation in inhibin-deficient mice

Inhibin and activin are members of the transforming growth factor beta (TGF-beta) family of ligands produced and secreted primarily by the gonads and adrenals. Inhibin-null (INH(-/-)) mice develop gonadal tumors and-when gonadectomized-adrenocortical carcinoma. The mechanisms leading to adrenal tumorigenesis have been proposed to involve the lack of a gonadal factor and/or a compensatory increase in gonadotropins. In order to achieve elevation of gonadotropins without the concomitant loss of a gonadal hormone, we crossed INH(-/-) mice with a transgenic mouse strain that has chronically elevated luteinizing hormone (LH) levels (LH-CTP). Compound INH(-/-)-LH-CTP mice die within 6 weeks of age from severe cancer cachexia induced by large, activin-secreting ovarian tumors. Unexpectedly, INH(-/-)-LH-CTP mice not only fail to develop adrenal tumors but have smaller adrenals, with a regressed x zone, indicating that elevated LH levels are not sufficient to induce adrenal tumor formation. However, following gonadectomy, INH(-/-)-LH-CTP mice develop large, sex steroid-producing adrenal tumors that arise from the x zone, indicating a growth-promoting effect of high levels of LH on the adrenal cortex in the absence of ovarian tumors. In addition, in vivo and in vitro data indicate that activin induces apoptosis specifically in the adrenal x zone. The restricted expression of activin receptor subunits and Smad2 in cells of the adrenal x zone, together with the elevated activin levels in INH(-/-)-LH-CTP mice, supports the conclusion that activin inhibits adrenal tumor growth by inducing x-zone regression.     

Bicarbonate enhances the hydroxylation, nitration, and peroxidation reactions catalyzed by copper, zinc superoxide dismutase. Intermediacy of carbonate anion radical

The effect of bicarbonate anion (HCO(3)(-)) on the peroxidase activity of copper, zinc superoxide dismutase (SOD1) was investigated using three structurally different probes: 5, 5'-dimethyl-1-pyrroline N-oxide (DMPO), tyrosine, and 2, 2'-azino-bis-[3-ethylbenzothiazoline]-6-sulfonic acid (ABTS). Results indicate that HCO(3)(-) enhanced SOD/H(2)O(2)-dependent (i) hydroxylation of DMPO to DMPO-OH as measured by electron spin resonance, (ii) oxidation and nitration of tyrosine to dityrosine, nitrotyrosine, and nitrodityrosine as measured by high pressure liquid chromatography, and (iii) oxidation of ABTS to the ABTS cation radical as measured by UV-visible spectroscopy. Using oxygen-17-labeled water, it was determined that the oxygen atom present in the DMPO-OH adduct originated from H(2)O and not from H(2)O(2). This result proves that neither free hydroxyl radical nor enzyme-bound hydroxyl radical was involved in the hydroxylation of DMPO. We postulate that HCO(3)(-) enhances SOD1 peroxidase activity via formation of a putative carbonate radical anion. This new and different perspective on HCO(3)(-)-mediated oxidative reactions of SOD1 may help us understand the free radical mechanism of SOD1 and related mutants linked to amyotrophic lateral sclerosis.