Austinogebia, a new genus in the Upogebiidae and rediagnosis of its close relative, Gebiacantha Ngoc-Ho, 1989 (Crustacea: Decapoda: Thalassinidea)

New material described recently permits the separation of six upogebiid species into the new genus Austinogebia, for which the diagnostic characters and a key are presented. The new taxon is compared to its close relative, Gebiacantha Ngoc-Ho, 1989, and the opportunity is taken to rediagnose the latter.     

CD45 modulates galectin-1-induced T cell death: regulation by expression of core 2 O-glycans.

Galectin-1 induces death of immature thymocytes and activated T cells. Galectin-1 binds to T cell-surface glycoproteins CD45, CD43, and CD7, although the precise roles of each receptor in cell death are unknown. We have determined that CD45 can positively and negatively regulate galectin-1-induced T cell death, depending on the glycosylation status of the cells. CD45(+) BW5147 T cells lacking the core 2 beta-1,6-N-acetylglucosaminyltransferase (C2GnT) were resistant to galectin-1 death. The inhibitory effect of CD45 in C2GnT(-) cells appeared to require the CD45 cytoplasmic domain, because Rev1.1 cells expressing only CD45 transmembrane and extracellular domains were susceptible to galectin-1 death. Moreover, treatment with the phosphotyrosine-phosphatase inhibitor potassium bisperoxo(1,10-phenanthroline)oxovanadate(V) enhanced galectin-1 susceptibility of CD45(+) T cell lines, but had no effect on the death of CD45(-) T cells, indicating that the CD45 inhibitory effect involved the phosphatase domain. Expression of the C2GnT in CD45(+) T cell lines rendered the cells susceptible to galectin-1, while expression of the C2GnT in CD45(-) cells had no effect on galectin-1 susceptibility. When CD45(+) T cells bound to galectin-1 on murine thymic stromal cells, only C2GnT(+) T cells underwent death. On C2GnT(+) cells, CD45 and galectin-1 co-localized in patches on membrane blebs while no segregation of CD45 was seen on C2GnT(-) T cells, suggesting that oligosaccharide-mediated clustering of CD45 facilitated galectin-1-induced cell death.     

Differential Histopathology and Chemokine Gene Expression in Lung Tissues following Respiratory Syncytial Virus (RSV) Challenge of Formalin-Inactivated RSV- or BBG2Na-Immunized Mice

A BALB/c mouse model of enhanced pulmonary pathology following vaccination with formalin-inactivated alum-adsorbed respiratory syncytial virus (FI-RSV) and live RSV challenge was used to determine the type and kinetics of histopathologic lesions induced and chemokine gene expression profiles in lung tissues. These data were compared and contrasted with data generated following primary and/or secondary RSV infection or RSV challenge following vaccination with a promising subunit vaccine, BBG2Na. Severe peribronchiolitis and perivascularitis coupled with alveolitis and interstitial inflammation were the hallmarks of lesions in the lungs of FI-RSV-primed mice, with peak histopathology evident on days 5 and 9. In contrast, primary RSV infection resulted in no discernible lesions, while challenge of RSV-primed mice resulted in rare but mild peribronchiolitis and perivascularitis, with no evidence of alveolitis or interstitial inflammation. Importantly, mice vaccinated with a broad dose range (20 to 0.02 microg) of a clinical formulation of BBG2Na in aluminium phosphate demonstrated histopathology similar to that observed in secondary RSV infection. At the molecular level, FI-RSV priming was characterized by a rapid and strong up-regulation of eotaxin and monocyte chemotactic protein 3 (MCP-3) relative gene expression (potent lymphocyte and eosinophil chemoattractants) that was sustained through late time points, early but intermittent up-regulation of GRO/melanoma growth stimulatory activity gene and inducible protein 10 gene expression, while macrophage inflammatory protein 2 (MIP-2) and especially MCP-1 were up-regulated only at late time points. By comparison, primary RSV infection or BBG2Na priming resulted in considerably lower eotaxin and MCP-3 gene expression increases postchallenge, while expression of lymphocyte or monocyte chemoattractant chemokine genes (MIP-1beta, MCP-1, and MIP-2) were of higher magnitude and kinetics at early, but not late, time points. Our combined histopathologic and chemokine gene expression data provide a basis for differentiating between aberrant FI-RSV-induced immune responses and normal responses associated with RSV infection in the mouse model. Consequently, our data suggest that BBG2Na may constitute a safe RSV subunit vaccine for use in seronegative infants.     

Common Antiviral Cytotoxic T-Lymphocyte Epitope for Diverse Arenaviruses

Members of the Arenaviridae family have been isolated from mammalian hosts in disparate geographic locations, leading to their grouping as Old World types (i.e., lymphocytic choriomeningitis virus [LCMV], Lassa fever virus [LFV], Mopeia virus, and Mobala virus) and New World types (i.e., Junin, Machupo, Tacaribe, and Sabia viruses) (C. J. Peters, M. J. Buchmeier, P. E. Rollin, and T. G. Ksiazek, p. 1521-1551, in B. N. Fields, D. M. Knipe, and P. M. Howley [ed.], Fields virology, 3rd ed., 1996; P. J. Southern, p. 1505-1519, in B. N. Fields, D. M. Knipe, and P. M. Howley [ed.], Fields virology, 3rd ed., 1996). Several types in both groups-LFV, Junin, Machupo, and Sabia viruses-cause severe and often lethal human diseases. By sequence comparison, we noted that eight Old World and New World arenaviruses share several amino acids with the nucleoprotein (NP) that consists of amino acids (aa) 118 to 126 (NP 118-126) (RPQASGVYM) of LCMV that comprise the immunodominant cytotoxic T-lymphocyte (CTL) epitope for H-2(d) mice (32). This L(d)-restricted epitope constituted >97% of the total bulk CTLs produced in the specific antiviral or clonal responses of H-2(d) BALB mice. NP 118-126 of the Old World arenaviruses LFV, Mopeia virus, and LCMV and the New World arenavirus Sabia virus bound at high affinity to L(d). The primary H-2(d) CTL anti-LCMV response as well as that of a CTL clone responsive to LCMV NP 118-126 recognized target cells coated with NP 118-126 peptides derived from LCMV, LFV, and Mopeia virus but not Sabia virus, indicating that a common functional NP epitope exists among Old World arenaviruses. Use of site-specific amino acid exchanges in the NP CTL epitope among these arenaviruses identified amino acids involved in major histocompatibility complex binding and CTL recognition.     

Requirement of Cysteines and Length of the Human Respiratory Syncytial Virus M2-1 Protein for Protein Function and Virus Viability

The M2-1 protein of human respiratory syncytial virus (hRSV) promotes processive RNA synthesis and readthrough at RSV gene junctions. It contains four highly conserved cysteines, three of which are located in the Cys(3)-His(1) motif at the N terminus of M2-1. Each of the four cysteines, at positions 7, 15, 21, and 96, in the M2-1 protein of hRSV A2 strain was individually replaced by glycines. When tested in an RSV minigenome replicon system using beta-galactosidase as a reporter gene, C7G, C15G, and C21G located in the Cys(3)-His(1) motif showed a significant reduction in processive RNA synthesis compared to wild-type (wt) M2-1. C96G, which lies outside the Cys(3)-His(1) motif, was fully functional in supporting processive RNA synthesis in vitro. Each of these cysteine substitutions was introduced into an infectious antigenomic cDNA clone derived from hRSV A2 strain. Except for C96G, which resulted in a viable virus, no viruses were recovered with mutations in the Cys(3)-His(1) motif. This indicates that the Cys(3)-His(1) motif is critical for M2-1 function and for RSV replication. The functional requirement of the C terminus of the M2-1 protein was examined by engineering premature stop codons that caused truncations of 17, 46, or 67 amino acids from the C terminus. A deletion of 46 or 67 amino acids abolished the synthesis of full-length beta-galactosidase mRNA and did not result in the recovery of viable viruses. However, a deletion of 17 amino acids from the C terminus of M2-1 reduced processive RNA synthesis in vitro and was well tolerated by RSV. Relocation of the M2-1 termination codon upstream of the M2-2 initiation codons did not significantly affect the expression of the M2-2 protein. Both rA2-Tr17 and rA2-C96G did not replicate as efficiently as wt rA2 in HEp-2 cells and was restricted in replication in the respiratory tracts of cotton rats.     

Crystal structure and mechanism of catalysis of a pyrazinamidase from Pyrococcus horikoshii.

Bacterial pyrazinamidase (PZAase)/nicotinamidase converts pyrazinamide (PZA) to ammonia and pyrazinoic acid, which is active against Mycobacterium tuberculosis. Loss of PZAase activity is the major mechanism of pyrazinamide-resistance by M. tuberculosis. We have determined the crystal structure of the gene product of Pyrococcus horikoshii 999 (PH999), a PZAase, and its complex with zinc ion by X-ray crystallography. The overall fold of PH999 is similar to that of N-carbamoylsarcosine amidohydrolase (CSHase) of Arthrobacter sp. and YcaC of Escherichia coli, a protein with unknown physiological function. The active site of PH999 was identified by structural features that are also present in the active sites of CSHase and YcaC: a triad (D10, K94, and C133) and a cis-peptide (between V128 and A129). Surprisingly, a metal ion-binding site was revealed in the active site and subsequently confirmed by crystal structure of PH999 in complex with Zn(2+). The roles of the triad, cis-peptide, and metal ion in the catalysis are proposed. Because of extensive homology between PH999 and PZAase of M. tuberculosis (37% sequence identity), the structure of PH999 provides a structural basis for understanding PZA-resistance by M. tuberculosis harboring PZAase mutations.     

Quantitative trait loci influencing drought tolerance in grain sorghum (Sorghum bicolor L. Moench)

Drought is a major constraint in sorghum production worldwide. Drought-stress in sorghum has been characterized at both pre-flowering and post-flowering stages resulting in a drastic reduction in grain yield. In the case of post-flowering drought stress, lodging further aggravates the problem resulting in total loss of crop yield in mechanized agriculture. The present study was conducted to identify quantitative trait loci (QTLs) controlling post-flowering drought tolerance (stay green), pre-flowering drought tolerance and lodging tolerance in sorghum using an F₇ recombinant inbred line (RIL) population derived from the cross SC56×Tx7000. The RIL lines, along with parents, were evaluated for the above traits in multiple environments. With the help of a restriction fragment length polymorphism (RFLP) map, which spans 1,355 cM and consists of 144 loci, nine QTLs, located over seven linkage groups were detected for stay green in several environments using the method of composite interval mapping. Comparison of the QTL locations with the published results indicated that three QTLs located on linkage groups A, G and J were consistent. This is considered significant since the stay green line SC56 used in our investigation is from a different source compared to B35 that was used in all the earlier investigations. Comparative mapping has shown that two stay green QTLs identified in this study corresponded to stay green QTL regions in maize. These genomic regions were also reported to be congruent with other drought-related agronomic and physiological traits in maize and rice, suggesting that these syntenic regions might be hosting a cluster of genes with pleiotropic effects implicated in several drought tolerance mechanisms in these grass species. In addition, three and four major QTLs responsible for lodging tolerance and pre-flowering drought tolerance, respectively, were detected. This investigation clearly revealed the important and consistent stay green QTLs in a different stay green source that can logically be targeted for positional cloning. The identification of QTLs and markers for pre-flowering drought tolerance and lodging tolerance will help plant breeders in manipulating and pyramiding those traits along with stay green to improve drought tolerance in sorghum. Received: 2 June 2000 / Accepted: 15 November 2000     

Photoautotrophic growth response of in vitro cultured coffee plantlets to ventilation methods and photosynthetic photon fluxes under carbon dioxide enriched condition

Effects of two ventilation methods (forced and natural) and two photosynthetic photon fluxes (PPF, 150 and 250 μmol m⁻² s⁻¹) on the photoautotrophic growth of in vitro cultured coffee (Coffea arabusta) plantlets were investigated. Number of air exchanges was 2.7, 5.9 and 3.9 h⁻¹ for forced low rate, forced high rate and natural ventilation, respectively. Single node cuttings of in vitro cultured coffee plantlets were cultured on Florialite, a mixture of vermiculite and cellulose fibers with high air porosity, emerged in liquid half strength basal MS medium, without sucrose, vitamins and plant growth regulators. The study included 40 days in the in vitro stage and 10 days in the ex vitro stage. Mean fresh and dry weights, leaf area, shoot and root lengths and net photosynthetic rate per plantlet were significantly greater in forced high rate treatments compared with those in natural and forced low rate treatments. PPF had a distinct effect on shoot length suppression and root elongation of coffee plantlets in forced high rate treatments. The control of carbon dioxide concentration inside the culture box according to the plant demand when growing was easy with the forced ventilation method in photoautotrophic micropropagation. This revised version was published online in June 2006 with corrections to the Cover Date.     

The resident endoplasmic reticulum protein, BAP31, associates with gamma-actin and myosin B heavy chain.

BAP31 is a 28-kDa integral membrane protein of the endoplasmic reticulum whose cytosolic domain contains two caspase recognition sites that are preferentially cleaved by initiator caspases, such as caspase-8. Recently, we reported that the caspase-resistant BAP31 inhibited Fas-mediated apoptotic membrane fragmentation and the release of cytochrome c from mitochondria in KB epithelial cells (Nguyen M., Breckenridge G., Ducret A & Shore G. (2000) Mol. Cell. Biol.20, 6731-6740). We describe here the characterization by capillary liquid chromatography microelectrospray tandem MS of a BAP31 immunocomplex isolated from a HepG2 cell lysate in the absence of a death signal. We show that BAP31 specifically associates with nonmuscle myosin heavy chain B and nonmuscle gamma-actin, two components of the cytoskeleton actomyosin complex. Collectively, these data confirm that BAP31, in addition to its potential role as a chaperone, may play a fundamental role in the structural organization of the cytoplasm. Here we also show that Fas stimulation of apoptosis releases BAP31 associations with these motor proteins, a step that may contribute to extranuclear events, such as membrane remodelling, during the execution phase of apoptosis.     

Accumulation of some nitrogen compounds in response to salt stress and their relationships with salt tolerance in rice (Oryza sativa L.) seedlings

The salt-induced accumulation of some nitrogen compounds (free amino acids, ammonium and urea) in shoots of eight rice cultivars differing in salt tolerance was investigated. Salt treatment (100 mM, 6 days) significantly increased the proline content of shoots but this appeared to be a reaction to stress damage and not associated with salt tolerance, because proline contents were higher in the more sensitive cultivars. Besides proline, some other free amino acids also accumulated leading to a significant increase in the total amino acid content of the stressed seedlings. High levels of free ammonium also accumulated under conditions of stress; this was highly correlated with the accumulation of Na⁺ in the shoots and negatively correlated with salt tolerance. The accumulation of ammonium was positively correlated with the accumulation of many free amino acids, and also associated with the production of urea in the stressed seedlings. Results from the present investigations suggest that an increase in the concentration of some free amino acids including proline, may be a result of the reassimilation of the stress-induced ammonium. A high capacity to assimilate ammonium may be an important factor in alleviating the consequence of stress because ammonium can be toxic at high concentrations.