Survival of water stress in annual fish embryos: dehydration avoidance and egg envelope amyloid fibers

Abstract

Diapausing embryos of Austrofundulus limnaeus survive desiccating conditions by reducing evaporative water loss. Over 40% of diapause II embryos survive 113 days of exposure to 75.5% relative humidity. An early loss of water from the perivitelline space occurs during days 1–2, but thereafter, rates of water loss are reduced to near zero. No dehydration of the embryonic tissue is indicated based on microscopic observations and the retention of bulk (freezable) water in embryos as judged by differential scanning calorimetry. Such high resistance to desiccation is unprecedented among aquatic vertebrates. Infrared spectroscopy indicates frequent intermolecular contacts via β-sheet (14%) in hydrated egg envelopes (chorions). These β-sheet contacts increase to 36% on dehydration of the egg envelope. Interestingly, the egg envelope is composed of protein fibrils with characteristics of amyloid fibrils usually associated with human disease. These features include a high proportion of intermolecular β-sheet, positive staining and green birefringence with Congo red, and detection of long, unbranched fibrils with a diameter of 4–6 nm. The high resistance of diapause II embryos to water stress is not correlated with ontogenetic changes in the egg envelope.