The transmembrane domain of the rice small protein OsS1Fa1 is responsible for subcellular localization and drought tolerance.

OsS1Fa1, a homologue of spinach S1Fa, is a small protein in rice that contains four distinct conserved motifs and participates in drought tolerance. However, the biological functions of these conserved motifs have not been characterized to date. Therefore, we investigated the roles of these conserved domains in the localization and cellular function of OsS1Fa1. We analysed the subcellular localization of OsS1Fa1 using confocal laser scanning microscopy (CLSM), following particle bombardment and bacterial infiltration. An E. coli in vivo reconstituted sumoylation assay was conducted to investigate sumoylation of OsS1Fa1. We characterized the function of the transmembrane domain of OsS1Fa1 in drought tolerance using transgenic Arabidopsis plants. Fluorescence analysis showed that OsS1Fa1 localized to the nuclear and cytoplasmic membranes. Mutation and cell fractionation analyses revealed that the membrane localization domain determined the subcellular localization of OsS1Fa1. The rice homologue OsS1Fa2 and Arabidopsis orthologs AtS1Fa1, AtS1Fa2, and AtS1Fa3 also exhibited similar localization patterns as OsS1Fa1. Sumoylation analysis demonstrated that OsS1Fa1 was conjugated with the small ubiquitin-related modifier (SUMO). Transgenic analysis showed that overexpression of OsS1Fa1(TMm1), a mutant form of the transmembrane domain of OsS1Fa1, in Arabidopsis did not enhance drought stress tolerance, whereas OsS1Fa1 overexpression improved the drought tolerance of transgenic Arabidopsis. Our data indicate that rice and Arabidopsis S1Fa1 proteins localize in the nuclear and cytoplasmic membranes, and that transmembrane domain determines subcellular localization and plays an important role in drought stress tolerance.

[The cases of tracing the source of patients infected with Omicron variant of SARS-CoV-2 based on wastewater-based epidemiology in Shenzhen].

Wastewater-based epidemiology (WBE) is an emerging discipline, which has been applied to drug abuse tracking and infectious disease pathogen surveillance. During the COVID-19 epidemic, WBE has been applied to monitor the epidemic trend and SARS-CoV-2 variants etc. In order to detect hidden COVID-19 cases and prevent transmission in the community, wastewater surveillance system for monitoring SARS-CoV-2 RNA was developed in Shenzhen. The sewage sampling sites were set up in key places such as the port areas, urban villages and residential communities of Futian, Nanshan, Luohu and Yantian districts. From July 26 to November 30, 2022, a total of 369 sewage sampling sites were set up, covering 1.93 million people. Continuous sampling was carried out for 3 hours in the peak period of water use every day. Sewage virus enrichment and SARS-CoV-2 nucleic acid detection were carried out by polyethylene glycol precipitation method and RT-qPCR, and a positive water sample disposal process was molded. This article aims to introduce the case of source tracing of COVID-19 infected patients based on urban sewage in Shenzhen. The sewage monitoring of Honghu water treatment plant in Luohu District played an early warning role, and the source of infection was traced. In the disposal of positive water samples in Futian South Road, Futian District, the important experience of monitoring point layout was obtained. In the sewage monitoring of Nanshan village, Nanshan District, the existence of occult infection was revealed. Sharing the experience of tracing the source of COVID-19 patients to avoid the spread of COVID-19 in the community based on wastewater surveillance of SARS-CoV-2 RNA in Shenzhen, and summarizing the advantages and application prospects of sewage surveillance can provide new ideas for monitoring emerging or re-emerging pathogens that are known to exhibit gastrointestinal excretion in the future.

[Clinical evaluation of in-plane ultrasound-guided thoracic paravertebral block using laterally intercostal approach].

OBJECTIVE: To evaluate the feasibility and success rate of in-plane ultrasound-guided paravertebral block using laterally intercostal approach. METHODS: In the study, 27 patients undergoing elective thoracic surgery were selected to do paravertebral block preoperatively. The fifth intercostal space was scanned by ultrasound probe which was placed along the long axis of the rib and 8 cm lateral to the midline of the spine. The needle was advanced in increments aiming at the space between the internal and innermost intercostal muscles. Once the space between the muscles was achieved, 20 mL of 0.5% (mass fraction) ropivacaine was injected and a catheter was inserted. Whether the tip of catheter was in right place was evaluated by ultrasound image. The block dermatomes of cold sensation were recorded 10, 20 and 30 min after the bolus drug was given. Then 0.2% ropivacaine was infused with 6 mL/h via the catheter by an analgesia pump postoperatively. The block dermatomes of cold sensation and pain score were recorded 1, 6, 24 and 48 h postoperatively. RESULTS: The first attempt success rate of catheteration was 81.48% (22/27); the tips of catheter were proved in right places after the second or third attempt in 5 patients. The median numbers of the block dermatomes 10, 20 and 30 min after the bolus drug was given were 2, 3, 4; the median numbers of block dermatomes were 5, 5, 5, 4, and of pain score were 1, 1, 2, 2 at 1, 6, 24, 48 h postoperatively; no case of bilateral block, pneumothorax or vessel puncture occurred. CONCLUSION: Thoracic paravertebral block using laterally intercostal approach is feasible, which has high success rate of block and low rate of complications.

Transport through quantum wells and superlattices on topological insulator surfaces.

We investigate electron transmission coefficients through quantum wells and quantum superlattices on topological insulator surfaces. The quantum well or superlattice is not constituted by general electronic potential barriers but by Fermi velocity barriers which originate in the different topological insulator surfaces. It is found that electron resonant modes can be renormalized by quantum wells and more clearly by quantum superlattices. The depth and width of a quantum well and superlattice, the incident angle of an electron, and the Fermi energy can be used to effectively tune the electron resonant modes. In particular, the number N of periodic structures that constitute a superlattice can further strengthen these regulating effects. These results suggest that a device could be developed to select and regulate electron propagation modes on topological insulator surfaces. Finally, we also study the conductance and the Fano factor through quantum wells and quantum superlattices. In contrast to what has been reported before, the suppression factors of 0.4 in the conductance and 0.85 in the Fano factor are observed in a quantum well, while the transport for a quantum superlattice shows strong oscillating behavior at low energy and reaches the same saturated values as in the case of a quantum well at sufficiently large energies.

Anomalous electron collimation in HgTe quantum wells with inverted band structure.

We investigate the electron collimation behavior in HgTe quantum wells (QWs) with a magnetic-electric barrier induced by a ferromagnetic metal stripe. We find that electrons can transmit perfectly through the magnetic-electric barrier at some specific incidence angles. These angles can be controlled by the tuning gate voltage, local magnetic field and Fermi energy of incident electrons in QWs with appropriate barrier length. This collimation feature can be used to construct momentum filters in HgTe QWs and has potential application in nanodevices.

Jasmonic acid carboxyl methyltransferase: a key enzyme for jasmonate-regulated plant responses.

Methyl jasmonate is a plant volatile that acts as an important cellular regulator mediating diverse developmental processes and defense responses. We have cloned the novel gene JMT encoding an S-adenosyl-l-methionine:jasmonic acid carboxyl methyltransferase (JMT) from Arabidopsis thaliana. Recombinant JMT protein expressed in Escherichia coli catalyzed the formation of methyl jasmonate from jasmonic acid with K(m) value of 38.5 microM. JMT RNA was not detected in young seedlings but was detected in rosettes, cauline leaves, and developing flowers. In addition, expression of the gene was induced both locally and systemically by wounding or methyl jasmonate treatment. This result suggests that JMT can perceive and respond to local and systemic signals generated by external stimuli, and that the signals may include methyl jasmonate itself. Transgenic Arabidopsis overexpressing JMT had a 3-fold elevated level of endogenous methyl jasmonate without altering jasmonic acid content. The transgenic plants exhibited constitutive expression of jasmonate-responsive genes, including VSP and PDF1.2. Furthermore, the transgenic plants showed enhanced level of resistance against the virulent fungus Botrytis cinerea. Thus, our data suggest that the jasmonic acid carboxyl methyltransferase is a key enzyme for jasmonate-regulated plant responses. Activation of JMT expression leads to production of methyl jasmonate that could act as an intracellular regulator, a diffusible intercellular signal transducer, and an airborne signal mediating intra- and interplant communications.

Characterization of a bifunctional enzyme fusion of trehalose-6-phosphate synthetase and trehalose-6-phosphate phosphatase of Escherichia coli.

To test the effect of the physical proximity of two enzymes catalyzing sequential reactions, a bifunctional fusion enzyme, TPSP, was constructed by fusing the Escherichia coli genes for trehalose-6-phosphate (T6P) synthetase (TPS) and trehalose-6-phosphate phosphatase (TPP). TPSP catalyzes the sequential reaction in which T6P is formed and then dephosphorylated, leading to the synthesis of trehalose. The fused chimeric gene was overexpressed in E. coli and purified to near homogeneity; its molecular weight was 88,300, as expected. The K(m) values of the TPSP fusion enzyme for the sequential overall reaction from UDP-glucose and glucose 6-phosphate to trehalose were smaller than those of an equimolar mixture of TPS and TPP (TPS/TPP). However, the k(cat) values of TPSP were similar to those of TPS/TPP, resulting in a 3.5- to 4.0-fold increase in the catalytic efficiency (k(cat)/K(m)). The K(m) and k(cat) values of TPSP and TPP for the phosphatase reaction from T6P to trehalose were quite similar. This suggests that the increased catalytic efficiency results from the proximity of TPS and TPP in the TPSP fusion enzyme. The thermal stability of the TPSP fusion enzyme was quite similar to that of the TPS/TPP mixture, suggesting that the structure of each enzyme moiety in TPSP is unperturbed by intramolecular constraint. These results clearly demonstrate that the bifunctional fusion enzyme TPSP catalyzing sequential reactions has kinetic advantages over a mixture of both enzymes (TPS and TPP). These results are also supported by the in vivo accumulation of up to 0.48 mg of trehalose per g of cells after isopropyl-beta-D-thiogalactopyranoside treatment of cells harboring the construct encoding TPSP.

NTR1 encodes a floral nectary-specific gene in Brassica campestris L. ssp. pekinensis.

We have characterized a gene specifically expressed in the floral nectaries of Brassica campestris L. ssp. pekinensis. Differential screening led to the isolation of a floral nectary-specific cDNA clone. Northern hybridization indicated that its mRNA transcript is 1450 nucleotides long and specific to the flower base. In situ hybridization and immunolocalization showed that its mRNA and protein are localized specifically to both the lateral and median nectaries of flowers. The cDNA codes for a 43.8 kDa polypeptide 392 amino acids long. The protein was named nectarin1 (NTR1) after floral nectary protein. NTR1 was located in the cytoplasm of nectariferous cells in the nectaries and was also observed in nuclei at a much lower level. The level of the transcript increases with flower development, especially during nectary development, but decreases abruptly with the opening of the flower. Genomic Southern blot analysis indicated that at least three copies of homologous genes were present in the genome of B. campestris, but that only a single copy was present in both Arabidopsis thaliana and Lycopersicon esculentum. The deduced amino acid sequence of NTR1 shows similarity to S-adenosyl-L-methionine:salicylic acid carboxyl methyltransferase of Clarkia breweri which is expressed mostly in petals. The function of the gene is speculated to be involved in the methylation of a plant secondary metabolite in the floral nectaries.

Molecular characterization of the garlic virus X genome.

The complete nucleotide sequence of the cDNA genome for garlic virus X (GVX), one of the major viruses infecting garlic plants, was determined. GVX is a single-stranded positive-sense RNA virus consisting of 8106 nucleotides excluding the 3'-end poly(A) tail and contains six open reading frames (ORFs) which encode putative proteins of 174 kDa (ORF1), 26 kDa (ORF2), 12 kDa (ORF3), 32 kDa (ORF4), 26 kDa (ORF5) and 15 kDa (ORF6). The putative viral proteins show similarity to those of carlaviruses and potexviruses but show the highest homology to shallot virus X (ShVX). Even though the GVX genome contains most of the structural elements common to carlaviruses and potexviruses, it is distinguished from them by the presence of an ORF4 which encodes an unusual protein. These results suggest that GVX may belong to an unassigned group of ShVX and GarV-type viruses rather than to the carlaviruses or potexviruses.

Identification of one of the major viruses infecting garlic plants, garlic virus X.

A partial cDNA clone for garlic virus X (GVX) was isolated. GVX was identified immunologically with an antibody raised against the recombinant coat protein (CP) and demonstrated to be one of the major viruses infecting garlic plants showing mosaic or streak symptoms. GVX belongs to an unassigned group of ShVX and GarV-type viruses rather than to carlaviruses or potexviruses. The recombinant CP of GVX was purified by Ni(2+)-NTA affinity chromatography. Anti-GVX CP antibody was raised against the purified recombinant CP. GVX particle is flexuous, rod-shaped, and about 750 nm long as determined by immunoelectron microscopy. The extent of infection by GVX of garlic plants was analyzed by Northern or immunoblot analyses of individual garlic plants cultivated in different regions. These results showed that almost all of the garlic plants tested from 40 different regions including America, China, Japan, and Korea are infected with GVX.

The loop B domain is physically separable from the loop A domain in the hairpin ribozyme.

In order to understand the catalysis mechanism of the hairpin ribozyme, mutant ribozymes were constructed. The distance between the loop A domain and the loop B domain was extended by inserting various lengths of nucleotide linkers at the hinge region in cis mutants, or the domains were separated physically in a trans mutant. All the mutant ribozymes, including the trans mutant, could cleave substrate RNA at the predicted site. A cis mutant with a single nucleotide insertion exhibited cleavage activity about twice as high as that of the wild-type (wt) ribozyme. The insertion of 2-5 nucleotides (nt) gradually reduced the activity to the level of the wt ribozyme. Insertion of a longer linker, up to 11 nt, resulted in the reduction of activity to one half of that of the wt ribozyme. The ribozyme with a single nucleotide insertion at the hinge region seems to form a more suitable conformation for catalysis by three-dimensional fold-back of the loop B to loop A containing the cleavage site. The trans mutant, in which the A and B domains were physically separated, maintained a significant level of activity, suggesting that both domains are necessary for catalysis, but separable. These results demonstrate that interaction between the A and B domains results in catalysis.

Ejaculation increases the serum prostate-specific antigen concentration.

OBJECTIVES: To determine the effect of ejaculation on the serum prostate-specific antigen (PSA) concentration in men at risk for developing prostate cancer. METHODS: A prospective, community-based study was conducted in which 64 men, aged 49 to 79 years, underwent a serum PSA determination immediately before ejaculation (baseline) and at 1 hour, 6 hours, and 24 hours following ejaculation. The serum PSA also was measured 48 hours and 1 week after ejaculation if the concentration had not returned to the baseline value by the previous time interval. All subjects abstained from ejaculation for a minimum of 7 days prior to the study and until the PSA concentration returned to the baseline level. Absolute and relative change in serum PSA concentration, as well as the time to return to baseline PSA concentration following ejaculation, were assessed. RESULTS: The serum PSA concentration increased following ejaculation in 87% of the subjects. The mean baseline PSA was 1.8 ng/mL (median, 0.7 ng/mL). The mean absolute PSA change +/- standard deviation 1 hour, 6 hours, 24 hours, and 48 hours after ejaculation was 0.8 +/- 1.32 ng/mL, 0.3 +/- 0.66 ng/mL, 0.2 +/- 0.33 ng/mL, and 0.4 +/- 0.40 ng/mL, respectively. The mean relative PSA change +/- standard error 1 hour, 6 hours, 24 hours, and 48 hours after ejaculation was 41 +/- 4%, 9 +/- 1.5%, 8 +/- 1.3%, and 10 +/- 2.3%, respectively. The absolute and relative changes in PSA concentration noted 1 hour, 6 hours, and 24 hours after ejaculation were statistically significant (P = 0.0001). A strong correlation was observed between absolute change in PSA and baseline serum PSA, at each time interval (1 hour: r = 0.68, 6 hours: r = 0.77, 24 hours: r = 0.70; P < 0.0001) after ejaculation. Similarly, a significant correlation was noted between absolute change in PSA and patient age at each time interval (1 hour: r = 0.37, 6 hours: r = 0.38; P = 0.002, 24 hours: r = 0.55; P < 0.0001). Ninety-two percent of subjects returned to baseline by 24 hours (95% confidence interval (Cl) = 83% to 97%), whereas 97% of subjects returned to baseline by 48 hours (95% Cl = 89% to 99%). CONCLUSIONS: Ejaculation causes a significant increase in the serum PSA concentration in men between 49 and 79 years of age that may persist for up to 48 hours. This change appears to correlate with age and baseline PSA. It is recommended that men abstain from ejaculation for 48 hours prior to having a serum PSA determination.

Stress leak point pressure: a simple and reproducible method utilizing a fiberoptic microtransducer.

OBJECTIVES: This work demonstrates a simple technique utilizing a fiberoptic microtransducer that provides statistically reproducible stress leak point pressure (SLPP) results without the use of fluorourodynamics. METHODS: Nineteen stress incontinent patients with varied clinical histories underwent two SLPP measurements on 2 separate days, totaling four data points. A 14 F catheter sheath was inserted to empty the bladder. Through this sheath, a 5 F fiberoptic microtransducer was inserted into the bladder and zeroed. Then, 250 cc of indigo-carmine solution was instilled, during which a filling cystometrogram was performed. The sheath was removed, leaving only the 5 F transducer in the bladder. A 2 by 2 inch gauze was placed at the meatus. As each participant performed a slow Valsalva maneuver, an event marker was used to note the pressure at which indigo solution was first seen to stain the gauze. RESULTS: SLPPs ranged from 15 to 140 cm water (H2O). A two-tailed paired t test demonstrated no statistical difference (P < 0.6) between the two SLPPs performed on day 1, with a mean difference of 1.05 +/- 2.61 (95% confidence interval [CI]). Comparison of the two SLPPs performed on day 2 also revealed no statistical difference (P < 0.8), with a mean difference of -0.17 +/- 5.65 (95% CI). Lastly, comparison of the mean SLPPs from day 1 with the mean SLPPs from day 2 revealed no statistically significant difference (P < 0.8), with the mean difference of -0.59 +/- 1.62 (95% CI). CONCLUSIONS: This study demonstrates a simple technique that produces reproducible SLPP measurements in a wide variety of clinical settings and avoids ionizing radiation.

Incidence of vesicoureteral reflux in children with unilateral renal agenesis.

We retrospectively reviewed 51 pediatric cases of unilateral renal agenesis to determine the incidence of contralateral vesicoureteral reflux. Initial diagnosis of unilateral renal agenesis was made by evaluation of associated congenital abnormality in 21 patients, evaluation of prenatally detected abnormality in 11, evaluation of urinary tract infection in 7, sibling screening in 3, hypertension in 2 and other methods in 7. A voiding cystourethrogram was obtained in 44 cases. Indications for the study included urinary tract infection in 11 patients, hydronephrosis in 18 and screening in 15. Overall, vesicoureteral reflux occurred in 19 of the 51 patients (37%). The highest incidence of contralateral reflux was in those with a prenatal abnormality with or without hydronephrosis (77%) although 5 of 15 patients (33%) who underwent a screening voiding cystourethrogram had reflux. Mean followup was 50 months. Of the patients with vesicoureteral reflux reimplantation was performed in 9, reflux spontaneously resolved in 3 and reflux persisted in 7. There is a high incidence of vesicoureteral reflux in children with unilateral renal agenesis and a voiding cystourethrogram is recommended even in the absence of hydronephrosis or urinary tract infection. Although 50% of children in our series underwent surgical intervention, a period of nonoperative observation is warranted.