CINCINNATI -- What molecular biologists used to call "heatshock proteins" (HSP) are set to invade environmental marketsunder their new, all-encompassing name, "cellular stressresponse proteins."

So said Richard M. Glickman, founder and president ofStressGen Biotechnologies Corp. of Victoria, British Columbia.

His two-and-one-half-year-old company co-sponsored a stressprotein symposium at the Society of Environmental Toxicologyand Chemistry's (SETAC) annual meeting here last week. Threebiotechnology-oriented strategies for early-warning systems todetect and measure hazardous contaminants attracted attentionin the symposium:

-- Eve Stringham, a biochemistry post-doc at the University ofBritish Columbia, Vancouver, reported to the SETAC symposiumon "Use of transgenic strains of the nematode Caenorhabditiselegans as monitors for environmental stress." Stringham,supported by a StressGen grant, is educating the tiny nematoderoundworm to do at the molecular level what canaries used todo in coal mines -- sense dangerous levels of contaminants.

Instead of methane, which canaries detect, Stringham'stransgenic C. elegans worms sense and measure heavy-metalcontaminants and such harmful herbicides as paraquat.

How do they do it? Stringham transforms their eggs withreporter genes containing a stress-protein promoter gene fusedto the coding region of the E. coli enzyme, b-galactosidase (b-gal) gene. When the adult transgenic animals are stressed -- asby a heavy metal contaminant -- they switch on the b-galenzymes along with their own stress proteins.

She then adds one of two colorless substrates for the b-gal geneto cleave. The first releases a soluble yellow product, whichmeasures the magnitude of the response (the concentration ofcontaminant, for example). The second produces an insolubleblue precipitate, which localizes it in the worm's tissues.

To test this twofold detection system, Stringham exposedtransgenic nematodes to water containing salts of arsenic,cadmium copper, lead, mercury or zinc, as well as paraquat.

Varying percentages of the pointer worms reacted to thesetoxic stresses by releasing blue and yellow markers intosusceptible tissues -- the gut for mercury and paraquat,pharynx for lead, copper and cadmium, hypodermic cells forzinc. Arsenic was "fairly evenly distributed throughout thetissues of the animals in a pattern reminiscent of classic heatshock induction."

This color-coded assay, Stringham told her audience, "wouldappear to be a more sensitive indicator of heavy metalcontamination than a standard LC50 (lethal concentration) testusing C. elegans.

-- At Georgia Institute of Technology in Atlanta biologist TerrySnell feeds fluorescently labeled latex microspheres to rotifers.These, like the soil-dwelling nematodes of which rotifers areaquatic cousins, are minute, multicellular and transparent.Taking rates of ingestion as a measure of stress response, Snellreported that fluorescence of rotifers exposed to water pollutedwith only 20 parts per billion of copper dropped from 160units to almost zero.

A current commercial test of such water pollution, he said, usesa similar organism, Cladocera, in which it tracks death andreproduction rates, rather than ingestion. A single suchstandard assay, he stated, costs about $1,000, and results takefour to seven days. Using lit-up rotifers, and ingestion ratherthan mortality as a gauge, he can do it for "50 times lessbecause less technician time is needed." A mere 15-minuteexposure is at least as good as the slower, costlier commercialtest, he added.

Snell also said that he plans to use firefly luciferase genes toproduce transgenic rotifers, fish and other animals, whichwould light up when unstressed and dim down when stressed.

-- Cell culture is the route chosen by molecular and cellbiologist Lawrence E. Hightower of the University ofConnecticut at Storrs. His marker source is a guppy-like fish,the Mexican desert topminnow, which has strong stressresponses. From this fish he has raised a hepatic carcinoma cellline, which has receptors to such toxins asdimethylbenz[a]anthracene (DMBA), a strong liver carcinogen.Instead of sacrificing fish to measure toxicity in their tissues,his team exposes the cell line in vitro to DMBA and recordsinduction of heat shock proteins.

"From my perspective," Hightower told BioWorld, "I seebiotechnology creeping into environmental biology. Mostprogress will occur from multidisciplinary synergy amongacademia, industry and government."

His work is sponsored by the U.S. Geologic Survey and the LongIsland Sound Fund of Connecticut's Department ofEnvironmental Protection.

-- David N. Leff Science Editor

(c) 1997 American Health Consultants. All rights reserved.

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