Optimizing nitrogen application position to change root distribution in soil and regulate maize growth and yield formation in a wide-narrow row cropping system: pot and field experiments.

The wide-and narrow-row cropping technology used for maize has the advantages of protecting cultivated soil and improving the population structure in maize fields. However, the relationship between nitrogen application position and root interactions has not been determined. Through pot and field experiments, we evaluated the effects of two nitrogen application positions ((narrow row nitrogen application (RC) and wide row nitrogen application (RN)) and two nitrogen application regimens ((high nitrogen(HN) and low nitrogen(LN)) on root growth and yield composition of wide-narrow row maize during the flowering and harvest stages. In field experiments, RC increased the biomass, length and surface area of competing roots (narrow-row roots, CR) at the flowering stage. The yield and agronomic efficiency of N(AEN) and partial factor productivity of N(PFPN) were increased by RN compared to RC under HN, However, the AEN under LN was significantly lower; There was no significant effect on maize growth and biomass allocation at the same level of application of N. At the flowering stage, the results of CR and non-competing roots (wide-row roots, NCR) was consistent under pot experiments and the field experiments, and the yield under RN was also higher than that under RC, although the difference was not significant. Furthermore, according to the principal component analysis and correlation analysis, the competing roots were the main factor influencing yield and AEN. In conclusion, our study showed that RN is a useful fertilization method to improve overall productivity. All in all, how roots coordinate neighbors and nitrogen spatial heterogeneity is a complex ecological process, and its trophic behavior deserves further study.

Transcriptome analysis revealed the expression levels of genes related to abscisic acid and auxin biosynthesis in grapevine (Vitis vinifera L.) under root restriction.

The root system is essential for the stable growth of plants. Roots help anchor plants in the soil and play a crucial role in water uptake, mineral nutrient absorption and endogenous phytohormone formation. Root-restriction (RR) cultivation, a powerful technique, confines plant roots to a specific soil space. In the present study, roots of one-year-old "Muscat Hamburg" grapevine under RR and control (nR) treatments harvested at 70 and 125 days after planting were used for transcriptome sequencing, and in total, 2031 (nR7 vs. nR12), 1445 (RR7 vs. RR12), 1532 (nR7 vs. RR7), and 2799 (nR12 vs. RR12) differentially expressed genes (DEGs) were identified. Gene Ontology (GO) enrichment analysis demonstrated that there were several genes involved in the response to different phytohormones, including abscisic acid (ABA), auxin (IAA), ethylene (ETH), gibberellins (GAs), and cytokinins (CTKs). Among them, multiple genes, such as PIN2 and ERF113, are involved in regulating vital plant movements by various phytohormone pathways. Moreover, following RR cultivation, DEGs were enriched in the biological processes of plant-type secondary cell wall biosynthesis, the defense response, programmed cell death involved in cell development, and the oxalate metabolic process. Furthermore, through a combined analysis of the transcriptome and previously published microRNA (miRNA) sequencing results, we found that multiple differentially expressed miRNAs (DEMs) and DEG combinations in different comparison groups exhibited opposite trends, indicating that the expression levels of miRNAs and their target genes were negatively correlated. Furthermore, RR treatment indeed significantly increased the ABA content at 125 days after planting and significantly decreased the IAA content at 70 days after planting. Under RR cultivation, most ABA biosynthesis-related genes were upregulated, while most IAA biosynthesis-related genes were downregulated. These findings lay a solid foundation for further establishing the network through which miRNAs regulate grapevine root development through target genes and for further exploring the molecular mechanism through which endogenous ABA and IAA regulate root architecture development in grapevine.

Effects of root restriction on phytohormone levels in different growth stages and grapevine organs.

Phytohormones play important roles in germination, blossom, senescence, abscission of plants by a series of signal transduction and molecular regulation. The purpose of this research was to investigate the influence of root restriction (RR) cultivation on plant endogenous hormone variation tendency at different growth stages in diverse organs or tissues. 'Muscat Hamburg' (Vitis 'Muscat of Alexandria' × Vitis 'Trollinger') grapevine was used as test material. High Performance Liquid Chromatography (HPLC) was used to quantify hormone levels, qRT-PCR was used to quantify the expression of genes related to hormone biosynthesis pathway, and determined parameters of growth and photosynthetic, aiming to investigate the influence of root restriction on the formation and metabolism of phytohormones, as well as the degree of correlation between phytohormones and plant growth and photosynthetic intensity under root restriction. By measuring the photosynthetic rate of leaves at the stages of core-hardening, veraison and maturity, it was found that root restriction could reduce most photosynthetic parameters. The results also revealed that RR treatment increased abscisic acid (ABA), salicylic acid (SA), zeatin riboside (ZR), N6-(delta 2-isopentenyl)-adenine nucleoside (iPR) concentrations, while reduced auxin (IAA), 3-indolepropionic acid (IPA), 3-indolebutyric acid (IBA), gibberellin A3 (GA3), zeatin (ZT), N6-(delta 2-Isopentenyl)-adenine (iP), kinetin (KT), jasmonic acid (JA) and methyl jasmonate (MeJA) concentrations in most organs and at most developmental stages. RT-qPCR was carried out to further explore the effect of root restriction on genes expression of ABA, SA and IAA biosynthesis pathways at molecular level. Meanwhile, through correlation analysis, we found that different phytohormones contributed differently to physiological indicators, there existed strong correlation of ABA, KT, MeJA, iPR, SA, JA with leaf photosynthesis, GA3, IBA, ZR, IAA, ZT with fruit quality. In addition, we also found that the shoot growth related parameters were closely correlated with JA, IPA and iP. To sum up, our results suggested that RR treatment could significantly increase soluble solid content, regulate the growth and photosynthesis of grapevine, by affecting the biosynthesis of phytohormones. It could further prove that root restriction was a feasible technique to ameliorate the phenomenon of low quality in grape berry in southern China.

Productive HIV infection in astrocytes can be established via a nonclassical mechanism.

OBJECTIVE: Astrocytes are proposed to be a critical reservoir of HIV in the brain. However, HIV infection of astrocytes is inefficient in vitro except for cell-to-cell transmission from HIV-infected cells. Here, we explore mechanisms by which cell-free HIV bypasses entry and postentry barriers leading to a productive infection. METHODS: HIV infection of astrocytes was investigated by a variety of techniques including transfection of CD4-expressing plasmid, treatment with lysosomotropic agents or using a transwell culture system loaded with HIV-infected lymphocytes. Infection was monitored by HIV-1 p24 in culture supernatants and integrated proviral DNA was quantified by Alu-PCR. RESULTS: Persistent HIV infection could be established in astrocytes by transfection of proviral DNA, transduction with VSV-G-pseudotyped viruses, transient expression of CD4 followed by HIV infection, or simultaneous treatment with lysosomotropic chloroquine or Tat-HA2 peptide with HIV infection. In absence of these treatments, HIV entered via endocytosis as seen by electronmicroscopy and underwent lysosomal degradation without proviral integration, indicating endocytosis is a dead end for HIV in astrocytes. Nevertheless, productive infection was observed when astrocytes were in close proximity but physically separated from HIV-infected lymphocytes in the transwell cultures. This occurred with X4 or dual tropic R5X4 viruses and was blocked by an antibody or antagonist to CXCR4. CONCLUSION: A CD4-independent, CXCR4-dependent mechanism of viral entry is proposed, by which immature HIV particles from infected lymphocytes might directly bind to CXCR4 on astrocytes and trigger virus--cell fusion during or after the process of viral maturation. This mechanism may contribute to the formation of brain HIV reservoirs.

Astrocytes as an HIV Reservoir: Mechanism of HIV Infection.

If we have any hope of achieving a cure for HIV infection, close attention to the cell types capable of getting infected with HIV is necessary. Of these cell types, astrocytes are the most ideal cell type for the formation of such a reservoir. These are long-lived cells with a very low turnover rate and are found in the brain and the gastrointestinal tract. Although astrocytes are evidently resistant to infection of cell-free HIV in vitro, these cells are efficiently infected via cell-tocell contact by which immature HIV virions bud off lymphocytes and have the ability to directly bind to CXCR4, triggering the process of fusion in the absence of CD4. In this review, we closely examine the evidence for HIV infection of astrocytes in the brain and the mechanisms for viral entry and regulation in this cell type, and discuss an approach for controlling this viral reservoir.

Cell-to-cell contact facilitates HIV transmission from lymphocytes to astrocytes via CXCR4.

OBJECTIVES: HIV reservoir in the brain represents a major barrier for curing HIV infection. As the most abundant, long-lived cell type, astrocytes play a critical role in maintaining the reservoir; however, the mechanism of infection remains unknown. Here, we determine how viral transmission occurs from HIV-infected lymphocytes to astrocytes by cell-to-cell contact. DESIGN AND METHODS: Human astrocytes were exposed to HIV-infected lymphocytes and monitored by live-imaging, confocal microscopy, transmission and three-dimensional electron microscopy. A panel of receptor antagonists was used to determine the mechanism of viral entry. RESULTS: We found that cell-to-cell contact resulted in efficient transmission of X4 or X4R5-using viruses from T lymphocytes to astrocytes. In co-cultures of astrocytes with HIV-infected lymphocytes, the interaction occurred through a dynamic process of attachment and detachment of the two cell types. Infected lymphocytes invaginated into astrocytes or the contacts occurred via filopodial extensions from either cell type, leading to the formation of virological synapses. In the synapses, budding of immature or incomplete HIV particles from lymphocytes occurred directly onto the membranes of astrocytes. This cell-to-cell transmission could be almost completely blocked by anti-CXCR4 antibody and its antagonist, but only partially inhibited by anti-CD4, ICAM1 antibodies. CONCLUSION: Cell-to-cell transmission was mediated by a unique mechanism by which immature viral particles initiated a fusion process in a CXCR4-dependent, CD4-independent manner. These observations have important implications for developing approaches to prevent formation of HIV reservoirs in the brain.

Development of fluorescent probes based on protection-deprotection of the key functional groups for biological imaging.

Recently, the strategy of protection-deprotection of functional groups has been widely employed to design fluorescent probes, as the protection-deprotection of functional groups often induces a marked change in electronic properties. Significant advances have been made in the development of analyte-responsive fluorescent probes based on the protection-deprotection strategy. In this tutorial review, we highlight the representative examples of small-molecule based fluorescent probes for bioimaging, which are operated via the protection-deprotection of key functional groups such as aldehyde, hydroxyl, and amino functional groups reported from 2010 to 2014. The discussion includes the general protection-deprotection methods for aldehyde, hydroxyl, or amino groups, as well as the design strategies, sensing mechanisms, and deprotection modes of the representative fluorescent imaging probes applied to bio-imaging.

Interaction of paroxetine with mitochondrial proteins mediates neuroprotection.

There are severe neurological complications that arise from HIV infection, ranging from peripheral sensory neuropathy to cognitive decline and dementia for which no specific treatments are available. The HIV proteins secreted from infected macrophages, gp120 and Tat, are neurotoxic. The goal of this study was to screen, identify and develop neuroprotective compounds relevant to HIV-associated neurocognitive disorders (HAND). We screened more than 2000 compounds that included FDA approved drugs for protective efficacy against oxidative stress-mediated neurodegeneration and identified selective serotonin reuptake inhibitors (SSRIs) as potential neuroprotectants. Numerous SSRIs were then extensively evaluated as protectants against neurotoxicity as measured by changes in neuronal cell death, mitochondrial potential, and axodendritic degeneration elicited by HIV Tat and gp120 and other mitochondrial toxins. While many SSRIs demonstrated neuroprotective actions, paroxetine was potently neuroprotective (100 nM potency) against these toxins in vitro and in vivo following systemic administration in a gp120 neurotoxicity model. Interestingly, the inhibition of serotonin reuptake by paroxetine was not required for neuroprotection, since depletion of the serotonin transporter had no effect on its neuroprotective properties. We determined that paroxetine interacts selectively and preferentially with brain mitochondrial proteins and blocks calcium-dependent swelling but had less effect on liver mitochondria. Additionally, paroxetine induced proliferation of neural progenitor cells in vitro and in vivo in gp120 transgenic animals. Therefore, SSRIs such as paroxetine may provide a novel adjunctive neuroprotective and neuroregenerative therapy to treat HIV-infected individuals.

Three-dimensional imaging of HIV-1 virological synapses reveals membrane architectures involved in virus transmission.

UNLABELLED: HIV transmission efficiency is greatly increased when viruses are transmitted at virological synapses formed between infected and uninfected cells. We have previously shown that virological synapses formed between HIV-pulsed mature dendritic cells (DCs) and uninfected T cells contain interdigitated membrane surfaces, with T cell filopodia extending toward virions sequestered deep inside invaginations formed on the DC membrane. To explore membrane structural changes relevant to HIV transmission across other types of intercellular conjugates, we used a combination of light and focused ion beam scanning electron microscopy (FIB-SEM) to determine the three-dimensional (3D) architectures of contact regions between HIV-1-infected CD4(+) T cells and either uninfected human CD4(+) T cells or human fetal astrocytes. We present evidence that in each case, membrane extensions that originate from the uninfected cells, either as membrane sheets or filopodial bridges, are present and may be involved in HIV transmission from infected to uninfected cells. We show that individual virions are distributed along the length of astrocyte filopodia, suggesting that virus transfer to the astrocytes is mediated, at least in part, by processes originating from the astrocyte itself. Mechanisms that selectively disrupt the polarization and formation of such membrane extensions could thus represent a possible target for reducing viral spread. IMPORTANCE: Our findings lead to new insights into unique aspects of HIV transmission in the brain and at T cell-T cell synapses, which are thought to be a predominant mode of rapid HIV transmission early in the infection process.

Molecular mechanisms in the dramatic enhancement of HIV-1 Tat transduction by cationic liposomes.

Human immunodeficiency virus type 1 (HIV-1) transactivator of transcription (Tat) protein possesses a unique membrane-transduction property. Interestingly, Tat transduction could be dramatically increased 1000-fold based on LTR-transactivation assay when complexed with cationic liposomes (lipo-Tat), compared with Tat alone. Therefore, underlining mechanisms were explored further. Microscopy and flow cytometry showed that this effect was associated with enhanced membrane binding, large particle formation (1-2 µm) and increased intracellular uptake of Tat fluorescent proteins. Using pharmacological assays and immune colocalizations, it was found that lipid raft-dependent endocytosis and macropinocytosis were major pathways involved in lipo-Tat uptake, and actin-filaments played a major role in intracellular trafficking of lipo-Tat to the nucleus. Furthermore, we found that the Tat hydrophobic domain (aa 36-47) mediated formation of two positively charged molecules into lipo-Tat complexes via hydrophobic bonds, based on LTR-transactivation inhibition assay. Thus, the hydrophobic domain may play an important role in Tat protein uptake and be useful for intracellular delivery of biomacromolecules if coupled together with Tat basic peptide, a cell-penetrating peptide.

Role of Tat protein in HIV neuropathogenesis.

The Tat protein of the human immunodeficiency virus (HIV) has been implicated in the pathophysiology of the neurocognitive deficits associated with HIV infection. This is the earliest protein to be produced by the proviral DNA in the infected cell. The protein not only drives the regulatory regions of the virus but may also be actively released from the cell and then interact with the cell surface receptors of other uninfected cells in the brain leading to cellular dysfunction. It may also be taken up by these cells and can then activate a number of host genes. The Tat protein is highly potent and has the unique ability to travel along neuronal pathways. Importantly, its production is not impacted by the use of antiretroviral drugs once the proviral DNA has been formed. This article reviews the pleomorphic actions of Tat protein and the evidence supporting its central role in the neuropathogenesis of the HIV infection.

Lipid Nanoparticles with Accessible Nickel as a Vaccine Delivery System for Single and Multiple His-tagged HIV Antigens.

Lipid-based nanoparticles (NPs) with a small amount of surface-chelated nickel (Ni-NPs) were developed to easily formulate the HIV his-tagged Tat protein, as well as to formulate and co-deliver two HIV antigens (his-p24 and his-Nef) on one particle. Female BALB/c mice were immunized by s.c. injection with his-Tat/Ni-NP formulation (1.5 µg Tat-his/mouse) and control formulations on day 0 and 14. The day 28 anti-Tat specific IgG titer with his-Tat/Ni-NP was significantly greater than that with Alum/his-Tat. Furthermore, splenocytes from his-Tat/Ni-NP immunized mice secreted significantly higher IFN-γ than those from mice immunized with Alum/his-Tat. Although Ni-NPs did not show better adjuvant activity than Tat-coated anionic NPs made with sodium dodecyl sulfate (SDS/NPs), they were less toxic than SDS/NPs. The initial results indicated that co-immunization of mice using his-p24/his-Nef/Ni-NP induced greater antibody response compared to using Alum/his-p24/his-Nef. Co-delivery of two antigens using Ni-NPs also increased the immunogenicity of individual antigens compared to delivery of a single antigen by Ni-NPs. In conclusion, Ni-NPs are an efficient delivery system for HIV vaccines including both single antigen delivery and multiple antigen co-delivery.

Ceftriaxone protects against the neurotoxicity of human immunodeficiency virus proteins.

Human immunodeficiency virus (HIV) proteins Tat and gp120 have been implicated in the pathogenesis of HIV dementia by various mechanisms, including down-regulation of excitatory amino acid transporter-2 (EAAT2), which is responsible for inactivation of synaptic glutamate. Recent work indicates that beta-lactam antibiotics are potent stimulators of EAAT2 expression. The authors treated mixed human fetal neuronal cultures with recombinant gp120 or Tat, in the presence or absence of ceftriaxone, and determined neurotoxicity by measuring mitochondrial membrane potential and neuronal cell death. Ceftriaxone produced dose-dependent attenuation of the neurotoxicity and neuronal cell death caused by both viral proteins. This study demonstrates that this class of drugs may have therapeutic efficacy in HIV dementia.

[Study on the distribution of human immunodeficiency virus-1 subtypes in different regions of China and mother-to-child transmission].

OBJECTIVE: To study the distribution of human immunodeficiency virus (HIV)-1 genotypes in major prevalent regions of China and to illustrate the relationship between HIV-1 subtypes and mother-to-child transmission in a retrospective cohort. METHODS: HIV-1 gag p17 and env C2-V4 region were amplified by nested-polymerase chain reaction (nPCR) and the sequences were obtained by sequencing gag nPCR products or clones of env gene. RESULTS: 60 HIV-1 positive individuals were subject to typing for gag p17 and 69 for env C2-V4 region. Single clade was only found in Henan (subtype B') and Xinjiang (subtype C), and subtypes C and E were demonstrated in Yunnan. These regions represented most of the HIV-1 infections in China. Multiple subtypes (A, B, C, E, etc.) were found in Beijing and Shanghai, where HIV infections were still in low level. The sequences of subtype C were less diversive in Xinjiang (p17: 0.0192 +/- 0.0078, C2-V4: 0.0455 +/- 0.0145) than in Yunnan (p17: 0.0279 +/- 0.0102, C2-V4: 0.0482 +/- 0.0171), but all of them clustered in "C" branch in phylogenetic trees. Trafficking of subtype C from Yunnan to Xinjiang was found but had already been reported by others. Compared to subtype C, subtype E was quite divergent (p17: 0.0473 +/- 0.0105, C2-V4: 0.1114 +/- 0.0112) in Yunnan, but no recombination was found in the C2-V4 region of env gene. Highe divergence of subtype B' was found in Henan and the peripheral provinces (p17: 0.0381 +/- 0.0101, C2-V4: 0.0691 +/- 0.0166), which might be attributed to the early epidemics of HIV-1 in these areas (early 1990's). In maternal-child cohort, subtypes B (7/21), C (11/21), E (1/21) and undefined types (2/21) were identified in non-transmitting HIV-1 positive mothers, while only subtype B (7/11) and C (4/11) appeared in transmitting HIV-1 positive mothers. The rate of transmission was 53.8% (7/13) in mothers infected with subtype B and 30.8% (4/13) in those infected with subtype C, but with no significant difference (P = 0.196). The imbalancing distribution of subtypes might be explained by the fact that transfusion or illegal blood would increased mother-to-child transmission on HIV-1 and most of mothers with clade B were infected by illegal blood transfusion in this cohort. In addition, most of the maternal-child pair's sequences clustered in gag or env phylogenetic trees but only a few did disperse among the unrelated patients because children were older (>/= 4 years). CONCLUSION: The characteristics of HIV-1 clade's distribution differed over most parts of China but no difference was demonstrated between subtype B and C in mother-to-child transmission on HIV-1.