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EnvZ: Transitioning Between Virulence and Symbiosis in Xenorhabdus nematophila
Wauwatosa West SMART Team: Waj Ali, Jimmy Kralj, Jordan Llanas, Leah Rogers, Mariah Rogers
Advisor: Mary Anne Haasch Mentor: Steven Forst, Ph.D., University of Wisconsin - Milwaukee
To Phosphorylate or Not To Phosphorylate
Abstract
The bacterium Xenorhabdus nematophilia participates in an unusual and
fascinating mutualistic relationship with the nematode, Steinernema carpocapsae,
which could not complete its lifecycle without the bacteria’s help. EnvZ, a kinase
protein located in the cell membrane of the bacterium, is critical to both
organisms’ success. Xenorhabdus resides quietly in a specialized pouch in
Steinernema’s intestines. To reproduce the juvenile nematode enters an insect
host via the anus or the mouth, and bores a hole though the intestine wall to get
into the insect’s hemolymph. In response to the environmental signals in the
hemolymph, the nematode’s pharynx begins to pump, forcing the Xenorhabdus out
of the intestinal pouch and into the insect’s hemolymph. Xenorhabdus begins to
act as an insect pathogen— killing the host insect, while simultaneously secreting
antibiotics to eliminate its bacterial competitors. In killing the insect host
Xenorhabdus provides its symbiotic partner, Steinernema, with a carcass in which
to reproduce. EnvZ helps Xenorhabdus sense the higher solute concentrations in
the insect’s hemolymph through a currently unknown mechanism. EnvZ functions
as a dimer. In response to appropriate environmental signals, a phosphate from
ATP bound to the cytoplasm section of one EnvZ molecule is transferred to the
HIS243 of another EnvZ. The phosphate group on HIS243 is then transferred to
OmpR, a gene-regulating protein in Xenorhabdus that regulates genes that produce
the antibiotics used to kill bacterial competitors. OmpR also regulates genes for
exoenzymes that degrade insect tissues providing nutrients that help the
nematode to reproduce.
EnvZ
P
HIS
1
ADP
P
Phos
DNA
binding
http://www.uic.edu/labs/kenneyl/img/ompR.JPG
(with EnvZ)
Antibiotics are inhibited.
• At this early stage
Xenorhabdus has an
abundance of nutrients
and few competitors.
•Energy is conserved.
OmpR
Wild type
To the DNA
Hemolymph
Antibiotics are produced.
•These wipe out other
bacteria which compete for
nutrients.
Flagella are produced.
•With less nutrients,
flagella provide transport
to insect’s un-eaten areas.
1joy.pdb
This is HIS243, the
amino acid in EnvZ
which transfers the
phosphate to OmpR.
Life Cycle
A nematode
enters the insect
and goes into the
hemolymph
5
After the two generations,
the nematodes reuptake
the Xenorhabdus bacteria
and break out of the
insect’s carcass to restart
the cycle.
EnvZ-
If OmpR is not
Phosphorylated:
Exotoxins are produced.
•These break down insect
tissues, releasing more
nutrients for Xenorhabdus.
The nematodes go through about
two generations, eating the dead
insect, while the Xenorhabdus use
EnvZ to re-switch their genes to
produce the antibiotics necessary to
kill bacterial competitors as well as
the exoenzymes to break down the
nutrients in the dead insect.
(No EnvZ)
Exotoxins are inhibited.
•These break down insect
tissues to release nutrients
which are not needed at
this time.
Inside EnvZ
In the hemolymph, the bacteria
initially find a nutrient rich
environment. Here, the
bacteria rapidly divide, bind to
the tissue (and therefore no
longer need their flagella), and
focus on nutrient uptake.
4
EnvZ-
The EnvZ- strain produces more
antibiotics early on, as shown by the
larger zone of inhibition.
Flagella is inhibited.
•Movement is not needed
to find more nutrients.
EnvZ in membrane of
Xenorhabdus nematophilia
2
Wild type
If OmpR is Phosphorylated:
OmpR
DNA
3
The nematode’s pharynx
contains a specialized pouch
which holds Xenorhabdus
nematophila bacteria. The
nematode contracts its
pharynx and releases the
bacteria into the hemolymph.
ATP
Experiments
Uses for Xenorhabdus nematophila
and Steinernema carpocapsae
The nematodes, Steinernema carpocapsae,
are used as biological pesticides against
insect pests.
First row of pictures show flagella
presence at 6 hours since Xenorhabdus
nematophilia’s presence in the insect.
Second and third rows correspond to
8.5 hours and 11 hours respectfully.
Credit: Drs. Dongjin Park and Steven Forst .
The EnvZ- strain does not fully
inhibit the production of flagella at
6 hours. Flagella should be inhibited
as they are not needed yet.
Wild type inhibits flagella at 6
hours. At 8.5 hours flagella grow as
OmpR is not phosphorylated by
EnvZ and movement is needed.
Top Right: S. carpocapsae
escaping
an insect carcass
Lower Left: X. nematophilia
Lower Right: a harmful
caterpillar destroying foliage.
http//www.ukorganic.co.ukr
Xenorhabdus nematophilia could possibly
be used for its production of antibiotics and
other useful natural products, but so far no
such use has been commercialized.
Life Cycle images #2,4,5: Dr. Steven Forst , UW—Milwaukee and Dr. Donald Danforth Plant Science Center
Life Cycle image #3 bacteria with flagella: http://www.3Dscience.com
Supported by the National Institutes of Health (NIH) – National Center for Research Resources Science Education Partnership Award (NCRR-SEPA)
www.ARBICO-organics.com