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Cnr (colorless non-ripening) is a pleiotropic tomato (Lycopersicon esculentum) fruit ripening mutant with altered tissue properties including weaker cell-to-cell contacts in the pericarp (A.J. Thompson, M. Tor, C.S. Barry, J. Vrebalov, C. Orfila, M.C. Jarvis, J.J. Giovannoni, D. Grierson, G.B. Seymour [1999] Plant Physiol 120: 383-390). Whereas the genetic basis of the Cnr mutation is being identified by molecular analyses, here we report the identification of cell biological factors underlying the Cnr texture phenotype. In comparison with wild type, ripe-stage Cnr fruits have stronger, non-swollen cell walls (CW) throughout the pericarp and extensive intercellular space in the inner pericarp. Using electron energy loss spectroscopy imaging of calcium-binding capacity and anti-homogalacturonan (HG) antibody probes (PAM1 and JIM5) we demonstrate that maturation processes involving middle lamella HG are altered in Cnr fruit, resulting in the absence or a low level of HG-/calcium-based cell adhesion. We also demonstrate that the deposition of (135)-␣-l-arabinan is disrupted in Cnr pericarp CW and that this disruption occurs prior to fruit ripening. The relationship between the disruption of (135)-␣-larabinan deposition in pericarp CW and the Cnr phenotype is discussed.The modification of cell walls (CW) is an important aspect of plant cell development. During fruit ripening the regulated swelling and dissolution of primary CW and the modification of middle lamellae (ML) between adherent primary CW are important factors contributing to tissue softening (Brady, 1987; Fischer and Bennett, 1991). The biochemistry and the spatial regulation of the dissolution of primary CW and ML are not fully understood, but in all cases appear to involve modifications to the network of pectic polysaccharides.The multi-functional pectic polysaccharides are the most complex class of polysaccharides in primary plant CW (Jarvis, 1984). Core backbone structures of contiguous 1,4-linked ␣-d-galacturonic acid (homogalacturonan, HG) or repeats of the disaccharide [34)-␣-d-GalA-(132)-␣-l-Rha-(13] (rhamnogalacturonan, RG) are elaborated with a range of modifications and substitutions. These include methylesterification, acetylation, and the addition of neutral polysaccharide side chains. Side chains may be attached to HG and RG to form the branched polysaccharides RG-II and RG-I, respectively, the latter often rich in (135)-␣-l-arabinan and (134)-␤-d-galactan components (O'Neill et al., 1990; Albersheim et al., 1996; Mohnen 1999). Several of these pectic structures appear to be capable of being enzymatically modified in muro. For example, the de-esterification of HG by pectin methyl esterases (PMEs) influences its capacity to form calcium cross-linked gels. The relationships of pectic polysaccharide domains within large polymer structures and their functional properties in relation to factors such as porosity, cell adhesion and expansion, ionic and hydration status, and cell signaling, are far from clear.Ripening-related textural changes in t...

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Multicellular plants depend for their integrity on effective adhesion between their component cells. This adhesion depends upon various cross-links; ionic, covalent or weak interactions between the macromolecules of the adjacent cell walls. In sugar-beet (Beta vulgaris L. Aztec) root parenchyma, cell-cell adhesion is disrupted by successive extractions with a calcium-chelating agent (imidazole) and a de-esterifying agent (sodium carbonate) but not by the calciumchelating agent or the de-esterifying agent alone. Cell-cell adhesion in sugarbeet parenchyma thus depends upon both ester and Ca 2þ cross-linked polymers. Pectic polysaccharides are removed by these treatments. Both parallel-electron energy-loss spectroscopy (PEELS) and Image-EELS show that calcium-binding sites are removed from the wall by imidazole. Using a monoclonal antibody that recognizes a relatively unesterified epitope of homogalacturonan, JIM 5, we show that a subset of JIM 5-reactive antigens remain in the middle lamella after Ca 2þ chelation and that this subset is removed by cold (4 C) Na 2 CO 3 -induced breakage of ester bonds. Fourier transform infrared, nuclear magnetic resonance, and spectrophotometric assays show that methyl and feruloyl esters are removed from the wall by Na 2 CO 3 but acetyl esters remain. Sodium carbonate extraction at 20 C removes cell wall autofluorescence and most of the feruloylated moieties from the wall. We propose that the chelator-resistant subset of ester-linked JIM 5-reactive pectins are important for cell-cell adhesion.

The targeted radiotherapy of neuroblastoma with 131l-labelled metaiodobenzyl guanidine (mIBG) is now the subject of several clinical studies. The precise intracellular localization of mIBG, necessary for nuclear microdosimetry, has not previously been described. We report the use of electron-energy-loss spectroscopy and electron spectroscopic imaging to establish the intracellular distribution of mIBG in cells from the human neuroblastoma line NBI-G which had been incubated with the drug by mapping iodine in ultra-thin sections of tumours. Most of the iodine is found within the mitochondria, with lesser amounts in the vesicles and on the nuclear membrane. The use of alternative radionuclides with different physical characteristics has been suggested to optimize the efficacy of this therapeutic strategy. The lack of penetration of mIBG into the nucleoplasm means that ultra-short-range Auger electron emitters such as 125I are not likely to prove more cytotoxic than 131I.