Development of transgenic

Elysia chlorotica as an
Effective Living
Bioindicator for Detecting
Environmental Toxicants
JONATHAN LING

Introduction
Many threats are endangering aquatic
environments
The discharge of industrial and domestic
wastewater has the most significant impacts
on freshwater ecosystems, as well as
agricultural production and human health
Constant monitoring is essential to ensure
timely response whenever damaging waste
discharge events happen

Nature is full of thieves:

Elysia chlorotica
Novel organism that
demonstrates high
feasibility for horizontal
gene transfer
Observed phenomenon
of kleptoplasty
Biological burglars
Natural genetic
engineering Example
of a natural transgenic
process in the so-called
higher life forms

Justifications of model
Ability to maintain the chloroplasts acquired from
Vaucheria litorea in a functional state
demonstrates:
Elysia chlorotica must possess photosynthesissupporting genes within its own nuclear genome,
most likely acquired through horizontal gene
transfer
Elysia chlorotica has incorporated the required
algal DNA into its own DNA so the algal plastids
(now Sea Slug plastids) can receive the proteins
they need to function according to specification
Vital algal gene, psbO, found in the sea slug's
DNA. Found to be identical to the algal version

Methods
Introduction of GFP transgene regulated by the
upstream open reading frame (uORF) fragment of
human DNA-damage-inducible transcript 3 (ddit3,
previously named chop) cDNA
uORF-based translational regulation plays
significant, if not primary, roles in the production
of CHOP protein, while the chop gene is one of
the most commonly used biomarkers for
endoplasmic reticulum (ER) stress
Gene can be introduced into organisms and
maintained in their genome through breeding,
injection with a viral vector, or cell transformation

CHOP gene
The CHOP gene is induced by cellular stress and is
involved in mediating apoptosis
Implicated in adipogenesis and erythropoiesis, is
activated by endoplasmic reticulum stress, and
promotes apoptosis
Conserved region in the promoter of the CHOP gene
responsible for its inducibility by endoplasmic
reticulum (ER) stress
The unfolded protein response (UPR) is a cellular
stress response related to the endoplasmic reticulum.
It is a stress response that has been found to be
conserved between all mammalian species, as well
as yeast and worm organisms

GFP Transgene
GFP gene is frequently used as a reporter of
expression
GFP gene has been introduced and expressed
in many Bacteria, Yeast and other Fungi, fish,
plants, flies, and mammalian cells
Different stresses could cause different GFP
expression patterns in a dose-dependent
manner
Holds considerable promise as a novel
fluorescent biomonitoring method

Methods
Embryos will be used for preliminary in vitro
research
Embryos derived from the transgenic Elysia
chloroticaonly to display fluorescent signals upon
encountering stresses, with no detectable
leakage under normal conditions
Embryos can give a faithful account of cellular
stresses
Could potentially be used to detect various
environmental contaminants, including heavy
metals and endocrine-disrupting chemicals
(EDCs)

Why transgenic?
When
Normal conditions, such as growth, survival
rates and egg hatchability, can be used as
monitoring parameters.
Quantifying the activity of enzymatic
defenses in is a common approach to assess
water quality
Interpretation of the data obtained from
these methods is limited by the fact
that multiple physiological, genetic, and
metabolic factors may simultaneously
affect these multifunctional enzymes

Why transgenic?
Why not native gene promoters?
the cyp1a1 promoter, which is induced
by polycyclic aromatic hydrocarbons
the heat-shock promoter, which is
induced by heat and other stressors

Why transgenic?
These promoters only respond to specific forms
of stress, their advantages over traditional
chemical analysis are not particularly significant
A given stress with little harm to the animal
may still induce the expression of a reporter
gene controlled by the heat-shock promoter
The reporter activity would have little
relationship to the actual physiological stresses
Hence, for an animal model to be a practical
biomonitor, it must (1) respond to a wide range
of pollutants with accuracy and sensitivity and
(2) dynamically trace physiological stresses

Limitations
We cannot discriminate natural variability
from changes due to human impacts, thus
limiting the applicability of bioindicators in
heterogeneous environments
Ethical issues
Issue of releasing a transgene into the wild if
the slugs are to be used as living
bioindicators