The relationship between endosperm cap weakening and endo-β-mannanase activity during priming and the time to germination after priming was studied in tomato (Lycopersicon esculentum) seeds. During priming of seeds in −0.4 MPa PEG, the mechanical restraint of the endosperm cap decreased while the endo-β-mannanase activity in the endosperm cap increased. There was no decrease in required puncture force and no increase in endo-β-mannanase activity in seeds during priming in −1.0 MPa PEG. Two classes of seeds could be distinguished during priming in −0.7 MPa PEG: one with decreased required puncture force and one without. A strong correlation was found between the lowering of the mechanical restraint and endo-β-mannanase activity. It was concluded that individual seeds have to cross a threshold water potential in order to develop enzyme activity and lower their mechanical restraint. A decrease in required puncture force and increase in endo-β-mannanase activity correlated with ice crystal-induced porosity in the endosperm cap cell walls in scanning micrographs. It was presumed that ice crystal-induced porosity reflects cell-wall hydrolysis. Germination time after priming correlated positively with required puncture force during priming, depending on the osmotic potential. Seeds in −1.0 MPa PEG improved their time to germination, without a decrease in the required puncture force. Therefore it was concluded that lowering of the endosperm restraint during priming positively affects the time to germination of primed seeds but is not a prerequisite for rapid germination.

Imbibitional stress was imposed on neem (Azadirachta indica) seeds by letting them soak for 1 h in water at unfavourable, low temperatures before further incubation at 30°C. Sensitivity to low imbibition temperatures increased with a decrease in seed moisture content (MC). To investigate a possible involvement of seed age in the extent of imbibitional damage, initially high-quality seed lots that differed in storage history (10 weeks versus 10 months) were examined at 4 and 7% MC (fresh weight basis). After 10 months of storage, the 7% MC seeds had become sensitive to imbibitional stress. Further drying (1 week) to 4% MC affected aged seeds more than non-aged seeds. Barrier properties of cellular membranes in axes excised after 1 d of rehydration were estimated using a spin-probe technique. The proportion of cells with intact membranes increased with increasing imbibition temperature. For each temperature tested, there were more cells with leaky membranes after 10 months than after 10 weeks of dry storage. Localization of embryo cells displaying loss of turgor and abnormal cellular structure was accomplished using cryo-planing, followed by cryo-scanning electron microscopy. Inspection of the cryo-planed surfaces confirmed that imbibitional damage was temperature dependent, occurring at the periphery. Ageing increased the number of imbibitionally damaged, peripheral cell layers. Germination was estimated to fail when less than 70% of axis cells were alive. We conclude that ageing increases the sensitivity to imbibitional stress. Both the fast ageing and the sensitivity to imbibitional stress might explain the apparent controversies about neem seed desiccation tolerance and storage behaviour.

Cryo-high-resolution scanning electron microscopy was used to analyze the conformation of fibronectin fibrils formed in human skin fibroblast cultures or in a cell-free system by treating soluble plasma fibronectin with guanidine. Structurally, fibrils assembled in the cell-free system and in culture were similar. Assembly of both fibrillar networks involves interactions with the III1 and amino terminal repeats of fibronectin; their conformations consist of either smooth surfaces or patches of smooth surfaces and nodules randomly spaced along the fibril. The random distribution of these two conformations in fibrils indicates that fibronectin fibrils are capable of undergoing localized conformational changes. The nodules may be discrete domains of 3 to 4 type III repeats, as they can be labeled with the monoclonal antibody IST-2 to the III13-14 repeats in fibronectin and are found in 160 kDa and 85 kDa fragments of fibronectin that only contain type III repeats. In our study, smooth regions of fibrils were never recognized by the IST-2 antibody, suggesting that the epitope in the III13-14 repeats is masked in these regions. These results demonstrate that fibronectin fibrils are flexible and certain epitopes of fibronectin may be buried, or exposed, depending on the conformation of the fibril. They also show that fibrils assembled in cell-free conditions can be a powerful tool for studying fibril formation.