Retrotransposon-like VL30 elements are efficiently induced in anoxic rat fibroblasts.

VL30 elements are a multigene family within the class of retroviruses and retrotransposons. We have characterized the response of normal rat fibroblasts to anoxia, in which endogenous VL30 element expression is strongly induced. Optimal induction up to 500-fold occurs under complete anoxia, although a lesser response is seen under atmospheres up to 2% oxygen. Phorbol esters and diacylglycerol, which induce mouse VL30 RNA approximately eightfold, show no effect on the rat VL30 system. The hypoxic conditions optimal for rat VL30 induction represent a mild cellular stress, with no reduction in cell viability during the induction period. Although the precise physiological role of this fibroblast response to temporary anoxia is unknown, it may occur during wound healing. The induction of VL30 by anoxia provides a unique model system wherein a member of the mammalian retrovirus/retrotransposon family is highly responsive to a common physiological signal.

Kinetochore microtubules in crane-fly spermatocytes: untreated, 2 degrees C-treated, and 6 degrees C-grown spindles.

We investigated the involvement of kinetochore microtubules (kMTs) in mediating chromosome-to-pole connections in crane-fly (Nephrotoma suturalis and Nephrotoma ferruginea) spermatocytes. Two experimental treatments were used to yield spindles with reduced numbers of nonkinetochore microtubules (nkMTs). Short-term (10-15 min) exposure of spermatocytes to 2 degrees C caused depolymerization of the majority of nkMTs, resulting in a kMT:(kMT + nkMT) ratio of 0.76. Long-term (24h) exposure to 2 degrees C followed by recovery at 6 degrees C resulted in a kMT:(kMT + nkMT) ratio of 0.55, the spindle having more nkMTs than a 2 degrees C-treated spindle but fewer than an untreated spindle, in which the kMT:(kMT + nkMT) ratio was 0.27. The numbers and lengths of kMTs in 6 degrees C-grown spindles were similar to those in untreated cells, suggesting that the overall inhibition of MT assembly at 6 degrees C apparently did not affect the mechanism by which kMTs are formed. We observed most kMTs of early anaphase spindles to be long (greater than 3 microns), and many extended to the polar regions of the spindle. Thus, the crane-fly spindle appears not to be as atypical as it was previously suggested to be.

Normal fibroblasts responding to anoxia exhibit features of the malignant phenotype.

Cancer has two fundamental features: neoplasia, representing the aberrant expression of a normal cell proliferation response, and malignancy, an ability to penetrate normal tissue boundaries. Although the role of oncogenes in neoplastic transformation is becoming clearer, malignancy remains far less well understood. Normal rat fibroblasts exhibit a staged response to anoxia which, if expressed in an uncontrolled fashion, may contribute vital aspects to the malignant phenotype. The response begins with induction of retrotransposon-like VL30 element transcription, progresses through induction of several intracellular proteins, and is followed by secretion of three major proteins including the protease cathepsin L. The induced VL30 element RNA encodes a 61-kDa secretory protein of unknown function. The response of fibroblasts to anoxia is evidently not a survival response. Instead, the response represents a close match to the role of fibroblasts during the early stages of wound healing where they are active under near-anoxic conditions. Malignant cancer cells are known to exhibit several of the characteristics we find induced in fibroblasts by anoxia. Conversion to the malignant phenotype may represent coordinate loss of control of this normal cellular response.