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Acidification of intracellular compartments by the vacuolar-type H+-ATPases (VHA) is known to energize ion and metabolite transport, though cellular processes influenced by this activity are poorly understood. At least 26 VHA genes encode 12 subunits of the V1Vo-ATPase complex in Arabidopsis, and how the expression, assembly, and activity of the pump are integrated into signaling networks that govern growth and adaptation are largely unknown. The role of multiple VHA-c genes encoding the 16-kD subunit of the membrane Vo sector was investigated. Expression of VHA-c1, monitored by promoter-driven β-glucuronidase (GUS) activity was responsive to light or dark in an organ-specific manner. VHA-c1 expression in expanding cotyledons, hypocotyls of etiolated seedlings, and elongation zone of roots supported a role for V-ATPase in cell enlargement. Mutants reduced in VHA-c1 transcript using dsRNA-mediated interference showed reduction in root growth relative to wildtype seedlings. In contrast, VHA-c3 promoter::GUS expression was undetectable in most organs of seedlings, but strong in the root cap. Interestingly, dsRNA-mediated mutants of vha-c3 also showed reduced root length and decreased tolerance to moderate salt stress. The results suggest that V-ATPase functions in the root cap influenced root growth. Expression of VHA-c1 and VHA-c3 in tissues with active membrane flow, including root cap, vascular strands, and floral style would support a model for participation of the Vo sector and V1Vo-ATPase in membrane trafficking and fusion. Two VHA-c genes are thus differentially expressed to support growth in expanding cells and to supply increased demand for V-ATPase in cells with active exocytosis. [ABSTRACT FROM AUTHOR]

Presents the sequence of chromosome 2 of the plant Arabidopsis thaliana. Analysis of the chromosome in two gap-free assemblies of 3.6 and 16 megabases; Features of chromosome 2; Gene and chromosomal duplications; Comparative and evolutionary genomics; Mitochondrial genome in the nucleus; Pericentromeric region; Representation of transposable elements on chromosome 2.

It has recently been established that synthesis of double-stranded cDNA can be done from a single cell for use in DNA sequencing. Global gene expression can be quantified from the number of reads mapping to each gene, and mutations and mRNA splicing variants determined from the sequence reads. Here we demonstrate that this method of transcriptomic analysis can be done using the extremely low levels of mRNA in a single nucleus, isolated from a mouse neural progenitor cell line and from dissected hippocampal tissue. This method is characterized by excellent coverage and technical reproducibility. On average, more than 16,000 of the 24,057 mouse protein-coding genes were detected from single nuclei, and the amount of gene-expression variation was similar when measured between single nuclei and single cells. Several major advantages of the method exist: first, nuclei, compared with whole cells, have the advantage of being easily isolated from complex tissues and organs, such as those in the CNS. Second, the method can be widely applied to eukaryotic species, including those of different kingdoms. The method also provides insight into regulatory mechanisms specific to the nucleus. Finally, the method enables dissection of regulatory events at the single-cell level; pooling of 10 nuclei or 10 cells obscures some of the variability measured in transcript levels, implying that single nuclei and cells will be extremely useful in revealing the physiological state and interconnectedness of gene regulation in a manner that avoids the masking inherent to conventional transcriptomics using bulk cells or tissues. [ABSTRACT FROM AUTHOR]

Epidermal growth factor receptor tyrosine kinase inhibitors gefitinib and erlotinib have been widely used in patients with non-small-cell lung cancer. Unfortunately, the efficacy of EGFR-TKIs is limited because of natural and acquired resistance. As a novel cytoprotective mechanism for tumor cell to survive under unfavorable conditions, autophagy has been proposed to play a role in drug resistance of tumor cells. Whether autophagy can be activated by gefitinib or erlotinib and thereby impair the sensitivity of targeted therapy to lung cancer cells remains unknown. Here, we first report that gefitinib or erlotinib can induce a high level of autophagy, which was accompanied by the inhibition of the PI3K/Akt/mTOR signaling pathway. Moreover, cytotoxicity induced by gefitinib or erlotinib was greatly enhanced after autophagy inhibition by the pharmacological inhibitor chloroquine (CQ) and siRNAs targeting ATG5 and ATG7, the most important components for the formation of autophagosome. Interestingly, EGFR-TKIs can still induce cell autophagy even after EGFR expression was reduced by EGFR specific siRNAs. In conclusion, we found that autophagy can be activated by EGFR-TKIs in lung cancer cells and inhibition of autophagy augmented the growth inhibitory effect of EGFR-TKIs. Autophagy inhibition thus represents a promising approach to improve the efficacy of EGFR-TKIs in the treatment of patients with advanced non-small-cell lung cancer. [ABSTRACT FROM AUTHOR]