Renewal of nongermative epithelia is poorly understood. The novel mitogen “lacritin” is apically secreted by several nongermative epithelia. We tested 17 different cell types and discovered that lacritin is preferentially mitogenic or prosecretory for those types that normally contact lacritin during its glandular outward flow. Mitogenesis is dependent on lacritin's C-terminal domain, which can form an α-helix with a hydrophobic face, as per VEGF's and PTHLP's respective dimerization or receptor-binding domain. Lacritin targets downstream NFATC1 and mTOR. The use of inhibitors or siRNA suggests that lacritin mitogenic signaling involves Gαi or Gαo–PKCα-PLC–Ca2+–calcineurin–NFATC1 and Gαi or Gαo–PKCα-PLC–phospholipase D (PLD)–mTOR in a bell-shaped, dose-dependent manner requiring the Ca2+ sensor STIM1, but not TRPC1. This pathway suggests the placement of transiently dephosphorylated and perinuclear Golgi–translocated PKCα upstream of both Ca2+ mobilization and PLD activation in a complex with PLCγ2. Outward flow of lacritin from secretory cells through ducts may generate a proliferative/secretory field as a different unit of cellular renewal in nongermative epithelia where luminal structures predominate.
We screened for mutations deleterious to the freezing tolerance of Arabidopsis fhaliana (1.) Heynh. ecotype Columbia. lolerance was assayed by the vigor and regrowth of intact plants after cold acclimation and freezing. From a chemically mutagenized population, we obtained 1 3 lines of mutants with highly penetrant phenotypes. In 5 of these, freezing sensitivity was attributable to chilling injury sustained during cold acclimation, but in the remaining 8 lines, the absence of injury prior to freezing suggested that they were affected specifically in the development of freezing tolerance. In backcrosses, freezing sensitivity from each line segregated as a single nuclear mutation. Complementation tests indicated that the 8 lines contained mutations in 7 different genes. l h e mutants' freezing sensitivity was also detectable in the leakage of electrolytes from frozen leaves. However, 1 mutant line that displayed a strong phenotype at the whole-plant leve1 showed a relatively weak phenotype by the electrolyte leakage assay.
Transposon Tn7 transposes at high frequency to a specific site, attTn7, in the Escherichia coli chromosome. We devised a quantitative assay for Tn7 transposition in which Tn7-end derivatives containing the cis-acting transposition sequences of Tn7 transpose from a bacteriophage lambda vector upon infection into cells containing the Tn7-encoded transposition proteins. We used this assay to identify a 68-base-pair DNA segment containing the sequences essential for atlTn7 target activity. This segment is positioned asymmetrically with respect to the specific point of Tn7 insertion in atLln7 and lacks obvious homolog to the sequences at the ends of Tn7 which participate directly in transposition. We also show that some sequences essential for attTn7 target activity are contained within the protein-coding sequence of a bacterial gene.
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