ProMetheusDB: An In-Depth Analysis of the High-Quality Human Methyl-proteome

Protein arginine (R) methylation is a post-translational modification involved in various biological processes, such as RNA splicing, DNA repair, immune response, signal transduction, and tumor development. Although several advancements were made in the study of this modification by mass spectrometry, researchers still face the problem of a high false discovery rate. We present a dataset of high-quality methylations obtained from several different heavy methyl stable isotope labeling with amino acids in cell culture experiments analyzed with a machine learning–based tool and show that this model allows for improved high-confidence identification of real methyl-peptides. Overall, our results are consistent with the notion that protein R methylation modulates protein–RNA interactions and suggest a role in rewiring protein–protein interactions, for which we provide experimental evidence for a representative case (i.e., NONO [non-POU domain–containing octamer-binding protein]–paraspeckle component 1 [PSPC1]). Upon intersecting our R-methyl-sites dataset with the PhosphoSitePlus phosphorylation dataset, we observed that R methylation correlates differently with S/T-Y phosphorylation in response to various stimuli. Finally, we explored the application of heavy methyl stable isotope labeling with amino acids in cell culture to identify unconventional methylated residues and successfully identified novel histone methylation marks on serine 28 and threonine 32 of H3. The database generated, named ProMetheusDB, is freely accessible at https://bioserver.ieo.it/shiny/app/prometheusdb.     

SDS-capped 1-pyrenecarboxaldehyde nanoprobe for selective detection of Cu2+ ion from water samples: Spectroscopic approach

Sodium dodecyl sulfate (SDS)-capped 1-pyrenecarboxaldehyde nanoparticles (PyalNPs) were prepared using a reprecipitation method in an aqueous medium and exhibited red-shifted aggregation-induced enhanced emission (AIEE). The dynamic light scattering (DLS) examination showed narrower particle size distribution with an average particle size of 41 nm, whereas −34.5 mV zeta potential value indicate the negative surface charge and good stability of nanoparticles (NPs) in an aqueous medium. The AIEE was seen at λ_max = 473 nm in a fluorescence spectrum of a PyalNP suspension. In the presence of Cu²⁺ ions, the fluorescence of PyalNPs quenches very significantly, even in the presence of other metal ions like Ba²⁺, Ca²⁺, Cd²⁺, Co²⁺, Al³⁺, Fe²⁺, Hg²⁺, Ni²⁺ and Mg²⁺. The changes in the fluorescence lifetime of PyalNPs in the presence of Cu²⁺ ions suggested that the type of quenching was dynamic. The fluorescence quenching data for the NPs suspension fitted well into a typical Stern–Volmer relationship in the concentration range 1.0–25 μg/ml of Cu²⁺ ions. The estimated value of the correlation coefficient R² = 0.9877 was close to 1 and showed the linear relationship between quenching data and Cu²⁺ ion concentration. The limit of detection (LOD) was found to be 0.94 ng/ml and is far below the tolerable intake limit value of 1.3 μg/ml accepted by the World Health Organization for Cu²⁺ ions in drinking water. The fluorescence quenching approach for a SDS-capped Pyal nanosuspension for copper ion quantification is of high specificity and coexisting ions were found to interfere very negligibly. The developed method was successfully applied for the estimation of copper ions in river water samples.     

SAUR proteins and PP2C.D phosphatases regulate H+-ATPases and K+ channels to control stomatal movements

Activation of plasma membrane (PM) H⁺-ATPase activity is crucial in guard cells to promote light-stimulated stomatal opening, and in growing organs to promote cell expansion. In growing organs, SMALL AUXIN UP RNA (SAUR) proteins inhibit the PP2C.D2, PP2C.D5, and PP2C.D6 (PP2C.D2/5/6) phosphatases, thereby preventing dephosphorylation of the penultimate phosphothreonine of PM H⁺-ATPases and trapping them in the activated state to promote cell expansion. To elucidate whether SAUR–PP2C.D regulatory modules also affect reversible cell expansion, we examined stomatal apertures and conductances of Arabidopsis thaliana plants with altered SAUR or PP2C.D activity. Here, we report that the pp2c.d2/5/6 triple knockout mutant plants and plant lines overexpressing SAUR fusion proteins exhibit enhanced stomatal apertures and conductances. Reciprocally, saur56 saur60 double mutants, lacking two SAUR genes normally expressed in guard cells, displayed reduced apertures and conductances, as did plants overexpressing PP2C.D5. Although altered PM H⁺-ATPase activity contributes to these stomatal phenotypes, voltage clamp analysis showed significant changes also in K⁺ channel gating in lines with altered SAUR and PP2C.D function. Together, our findings demonstrate that SAUR and PP2C.D proteins act antagonistically to facilitate stomatal movements through a concerted targeting of both ATP-dependent H⁺ pumping and channel-mediated K⁺ transport.

SMALL AUXIN UP RNA (SAUR) proteins and PP2C.D phosphatases antagonistically regulate stomatal aperture in Arabidopsis by modulating the activities of plasma membrane H⁺-ATPases and K⁺ channels.     

Active oxygen species metabolism in neem (<Emphasis Type="Italic">Azadirachta indica</Emphasis>) seeds exposed to natural ageing and controlled deterioration

Success of ex situ storage of germplasm for trade and conservation essentially depends upon the precision of the protocol employed for the assessment of germination potential. Active oxygen species and antioxidative enzymes during natural ageing (NA) and controlled deterioration (CD) was monitored during the loss of seed vigour and germination potential in neem seeds showing intermediate seed storage behaviour. Higher levels of SOD, CAT and APX were strongly and positively associated with germination and vigour. The loss of CAT and APX activity estimated quantitatively and number of isoenzymes were closely accompanied with the simultaneous increase in the amounts of H₂O₂ and OH-radical. The decline in germination and vigour was negatively related with the levels of H₂O₂ and OH-radical and enhanced electrolyte leakage. The amounts of OH-radical were positively correlated with the decline in DNA content and DNA damage. The levels of SOD isoenzymes initially increased as the germination index of seeds declined from 5250 to 762 and 882 under NA and CD conditions, respectively. Increasing activity of SOD in the ageing seeds were associated with the accumulation of H₂O₂. The role of antioxidative enzymes in maintaining signalling and damaging amounts of AOS as well as revelations of different pathways of ageing during NA and CD in the ageing neem seeds were discussed.     

Beneficial mutations for carotenoid production identified from laboratory-evolved Saccharomyces cerevisiae

Journal of Industrial Microbiology & Biotechnology - Adaptive laboratory evolution (ALE) is a powerful tool used to increase strain fitness in the presence of environmental stressors. If...     

Isolation and HPLC assisted quantification of two iridoid glycoside compounds and molecular DNA fingerprinting in critically endangered medicinal Picrorhiza kurroa Royle ex Benth: implications for conservation

Picrorhiza kurroa is a medicinally important, high altitude perennial herb, endemic to the Himalayas. It possesses strong hepato-protective bioactivity that is contributed by two iridoid picroside compounds viz Picroside-I (P-I) and Picroside-II (P-II). Commercially, many P. kurroa based hepato-stimulatory Ayurvedic drug brands that use different proportions of P-I and P-II are available in the market. To identify genetically heterozygous and high yielding genotypes for multiplication, sustained use and conservation, it is essential to assess genetic and phytochemical diversity and understand the population structure of P. kurroa. In the present study, isolation and HPLC based quantification of picrosides P-I and P-II and molecular DNA fingerprinting using RAPD, AFLP and ISSR markers have been undertaken in 124 and 91 genotypes, respectively. The analyzed samples were collected from 10 natural P. kurroa Himalayan populations spread across four states (Jammu & Kashmir, Sikkim, Uttarakhand and Himachal Pradesh) of India. Genotypes used in this study covered around 1000 km geographical area of the total Indian Himalayan habitat range of P. kurroa. Significant quantitative variation ranging from 0.01 per cent to 4.15% for P-I, and from 0.01% to 3.18% in P-II picroside was observed in the analyzed samples. Three molecular DNA markers, RAPD (22 primers), ISSR (15 primers) and AFLP (07 primer combinations) also revealed a high level of genetic variation. The percentage polymorphism and effective number of alleles for RAPD, ISSR and AFLP analysis varied from 83.5%, 80.6% and 72.1%; 1.5722, 1.5787 and 1.5665, respectively. Further, the rate of gene flow (Nm) between populations was moderate for RAPD (0.8434), and AFLP (0.9882) and comparatively higher for ISSR (1.6093). Fst values were observed to be 0.56, 0.33, and 0.51 for RAPD, ISSR and AFLP markers, respectively. These values suggest that most of the observed genetic variation resided within populations. Neighbour joining (NJ), principal coordinate analysis (PCoA) and Bayesian based STRUCTURE grouped all the analyzed accessions into largely region-wise clusters and showed some inter-mixing between the populations, indicating the existence of distinct gene pools with limited gene flow/exchange. The present study has revealed a high level of genetic diversity in the analyzed populations. The analysis has resulted in identification of genetically diverse and high picrosides containing P. kurroa genotypes from Sainj, Dayara, Tungnath, Furkia, Parsuthach, Arampatri, Manvarsar, Kedarnath, Thangu and Temza in the Indian Himalayan region. The inferences generated in this study can be used to devise future resource management and conservation strategies in P. kurroa.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12298-021-00972-w.     

Engineered BcZAT12 gene mitigates salt stress in tomato seedlings

In salt-prone areas, plant growth and productivity is adversely affected. In the present study, the ZT1-ZT6 transgenic tomato lines having BcZAT12 gene under the regulatory control of the stress inducible Bclea1 promoter were exposed to three salinity levels (50, 100 and 200 mM) at the four leaf stage for 10 days. The transgenic lines showed improved growth in stem height, leaf area, root length and shoot length under saline conditions, as compared to control. Moreover, ZT1 and ZT5 lines showed lower electrolyte leakage and decreased hydrogen peroxide formation, in combination with elevated relative water content, proline and chlorophyll levels. The enzyme activity of catalase was also enhanced in ZT1 and ZT5. These results poses the present lines as an attractive alternative for tomato cultivation in salinity-affected areas.Keyword: Abiotic stress, Antioxidant, Oxidative stress, Salinity, Seedlings, Tomato     

Pyridinium-2-carbaldoximes with quinolinium carboxamide moiety are simultaneous reactivators of acetylcholinesterase and butyrylcholinesterase inhibited by nerve agent surrogates

The pyridinium-2-carbaldoximes with quinolinium carboxamide moiety were designed and synthesised as cholinesterase reactivators. The prepared compounds showed intermediate-to-high inhibition of both cholinesterases when compared to standard oximes. Their reactivation ability was evaluated in vitro on human recombinant acetylcholinesterase (hrAChE) and human recombinant butyrylcholinesterase (hrBChE) inhibited by nerve agent surrogates (NIMP, NEMP, and NEDPA) or paraoxon. In the reactivation screening, one compound was able to reactivate hrAChE inhibited by all used organophosphates and two novel compounds were able to reactivate NIMP/NEMP-hrBChE. The reactivation kinetics revealed compound 11 that proved to be excellent reactivator of paraoxon-hrAChE better to obidoxime and showed increased reactivation of NIMP/NEMP-hrBChE, although worse to obidoxime. The molecular interactions of studied reactivators were further identified by in silico calculations. Molecular modelling results revealed the importance of creation of the pre-reactivation complex that could lead to better reactivation of both cholinesterases together with reducing particular interactions for lower intrinsic inhibition by the oxime.