In vitro regulation of morphogenesis in peanut (Arachis hypogaea L.)

Callusing, caulogenesis, in vitro flowering and somatic embryogenesis were induced from the base of leaflets derived from mature embryos of peanut (Arachis hypogaea) by altering the hormonal composition of the Murashige and Skoog's (MS) basal medium. A combination of 4 mg/l alpha napththaleneacetic acid (NAA) and 5 mg/l 6-benylaminopurine (BAP) was optimum for inducing caulogenic buds. The caulogenic buds proliferated in medium with 3 mg/l BAP. Differentiation of these buds to shoots was achieved in MS basal medium with 0.5 mg/l each of BAP and kinetin (KN). Shoot buds and flower buds were produced when caulogenic buds were cultured on medium containing 1 mg/l BAP and 1 mg/l KN, prior to elongation. Clonally propagated plantlets derived from axillary buds elongated, formed roots and were grown to maturity in soil. Embryogenic mass formation was induced from the leaf base in the presence of 20 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D). Somatic embryos developed upon reducing 2,4-D to 3 mg/l.     

High-frequency conversion of abnormal peanut somatic embryos

Peanuts (Arachis hypogaea L.) are widely cultivated as a rich source of protein and oil. Although protocols for the regeneration of peanut via somatic embryogenesis and organogenesis have been developed, most of them have resulted in low frequencies of plant recovery. In this report, we describe a protocol for plantlet formation at high frequency from somatic embryos. Morphologically abnormal somatic embryos germinated and produced roots only in medium devoid of growth regulators. Shoots emerged from the undeveloped plumule of these rooted embryos in medium containing both 6-benzyladenine (BA) and kinetin (KN), or in medium with thidiazuron (TDZ) alone. In Murashige and Skoog basal medium supplemented with 8.9 μm BA and 14 μm KN, 86% of the embryos developed shoots. Substitution of BA and KN with 22.7 μm TDZ increased plant recovery from 86% to 92%. Plants grown on TDZ had multiple shoots. Eighty-four percent of these plants survived in sandy soil and were grown to maturity. Received: 12 February 1996 / Revision received: 11 July 1996 / Accepted 30 April 1997     

Mangrove associates versus true mangroves: a comparative analysis of leaf litter decomposition in Sundarban

Mangrove species are broadly classified as ‘true mangroves’ and ‘mangrove associates’. We hypothesized that the leaf litter decomposition rates of true mangroves differ significantly from the mangrove associates under the same ecological and bio-climatic conditions. In order to test this hypothesis, the leaf litter decay rates of 24 true mangrove species and 10 mangrove associates along with the concomitant carbon and nitrogen dynamics of the litters were studied in the tropical mangrove forest of Sundarban by means of litter bags. The decomposition was monitored for six consecutive weeks in the pre-monsoon, monsoon and post-monsoon season. All the species in general went through a rapid decay phase in the first 2 weeks, however, the rate substantially decreased in the following 4 weeks. Most of the species studied had significant seasonal variability (p < 0.05) in the decay rate. Species-specific decay was highest throughout the monsoon and least during the post-monsoon season. The mean dry weight composition (i.e. percentage of dry weight of the leaf litters remaining at the end of weekly intervals) of the true mangroves was 10–12 % higher than the mangrove associates throughout the sampling period. The mean decay constants (K in week^?1) of the true mangroves were 0.15 ± 0.05, 0.20 ± 0.06 and 0.16 ± 0.05 in the pre-monsoon, monsoon and post-monsoon season respectively. The mangrove associates had significantly higher decay constants in the respective seasons that followed the order 0.23 ± 0.09, 0.25 ± 0.06 and 0.24 ± 0.09. As a consequence, the computed mean half-life period of the true mangrove litters (32 ± 11 days) was much higher than the mangrove associates (23 ± 11 days). This showed that collectively the leaf litters of mangrove associates degraded at a much faster rate than the true mangroves throughout the annual cycle and thus our hypothesis was justified.     

Diospyrin derivative, an anticancer quinonoid, regulates apoptosis at endoplasmic reticulum as well as mitochondria by modulating cytosolic calcium in human breast carcinoma cells

Diospyrin diethylether (D7), a bisnaphthoquinonoid derivative, exhibited an oxidative stress-dependent apoptosis in several human cancer cells and tumor models. The present study was aimed at evaluation of the increase in cytosolic calcium [Ca(2+)](c) leading to the apoptotic cell death triggered by D7 in MCF7 human breast carcinoma cells. A phosphotidylcholine-specific phospholipase C (PC-PLC) inhibitor, viz. U73122, and an antioxidant, viz. N-acetylcysteine, could significantly prevent the D7-induced rise in [Ca(2+)](c) and PC-PLC activity. Using an endoplasmic reticulum (ER)-Ca(2+) mobilizer (thapsigargin) and an ER-IP3R antagonist (heparin), results revealed ER as a major source of [Ca(2+)](c) which led to the activation of calpain and caspase12, and cleavage of fodrin. These effects including apoptosis were significantly inhibited by the pretreatment of Bapta-AM (a cell permeable Ca(2+)-specific chelator), or calpeptin (a calpain inhibitor). Furthermore, D7-induced [Ca(2+)](c) was found to alter mitochondrial membrane potential and induce cytochrome c release, which was inhibited by either Bapta-AM or ruthenium red (an inhibitor of mitochondrial Ca(2+) uniporter). Thus, these results provided a deeper insight into the D7-induced redox signaling which eventually integrated the calcium-dependent calpain/caspase12 activation and mitochondrial alterations to accentuate the induction of apoptotic cell death.     

RPA physically interacts with the human DNA glycosylase NEIL1 to regulate excision of oxidative DNA base damage in primer-template structures

The human DNA glycosylase NEIL1, activated during the S-phase, has been shown to excise oxidized base lesions in single-strand DNA substrates. Furthermore, our previous work demonstrating functional interaction of NEIL1 with PCNA and flap endonuclease 1 (FEN1) suggested its involvement in replication-associated repair. Here we show interaction of NEIL1 with replication protein A (RPA), the heterotrimeric single-strand DNA binding protein that is essential for replication and other DNA transactions. The NEIL1 immunocomplex isolated from human cells contains RPA, and its abundance in the complex increases after exposure to oxidative stress. NEIL1 directly interacts with the large subunit of RPA (K_d ～20 nM) via the common interacting interface (residues 312–349) in NEIL1's disordered C-terminal region. RPA inhibits the base excision activity of both wild-type NEIL1 (389 residues) and its C-terminal deletion CΔ78 mutant (lacking the interaction domain) for repairing 5-hydroxyuracil (5-OHU) in a primer-template structure mimicking the DNA replication fork. This inhibition is reduced when the damage is located near the primer-template junction. Contrarily, RPA moderately stimulates wild-type NEIL1 but not the CΔ78 mutant when 5-OHU is located within the duplex region. While NEIL1 is inhibited by both RPA and Escherichia coli single-strand DNA binding protein, only inhibition by RPA is relieved by PCNA. These results showing modulation of NEIL1's activity on single-stranded DNA substrate by RPA and PCNA support NEIL1's involvement in repairing the replicating genome.     

Role of rusticyanin in the electron transport process in Thiobacillus ferrooxidans.

Effect of diethyl dithiocarbamate (DEDC), an antimicrobial agent, on growth of Thiobacillus ferrooxidans, possibly by inhibiting rusticyanin present in the periplasmic space of the microorganism, has been studied to gain more insight into the electron transport chain in the bioleaching process. DEDC is found to form a stable complex with rusticyanin in solution and also in polyacrylamide gel. The spectrum of the complex is identical to that of Cu-DEDC complex, suggesting binding of DEDC with copper moiety of rusticyanin and resulting in inhibition of growth. In vitro reduction of purified rusticyanin by Fe(II) in absence of acid-stable cytochrome c is very slow, indicating the importance of cytochrome c in electron transport. Thus, in the iron oxidation process, acid-stable cytochrome c is the primary acceptor of electron, transferring the electron to rusticyanin at pH 2.0, which, in turn, affects electron transfer to iron-cytochrome c reductase around pH 5.5.     

Bidens pilosa L.: Exclusive report of vivipary in a non-mangrove taxa from the eastern Himalayas

Vivipary is an extremely evolved type of epigeal germination in seed plants and very common in tropical estuarine plant communities (mangroves), ensuring a greater number of viable settings of offspring. In addition to this, there are some other non-mangrove angiosperm taxa that have a successful existence, and because of their invasive nature, vivipary has been reported as an attribute of their uncongenial abiotic stress. The present observation confirms the incidence of true vivipary in Bidens pilosa L. (a member of the family Asteraceae) from the eastern Himalayan region.     

The Role of the Mammalian DNA End-processing Enzyme Polynucleotide Kinase 3’-Phosphatase in Spinocerebellar Ataxia Type 3 Pathogenesis

DNA strand-breaks (SBs) with non-ligatable ends are generated by ionizing radiation, oxidative stress, various chemotherapeutic agents, and also as base excision repair (BER) intermediates. Several neurological diseases have already been identified as being due to a deficiency in DNA end-processing activities. Two common dirty ends, 3’-P and 5’-OH, are processed by mammalian polynucleotide kinase 3’-phosphatase (PNKP), a bifunctional enzyme with 3’-phosphatase and 5’-kinase activities. We have made the unexpected observation that PNKP stably associates with Ataxin-3 (ATXN3), a polyglutamine repeat-containing protein mutated in spinocerebellar ataxia type 3 (SCA3), also known as Machado-Joseph Disease (MJD). This disease is one of the most common dominantly inherited ataxias worldwide; the defect in SCA3 is due to CAG repeat expansion (from the normal 14–41 to 55–82 repeats) in the ATXN3 coding region. However, how the expanded form gains its toxic function is still not clearly understood. Here we report that purified wild-type (WT) ATXN3 stimulates, and by contrast the mutant form specifically inhibits, PNKP’s 3’ phosphatase activity in vitro. ATXN3-deficient cells also show decreased PNKP activity. Furthermore, transgenic mice conditionally expressing the pathological form of human ATXN3 also showed decreased 3’-phosphatase activity of PNKP, mostly in the deep cerebellar nuclei, one of the most affected regions in MJD patients’ brain. Finally, long amplicon quantitative PCR analysis of human MJD patients’ brain samples showed a significant accumulation of DNA strand breaks. Our results thus indicate that the accumulation of DNA strand breaks due to functional deficiency of PNKP is etiologically linked to the pathogenesis of SCA3/MJD.