Extended leaf longevity in the ore4-1 mutant of Arabidopsis with a reduced expression of a plastid ribosomal protein gene

The longevity of plant leaf organs is genetically determined. However, the molecular mechanisms underlying the control of longevity are still largely unknown. Here, we describe a T-DNA-insertional mutation of Arabidopsis thaliana that confers extended leaf longevity. The mutation, termed ore4-1, delays a broad spectrum of age-dependent leaf senescence, but has little effect on leaf senescence artificially induced by darkness, abscisic acid (ABA), methyl jasmonate (MeJA), or ethylene. The T-DNA was inserted within the promoter region of the plastid ribosomal small subunit protein 17 (PRPS17) gene, and this insertion dramatically reduced PRPS17 mRNA expression. In the ore4-1 mutant, the leaf growth rate is decreased, while the maturation timing is similar to that of wild-type. In addition, the activity of the photosystem I (PSI) is significantly reduced in the ore4-1 mutant, as compared to wild-type. Thus, the ore4-1 mutation results in a deficiency in various chloroplast functions, including photosynthesis, which may decrease leaf growth. Our results suggest a possible link between reduced metabolism and extended longevity of the leaf organs in the ore4-1 mutation.     

Specific association of a small protein with the telomeric DNA-protein complex during the onset of leaf senescence in Arabidopsis thaliana

Telomeres and their changes in length throughout the life span of cells have been intensively investigated in different organisms. Telomere length is assumed to control replicative senescence in mammalian cells. However, only very few data are available on the developmental dynamics of plant telomeres. Here, changes of telomere length and DNA-protein structure of Arabidopsis thaliana telomeres were analysed in different stages of development, with the main focus resting on the transition from pre-senescent to senescent leaves. The lengths of the telomeres, ranging from ca. 2.0 to 6.5 kb, do not significantly change during plant development indicating that telomere length is not involved in differentiation and replicative senescence nor in post-mitotic senescence of A. thaliana. In dedifferentiated cultured cells a slight increase in length can be determined. The nucleoprotein structure of the telomeric DNA was investigated by gel mobility shift assays, with synthetic oligonucleotides and nuclear protein extracts derived from four defined stages of post-mitotic leaf senescence. In all four stages, a highly salt-resistant DNA-protein complex was formed with the double-stranded as well as with the single-stranded G-rich telomeric DNA. An additional DNA-protein complex was identified in nuclear protein extracts isolated from plants in the transition stage from pre-senescence to senescence. The protein components of the DNA-protein complexes were analysed on native PAGE and SDS-PAGE gels. A protein of 67 kDa (ATBP1) bound to the telomeric DNA in all developmental stages. An additional protein of merely 22 kDa (ATBP2) was associated via protein-protein interaction with ATBP1 to form a higher-order complex exclusively during the onset of senescence. DNA interaction of this higher-order protein complex seems to be restricted to double-stranded telomeric DNA. The defined period of ATBP1/ATBP2 complex formation with the telomeric DNA probably indicates that ATBP2 is involved in the onset of post-mitotic leaf senescence by either disturbing an established or establishing an additional function exhibited by the telomeres in the interphase nuclei.     

Chromosomal location and expression of the single-copy gene encoding high-mobility-group protein HMG-I/Y in Arabidopsis thaliana

A cDNA encoding the HMG-I/Y protein from Arabidopsis thaliana has been isolated and characterised by nucleotide sequencing. The 903 bp cDNA contains a 612 bp open reading frame encoding a protein of 204 amino acid residues showing homology to HMG-I/Y proteins from other plant species. The protein contains four copies of the AT-hook motif which is involved in binding A/T-rich DNA. Southern blotting showed that the HMG-I/Y gene was present in a single copy in the Arabidopsis genome. The gene was localised to the top of chromosome 1 by RFLP analysis of F₈ recombinant inbred lines. Northern blotting showed that the gene was expressed in all organs examined, with the highest expression in flowers and developing siliques.     

Isolation and partial characterization of a protein kinase NII from wheat germ chromatin

A protein kinase, type NII, has been purified from wheat germ chromatin. The enzyme, which uses both ATP and GTP as phosphoryl donors, catalyzes the phosphorylation of casein, phosvitin and E. coli RNA polymerase, but not of histone proteins. Polypeptide bands at 46 kDa, 37 kDa and 25 kDa were estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Autophosphorylation of the 25 kDa subunit was observed following incubation of the purified kinase with (-³²P)ATP and (-³²P)GTP.     

Differential changes in the synthesis and steady-state levels of thylakoid proteins during bean leaf senescence

During senescence of primary bean leaves (Phaseolus vulgaris), there are differential changes in the rates at which thylakoid proteins are synthesized. In particular, synthesis of the 32 kD herbicide-binding protein continues throughout senescence, whereas formation of the and subunits of ATPase, the 68 kD photosystem I reaction center polypeptide, cytochrome f, cytochrome b₆ and the structural apoprotein of the lightharvesting chlorophyll protein complex (LHCP) declines. Pulse-chase experiments with intact leaves indicated rapid degradation of the 32 kD protein, which is consistent with its known rapid rate of turnover. This degradation was light-dependent and inhibited by DCMU, and the kinetics of degradation were similar for young and senescent membranes. In Coomassie-stained gels, the 68 kD reaction center polypeptide of photosystem I, the and subunits of ATPase and the LHCP were the dominant proteins for all ages of membranes. Western blot analysis indicated that cytochrome f and cytochrome b₆ are selectively depleted during senescence. The data have been interpreted as indicating that translational disruptions in both the cytoplasmic and chloroplastic compartments may contribute to the decline in photosynthetic electron transport in the senescing leaf.     

Berberine ameliorates cellular senescence and extends the lifespan of mice via regulating p16 and cyclin protein expression

Although aging and senescence have been extensively studied in the past few decades, however, there is lack of clinical treatment available for anti‐aging. This study presents the effects of berberine (BBR) on the aging process resulting in a promising extension of lifespan in model organisms. BBR extended the replicative lifespan, improved the morphology, and boosted rejuvenation markers of replicative senescence in human fetal lung diploid fibroblasts (2BS and WI38). BBR also rescued senescent cells with late population doubling (PD). Furthermore, the senescence‐associated β‐galactosidase (SA‐β‐gal)‐positive cell rates of late PD cells grown in the BBR‐containing medium were ~72% lower than those of control cells, and its morphology resembled that of young cells. Mechanistically, BBR improved cell growth and proliferation by promoting entry of cell cycles from the G₀ or G₁ phase to S/G₂‐M phase. Most importantly, BBR extended the lifespan of chemotherapy‐treated mice and naturally aged mice by ~52% and ~16.49%, respectively. The residual lifespan of the naturally aged mice was extended by 80%, from 85.5 days to 154 days. The oral administration of BBR in mice resulted in significantly improved health span, fur density, and behavioral activity. Therefore, BBR may be an ideal candidate for the development of an anti‐aging medicine.     

A modified storage protein is synthesized,processed, and degraded in the seeds of transgenic plants

In vitro mutagenesis was used to supplement the sulfur amino acid codon content of a gene encoding -phaseolin, a Phaseolus vulgaris storage protein. The number of methionine codons in the phaseolin gene was increased from three to nine by insertion of a 45 base pair (bp) synthetic duplex. Either modified or normal phaseolin genes were integrated into the genome of tobacco plants through Agrobacterium tumefaciens-mediated transformation. Although similar levels of phaseolin RNA are detected in seeds of plants transformed with either the normal or modified (himet) gene, the quantity of himet protein is consistently much lower than normal -phaseolin. Himet phaseolin is expressed in a temporal- and organ-specific fashion, and is N-glycosylated and assembled into trimers in the manner of normal phaseolin. After germination, both types of phaseolin are hydrolyzed, but the himet protein is more quickly degraded. Electron microscopic immunocytochemical observations of developing seeds indicate that the himet protein is primarily localized in the endoplasmic reticulum (ER) and in Golgi apparatus secretion vesicles. Himet phaseolin is absent from protein storage vacuoles, termed protein bodies, where normal phaseolin is deposited in transgenic tobacco. We interpret the immunocytochemical data to indicate that himet phasolin is transported through the ER and Golgi apparatus and is then degraded in Golgi secretion vesicles or the protein bodies.Mention of trademark, proprietary product, or vendor does not constitute a guarantee of warranty of the product by the United States Department of Agriculture and does not imply its approval to the exclusion of other products or vendors that may also be suitable. Publication No. 70 from Agrigenetics Advanced Science Company.     

Protein sorting and expression of a unique soybean cotyledon protein,GmSBP, destined for the protein storage vacuole

The initial biochemical characterization of the soybean sucrose-binding protein, GmSBP, within our lab and others produced several incongruous characteristics that required a re-characterization of GmSBP via sequence homology, cell biology, immunolocalization, and semi-quantitative analysis. The GmSBP proteins share amino acid sequence homology as well as putative structural homology with globulin-like seed storage proteins. A comparison to the major soybean seed storage proteins, glycinin and -conglycinin established several storage protein-like characteristics for GmSBP. All three proteins were present in a prevacuolar compartment and protein storage vacuole. All three proteins increased in expression during seed development and are remobilized during germination. Quantitatively, the relative concentrations of GmSBP, -conglycinin (/ subunits), and glycinin (acidic subunits) indicated that GmSBP contributes 19-fold less to the stored nitrogen. The quantitative differences between GmSBP and glycinin may be attributed to the unconserved order and spacing of cis-acting regulatory elements present within the promoter regions. Ultimately, GmSBP is transported to the mature protein storage vacuole. The biological function of GmSBP within the protein storage vacuole remains uncertain, but its localization is a remnant of its evolutionary link to a globulin-like or vicilin-like ancestor that gave rise to the 7S family of storage proteins.     

Overexpression of the rice Osmyb4 gene increases chilling and freezing tolerance of Arabidopsis thaliana plants

The expression of the gene Osmyb4, detected at low level in rice (Oryza sativa) coleoptiles grown for 3 days at 29 degrees C, is strongly induced by treatments at 4 degrees C. At sublethal temperatures of 10 and 15 degrees C, its expression in rice seedlings is already evident, but this effect cannot be vicariated by other stresses or ABA treatment. We demonstrate by transient expression that Myb4 transactivates the PAL2, ScD9 SAD and COR15a cold-inducible promoters. The Osmyb4 function in vivo is demonstrated overexpressing its cDNA in Arabidopsis thaliana plants (ecotype Wassilewskija) under the control of the constitutive CaMV 35S promoter. Myb4 overexpressing plants show a significant increased cold and freezing tolerance, measured as membrane or Photosystem II (PSII) stability and as whole plant tolerance. Finally, in Osmyb4 transgenic plants, the expression of genes participating in different cold-induced pathways is affected, suggesting that Myb4 represents a master switch in cold tolerance.     

Mutations in the Arabidopsis SWC6 gene, encoding a component of the SWR1 chromatin remodelling complex, accelerate flowering time and alter leaf and flower development

Mutations affecting the Arabidopsis SWC6 gene encoding a putativeorthologue of a component of the SWR1 chromatin remodellingcomplex in plants have been characterized. swc6 mutations causeearly flowering, shortened inflorescence internodes, and alteredleaf and flower development. These phenotypic defects resemblethose of the photoperiod independent early flowering 1 (pie1)and early in short days 1 (esd1) mutants, also affected in homologuesof the SWR1 complex subunits. SWC6 is a ubiquitously expressednuclear HIT-Zn finger-containing protein, with the highest levelsfound in pollen. Double mutant analyses suggest that swc6 abolishesthe FLC-mediated late-flowering phenotype of plants carryingactive alleles of FRI and of mutants of the autonomous pathway.It was found that SWC6 is required for the expression of theFLC repressor to levels that inhibit flowering. However, theeffect of swc6 in an flc null background and the down-regulationof other FLC-like/MAF genes in swc6 mutants suggest that floweringinhibition mediated by SWC6 occurs through both FLC- and FLC-likegene-dependent pathways. Both genetic and physical interactionsbetween SWC6 and ESD1 have been demonstrated, suggesting thatboth proteins act in the same complex. Using chromatin immunoprecipitation,it has been determined that SWC6, as previously shown for ESD1,is required for both histone H3 acetylation and H3K4 trimethylationof the FLC chromatin. Altogether, these results suggest thatSWC6 and ESD1 are part of an Arabidopsis SWR1 chromatin remodellingcomplex involved in the regulation of diverse aspects of plantdevelopment, including floral repression through the activationof FLC and FLC-like genes. Key words: Arabidopsis, chromatin remodelling, floral repression, HIT-Zn finger, phase transition, SWR1 complex     

Overexpression of AtPTPA in Arabidopsis increases protein phosphatase 2A activity by promoting holoenzyme formation and ABA negatively affects holoenzyme formation

AtPTPA is a critical regulator for the holoenzyme assembling of protein phosphatase 2A (PP2A) in Arabidopsis. Characterization of AtPTPA improves our understanding of the function and regulation of PP2A in eukaryotes. Further analysis of AtPTPA-overexpressing plants indicates that AtPTPA increases PP2A activity by promoting PP2A''s AC dimer formation, thereby holoenzyme assembling. Plant hormone abscisic acid (ABA) reduces PP2A enzyme activity by negatively affects PP2A''s AC dimer formation. Therefore, AtPTPA is a positive factor that promotes PP2A holoenzyme assembly, and ABA is a negative factor that prevents PP2A holoenzyme assembly.     

Synthesis and degradation of a 28-kDa pod storage protein in French bean (Phaseolus vulgaris) plants

Pod storage protein (PSP) accumulated in developing pods of French bean (Phaseolus vulgaris L.) plants, and increasing the PSP mRNA level by pod removal resulted in the enhancement of PSP accumulation in pods that formed later. Pod storage protein was detected in flowers, young leaves and young stem internodes in addition to pods. Accumulation of PSP and its mRNA was induced by sink-removal in an organ-specific manner. In addition, wounding induced PSP accumulation systemically in leaves. Methyl jasmonate did not induce PSP synthesis but enhanced the synthesis that was induced by wounding. In senescing pods, PSP was degraded, and degradation products with molecular masses of 20 and 17 kDa were detected in the pods. The amount of 20-kDa degradation product was greater than that of the 17 kDa product. Received: 26 May 1999 / Accepted: 24 June 1999     

Crystal structure of the crenarchaeal conserved chromatin protein Cren7 and double-stranded DNA complex

Cren7 is a crenarchaeal conserved chromatin protein discovered recently. To explore the mechanism of the DNA packaging in Crenarchaeota, the crystal structure of Cren7–GCGATCGC complex has been determined and refined at 1.6 Å resolution. Cren7 kinks the dsDNA sharply similar to Sul7d, another chromatin protein existing only in Sulfolobales, which reveals that the “bending and unwinding” compacting mechanism is conserved in Crenarchaeota. Significant structural differences are revealed by comparing both protein–dsDNA complexes. The kinked sites on the same dsDNA in the complexes with Sul7d and Cren7 show one base pair shift. For Cren7, fewer charged residues in the β‐barrel structural region bind to DNA, and additionally, the flexible loop L_β3β4 is also involved in the binding. Electrophoretic mobility shift assays indicate that loop L_β3β4 is essential for DNA‐binding of Cren7. These differences provide insight into the functional difference of both chromatin proteins, suggesting that Cren7 may be more regulative than Sul7d in the DNA‐binding affinity by the methylation in the flexible loop L_β3β4 in vivo.     

Optimum leaf excision increases the biomass accumulation and seed yield of maize plants under different planting patterns

Without developing new agronomic practices, present rates of improvement in seed yields of cereal crops globally are insufficient to fulfil the estimated increasing food demand for 2050 and beyond. Intercropping is one of the agricultural practices that can lead to greater crop yields. However, there exists leaf redundancy for maize in intercropping systems, and the top canopy leaves shade more competent leaves at middle strata of maize plants. Therefore, this work aimed to elucidate the effect of leaf excision treatments in maize to understand the optimum leaf area of maize plants under a maize–soybean relay‐intercropping system (MS_R) and a sole cropping system (SM). The effects of four‐leaf excision treatments (T₁, 0; T₂, 2; T₃, 4; T₄, 6 leaves excised from the top of maize plants until 7 days after silking) on light interception, leaf area index (LAI), photosynthetic characteristics, total biomass accumulation at blistering stage (BS), dough stage (DS) and physiological maturity (PM), and seed yield of maize were investigated through field experiments for 2 years under MS_R and SM. Results showed that, under MS_R and SM, as compared to control (T₁), optimum excision of leaves (T₂) from the top of maize plants significantly improved the light interception (by 25, 18 and 16% at BS, DS and PM, respectively) to lower strata leaves and accelerated the biomass partitioning to maize seeds (by 13 and 12% at DS and PM, respectively). Importantly, plants under T₂ exhibited higher green leaf area than control, that is, excision the top two leaves led to an increase in LAI at PM by 10%, suggesting that leaf senescence under T₂ was delayed which enhanced the photosynthetic rate at PM by 7% in 2017 and 6% in 2018. Relative to T₁, maize under T₂ produced 19 and 13% higher maize yield under MS_R and SM, respectively, and relay‐cropped maize had 90% of SM seed yield. These results suggest that by manipulating the canopy structure of maize plants we can enhance the biomass accumulation and seed yield of maize crops under MS_R and SM.