RCY1, an Arabidopsis thaliana RPP8/HRT family resistance gene,conferring resistance to cucumber mosaic virus requires salicylic acid,ethylene and a novel signal transduction mechanism

The dominant locus, RCY1, in the Arabidopsis thaliana ecotype C24 confers resistance to the yellow strain of cucumber mosaic virus (CMV-Y). The RCY1 locus was mapped to a 150-kb region on chromosome 5. Sequence comparison of this region from C24 and a CMV-Y-susceptible C24 mutant predicts that the RCY1 gene encodes a 104-kDa CC-NBS-LRR-type protein. The RCY1 gene from C24, when expressed in the susceptible ecotype Wassilewskija (Ws), restricted the systemic spread of virus. RCY1 is allelic to the resistance genes RPP8 from the ecotype Landsberg erecta and HRT from the ecotype Dijon-17, which confer resistance to Peronospora parasitica biotype Emco5 and turnip crinkle virus (TCV), respectively. Examination of RCY1 plants defective in salicylic acid (SA), jasmonic acid (JA) and ethylene signaling revealed a requirement for SA and ethylene signaling in mounting a resistance response to CMV-Y. The RCY1 nahG etr1 double mutants exhibited an intermediate level of susceptibility to CMV-Y, compared to the resistant ecotype C24 and the susceptible ecotypes Columbia and Nossen. This suggests that in addition to SA and ethylene, a novel signaling mechanism is associated with the induction of resistance in CMV-Y-infected C24 plants. Moreover, our results suggest that the signaling pathways downstream of the RPP8, HRT, and RCY1 have evolved independently.     

Molecular cloning and expression of a Schistosoma japonicum tegumental membrane-associated antigen from Japanese strain

Diagnosis and vaccine development form the major focus in creating strategies for the control of schistosomiasis. In this study, we established an IgG1 mouse monoclonal antibody (MoAb), SJA111, which strongly reacted with 23–25-kDa Schistosoma japonicum tegumental-associated membrane proteins, but not with eight other parasitic antigens. A λgt 11 cDNA library from the Japanese strain of the Schistosoma japonicum adult worm was screened with SJA111 as a probe. A single positive clone was isolated and the nucleotide sequence of the isolated cDNA was determined. The cDNA clone consisted of 844 bp, and the coding region contained 576 bp which was translated to a 22.6-kDa protein. This region showed 99.0% and 99.3% significant homology with those of the Chinese and Philippine strains of Schistosoma japonicum, respectively. The deduced amino acid sequence of the protein was identical to that of the Philippine strain and only one residue differed from that of the Chinese strain. The recombinant form of the tegumental protein was expressed in Escherichia coli and purified by a combination of ion exchange and affinity chromatography, and the purified protein was found to react with the sera of patients infected with Schistosoma japonicum. This result suggests that this antigen may be useful in the immunodiagnosis of schistosomiasis as well as in the development of an effective vaccine.     

Involvement of autophagy in trypsinogen activation within the pancreatic acinar cells

Autophagy is mostly a nonselective bulk degradation system within cells. Recent reports indicate that autophagy can act both as a protector and killer of the cell depending on the stage of the disease or the surrounding cellular environment (for review see Cuervo, A.M. 2004. Trends Cell Biol. 14:70-77). We found that cytoplasmic vacuoles induced in pancreatic acinar cells by experimental pancreatitis were autophagic in origin, as demonstrated by microtubule-associated protein 1 light chain 3 expression and electron microscopy experiments. To analyze the role of macroautophagy in acute pancreatitis, we produced conditional knockout mice lacking the autophagy-related 5 gene in acinar cells. Acute pancreatitis was not observed, except for very mild edema in a restricted area, in conditional knockout mice. Unexpectedly, trypsinogen activation was greatly reduced in the absence of autophagy. These results suggest that autophagy exerts devastating effects in pancreatic acinar cells by activation of trypsinogen to trypsin in the early stage of acute pancreatitis through delivering trypsinogen to the lysosome.     

Synthesis and Enantioseparation Ability of Xylan Bisphenylcarbamate Derivatives as Chiral Stationary Phases in HPLC

Ten novel xylan bisphenylcarbamate derivatives bearing meta‐ and para‐substituents on their phenyl groups were synthesized and their chiral recognition abilities were evaluated as the chiral stationary phases (CSPs) for high‐performance liquid chromatography (HPLC) after coating them on macroporous silica. The chiral recognition abilities of these CSPs depended on the nature, position, and number of the substituents on the phenyl moieties. The introduction of an electron‐donating group was more attractive than an electron‐withdrawing group to improve the chiral recognition ability of the xylan phenylcarbamate derivatives. Among the CSPs discussed in this study, xylan bis(3,5‐dimethylphenylcarbamate)‐based CSP seems to possess the highest resolving power for many racemates, and the meta‐substituted CSPs showed relatively better chiral recognition than the para‐substituted ones. For some racemates, the xylan bis(3,5‐dimethylphenylcarbamate) derivative exhibited higher enantioselectivity than the CSP based on cellulose tris(3,5‐dimethylphenylcarbamate). Chirality 27:518–522, 2015 © 2015 Wiley Periodicals, Inc.     

Spontaneous transformation and immortalization of human endothelial cells

Summary A new cell line from the human umbilical vein has been established and maintained for more than 5 yr (180 generations; 900 population doublings). This strain, designated ECV304, is characterized by a cobblestone monolayer growth pattern, high proliferative potential without any specific growth factor requirement, and anchorage dependency with contact inhibition. Karyotype analysis of this cell line reveals it to be of human chromosomal constitution with a high trisomic karyotype (mode 80). Ultrastructurally, endothelium-specific Weibel-Palade bodies were identified. Although one of the endothelial cell markers, Factor VIII-related antigen (VIIIR:Ag) was negative in this cell line, immunocytochemical staining for the lectin Ulex europaeus I (UEA-I), and PHM5 (anti-human endothelium as well as glomerular epithelium monoclonal antibody) was positive, and angiotensin-converting enzyme (ACE) activity was also demonstrated. In addition, ECV304 displayed negativity for alkaline and acid phosphatase and for the epithelial marker keratin. All of these findings suggest that ECV304 cells originated from umbilical vein endothelial cells by spontaneous transformation. Ultrastructurally, no viruslike particles have been detected intracellularly. Nude mouse tumorigenicity and rabbit cornea tests were both positive. This is a report on a novel case of phenotypic alteration of normal venous endothelial cells of human origin in vitro, and generation of a transformant with indefinite life spans. This line may be useful in studies of some physiologically active factors available for medical use.     

Stimulation of platelet-derived growth factor-induced DNA synthesis by angiotensin II in rabbit vascular smooth muscle cells

In cultured rabbit vascular smooth muscle cells (VSMCs), angiotensin II by itself had little mitogenic effect even in the presence of cell-free plasma-derived serum (PDS), but markedly stimulated the platelet-derived growth factor (PDGF)-induced DNA synthesis in the presence of PDS. The maximal extent of DNA synthesis induced by PDGF plus angiotensin II was about twice that induced by PDGF alone. The stimulatory effect of angiotensin II was dose-dependent with the maximal response seen at 1 microM and was inhibited by the specific angiotensin II receptor antagonist, [Sar1, Ile8]angiotensin II. In VSMCs, both PDGF and angiotensin II induced expression of the c-fos gene in dose-dependent manners. In contrast to the synergistic effect of angiotensin II and PDGF on DNA synthesis, they induced expression of the c-fos gene in an additive manner. These results suggest that angiotensin II may act as a growth regulator for VSMCs in addition to acting as a vasoconstrictor.     

Functional analysis of guinea pig β1-adrenoceptor

Although similarity of pharmacological responses to certain stimuli between guinea pigs and humans has been reported, this has been poorly defined by a molecular biological approach. In this study, we cloned the gene of guinea pig ?1-adrenoceptor (ADRB1). The deduced amino acid sequence of guinea pig ADRB1 (467-aa) showed 91% and 92% identity with the human and rat ADRB1 sequences, respectively. Using HEK293T cells expressing guinea pig, human and rat ADRB1s independently, we elucidated the functional characteristics of each ADRB1. The ligand-binding profiles and the concentration-response relationships for isoprenaline-induced cyclic adenosine monophosphate (cAMP) production were similar among the three ADRB1s. Isoprenaline also induced phosphorylation of extracellular-signal related kinases (ERK) through ADRB1s in a concentration-dependent manner. The minimum effective concentration of isoprenaline for phosphorylation of ERK, through guinea pig ADRB1 was the same as through human ADRB1, but markedly lower than that of through rat ADRB1. ERK phosphorylation through guinea pig ADRB1 was sensitive to pertussis toxin, a dominant-negative ras and PD98059, indicating that a G(i)-mediated pathway is involved in the ADRB1/ERK signaling loop. These results suggest that the G(i)-coupling efficacy of guinea pig and human ADRB1s may be higher than that of rat ADRB1.     

Maturation and spawning processes of anadromous and resident pond smelt in lake ogawara

Pond smelt,Hypomesus nipponensis McAllister, in Lake Ogawara demonstrate alternative life history strategies, as evidenced by the coexistence of anadromous and resident fish. However, it is unknown if anadromous and resident groups interbreed. In this study, maturation and spawning processes were examined and compared between anadromous and resident groups. Histological observations indicated negligible variation in the maturational stage composition of oocytes, the frequency of oocyte diameter being unimodal for all specimens at different maturational stages. Oocytes were absent in the ovaries of spent fish. Accordingly, the species can be considered a semelparous spawner with unimodal oocyte diameter distribution. Temporal changes in the proportion of spent fish were compared between anadromous and resident groups. Spawning of both groups began in late March and peaked over April 8–12. Although both groups did not differ significantly in the period of peak spawning, anadromous fish finished spawning earlier than resident ones. Anadromous fish were not able to spawn upon migration into Lake Ogawara, and quickly matured after immigration, contrasting with resident fish.