Disease relevance of Protoplasts

• A transferred DNA (T-DNA) tagging vector with the potential to produce dominant mutations was used with cocultured Agrobacterium tumefaciens and protoplasts to tag genes involved in the action of the plant growth substance auxin [1].
• Efficiently regenerated single colonies from mixed multiply auxotrophic Bacillus subtilis protoplasts, fused with polyethylene glycol, reveal colonies carrying each of the parent types (biparentals) and recombinant colonies [2].
• Coexpression of HSP70h and the p61 protein with CPm in protoplasts restricted encapsidation to the 5' approximately 630 nucleotides, which is close to the normal boundary of the bipolar virion, whereas the presence of either HSP70h or the p61 protein alone did not limit encapsidation by CPm [3].
• One Ltk- cell in 300 to 500 was transformed to the TK+ phenotype after fusion with protoplasts carrying the chimeric plasmid pX1, which consists of pBR322 and the BamHI fragment coding for the herpes simplex virus type 1 thymidine kinase gene [4].
• The effect of the 5'-dephosphorylated 2',5'-adenylate trimer and its 2',5'-trimer core analogs on the inhibition of tobacco mosaic virus (TMV) replication was determined in tobacco leaf discs, protoplasts, and whole tobacco plants, using infectivity tests and enzyme-linked immunosorbent assays [5].

High impact information on Protoplasts

• Transient expression of eIF3g and L24 in plant protoplasts strongly affects TAV-mediated reinitiation activity [6].
• A factor in conditioned medium from B. subtilis cells engineered to produce SpollR during growth triggered processing in protoplasts of B. subtilis cells that had been engineered to produce SpollGA and pro-sigma E [7].
• Pol II-transcribed U3 RNA, containing the 5'-terminal cap different from that present in the wild-type counterpart, is packaged in transfected protoplasts into U3 snRNP precipitable with anti-fibrillarin antibodies [8].
• A synthetic intron of arbitrary sequence but incorporating splice site consensus sequences and a high proportion of U and A nucleotides, a characteristic feature of plant introns, was efficiently spliced in protoplasts [9].
• A complete human gamma 1 heavy chain gene (HIG1) was transferred into mouse cells by protoplast fusion [10].

Chemical compound and disease context of Protoplasts

• Tobacco protoplasts transformed with Agrobacterium tumefaciens carrying this vector yielded chlorsulfuron-resistant colonies [11].
• Latent ATPase, located on the inner surface of protoplast ghosts of Mycobacterium phlei, was unmasked either by trypsin or an impermeable form of trypsin, ethylene maleic anhydride-trypsin [12].
• A method is described for the preparation of protoplasts of Streptococcus mutans BHT [13].
• Transient expression assays in rice protoplasts indicate that a 134 bp fragment (-194 to -60 containing the ABRE) fused to a truncated cauliflower mosaic virus promoter (35S) is sufficient to confer ABA-responsiveness upon the GUS reporter gene [14].
• Mycelial protoplasts were transformed to geneticin-resistance with plasmids containing the Escherichia coli kanamycin-resistance gene as a selectable marker and autonomously replicating sequences (ARS) from Saccharomyces cerevisiae (ARS1, 2 mu ARS) [15].

Biological context of Protoplasts

• The genomic DNA, as well as a cognate complementary DNA, once transfected into protoplasts promote growth and cell division in vitro in the absence of exogenously added auxin [1].
• We show that VP1 overexpression and the hormone, abscisic acid (ABA), activate a reporter gene driven by the C1 promoter in maize protoplasts [16].
• Shortly after transfection, expression of this chimeric NPTII gene was negatively affected by high extracellular sucrose concentrations in the protoplast cultivation medium [17].
• A DNA fragment containing the conserved sequence acts as an auxin responsive element in transient expression assays in wild-type protoplasts and this response is higher in axi 159 protoplasts [18].
• An RNA3 mutant bearing a 20-base deletion yielding normal levels of aminoacylation and enhanced levels of replicase template activity and adenylation in vitro was able to replicate in protoplasts and plants; however, its accumulation in protoplasts was reduced relative to wild-type [19].

Anatomical context of Protoplasts

• Fusion of hen erythrocytes with yeast protoplasts induced by polyethylene glycol [20].
• Substrate specificity and inhibition by protein modifiers are similar to results obtained for sucrose transport into protoplasts and plasma membrane vesicles and for the spinach transporter, with the exception of a reduction in maltose affinity [21].
• The association of glycolytic enzymes with mitochondria was confirmed in vivo by the expression of enolase- and aldolase-yellow fluorescent protein fusions in Arabidopsis protoplasts [22].
• The technique involves digesting bacterial cell walls with lysozyme to produce protoplasts and then fusing the protoplasts to mammalian cells by treatment with polyethylene glycol [4].
• Subcellular fractions were obtained from protoplasts of a high penicillin-producing strain upon lysis by Triton X-100, and vacuoles purified from them [23].

Associations of Protoplasts with chemical compounds

• Tobacco protoplasts treated with cAMP, or the adenylyl cyclase activator forskolin, no longer require auxin to divide [24].
• Exposure to increased CO2 significantly reduced the cellular content of dimethylallyl diphosphate, the substrate for isoprene synthesis, in both leaves and leaf protoplasts [25].
• Dark-repair of ultraviolet-induced pyrimidine dimers in the DNA of wild carrot protoplasts [26].
• The ability of protoplasts to sustain growth in response to cytokinin glucosides persisted indefinitely after the likely disappearance of the expression vector [27].
• A high voltage electrical pulse was applied to the protoplasts, which were then grown on filters placed over feeder layers of maize suspension cells (Black Mexican Sweet) and selected for growth in the presence of kanamycin [28].

Gene context of Protoplasts

• Analyses of loss-of-function and ARR2-overexpressing lines as well as functional assays in protoplasts indicate an important role of ARR2 in mediating ethylene responses [29].
• Also, a T-DNA insertion mutation at the ADL1E gene caused abnormal mitochondrial elongation that was rescued by the transient expression of ADL1C and ADL1E in protoplasts [30].
• DREB2A domain analysis using Arabidopsis protoplasts identified a transcriptional activation domain between residues 254 and 335, and deletion of a region between residues 136 and 165 transforms DREB2A to a constitutive active form [31].
• Using transient expression in protoplasts of BRI1 and AtSERK3 fused to cyan and yellow fluorescent green fluorescent protein variants allowed us to localize each receptor independently in vivo [32].
• We find that ACD2 shields root protoplasts that lack chlorophyll from light- and PPIX-induced PCD [33].

Analytical, diagnostic and therapeutic context of Protoplasts

• Here we study a chimeric integral membrane reporter protein expressed in tobacco suspension culture protoplasts whose traffic was assessed biochemically by following acquisition of complex Asn-linked glycan modifications and proteolytic processing, and whose intracellular localization was determined with confocal immunofluorescence [34].
• By use of indirect immunofluorescence, anti-IFA stained formaldehyde-fixed onion meristematic cells and carrot protoplasts in patterns approximating those obtained with monoclonal anti-tubulins [35].
• In immunohistochemistry and in vivo targeting experiments in Arabidopsis protoplasts, ADL1C and ADL1E appeared as numerous speckles and the two proteins colocalized [30].
• The association of AC4 with miRNA was demonstrated by the association of A-AC4-GFP fusion protein, extracted from Arabidopsis protoplasts, with 2'-O-methyloligonucleotide complementary to miR159 (miR159*) and by the presence of miRNA with the A-AC4-GFP fusion protein after immunoprecipitation with antibody against GFP [36].
• In this paper, we show that T-strands disappear from acetosyringone-induced A. tumefaciens within 30 min of bacterial cocultivation with tobacco protoplasts [37].

References