The
evolution of programmed cell death in marine phytoplankton
Co-PI: Paul Falkowski (IMCS,
What factors influence the availability and relative
coupling of phytoplankton organic matter to ecosystem
pathways like the microbial loop? One
is clearly mortality. Biological
oceanographers and phytoplankton physiologists have made an intense effort over
the past few decades to elucidate the conditions, mechanisms and strategies
that control phytoplankton cell growth.
Unfortunately, very few studies have specifically focused on
phytoplankton mortality, even though it largely determines how other organisms
live and is essential to linking major biogeochemical
cycles in aquatic ecosystems.
Consequently, the mechanisms controlling the dramatic and abrupt bloom
termination in natural systems are not well understood.
This lack
of attention is in part due to the misconception among biological
oceanographers that phytoplankton are largely
immortal, unless eaten by zooplankton grazers. Phytoplankton
are not immortal. Like all marine organisms, microscopic
phytoplankton encounter adverse environmental conditions, which are not only
detrimental to growth but often result in death. Indeed, substantial cell death (via lysis) has been documented in
field studies with some estimates exceeding 50% of phytoplankton growth. This includes viral infection and autocatalyzed cell death triggered
by nutrient stress, a self-destruction analogous to programmed cell death (PCD) or apoptosis in multicellular organisms.
I am examining its establishment and retention in
evolutionarily and biogeochemically diverse lineages of phytoplankton as a
critical mortality mechanism in response to environmental stress. This work provides insight into the
mechanistic control of phytoplankton mortality and provides novel ecological
and evolutionary context for PCD as a potentially
important loss component in natural systems.
Our lab has discovered genetically-controlled,
autocatalytic cell death programs in a prokaryotic (Trichodesmium sp. IMS101,
cyanobacterium) and in representatives of "green" (Dunaliella tertiolecta, chlorophyte) and "red' (Emiliania huxleyi, coccolithophorid) eukaryotic phytoplankton lineages.
This phenomenon is
elicited in response to physiological and environmental stress (culture
age, oxidative stress, iron and phosphorus limitation)
and results in massive cell death. The characteristics of
autocatalytic cell death in these diverse phytoplankton display remarkable
biochemical similarities to metazoans, including the involvement of
caspases. It has been
suggested that caspase-like cysteine proteases may represent the
initial, ancestral core of executioners that allowed the emergence of the cell
death machinery. I am currently examining
whether caspase-like proteases have a direct role in executing autocatalyzed cell death in prokaryotic and eukaryotic
phytoplankton, organisms that evolved well before metazoans. So far, I have identified several protein
sequences in a variety of phytoplankton that contain a caspase domain structure
and displayed sequence similarities to metacaspases
in the unicellular protists Saccharomyces cerevisiae (Yca-1) and Trypanosoma brucei, including the histidine- and cysteine-containing catalytic diad found in true caspases (p20 subunit), paracaspases and metacaspases.
Cellular and biochemical characteristics of
autocatalytic cell death in the filamentous, prokaryotic phytoplankton, Trichodesmium tertiolecta. (A) Growth and demise of
Trichodesmium IMS101 cultures.
(B) TEM micrographs showing changes in morphology of aging cells, with
arrows indicating the phase from which cells were sampled.
Abbreviations: th, thylakoids; gv, gas vesicles. (C)
TUNEL staining of healthy cells (left) and
physiological stressed cells, with the latter display intense, localized
incorporation of the TUNEL fluorescent label
indicative of DNA fragmentation and a large population of 3-hydroxyl ends. (D) Western blot analysis of whole cell
protein extracts illustrates increased in caspase-3 immunoreactivity
in aging Trichodesmium sp. IMS101
cells. Lane numbers identify age of
culture in days. (E) Dependence of DEVD
cleavage on mortality rate in two separate experiments. Filled diamonds represent the samples
presented in Western analysis. Images
are adapted from Berman-Frank et al. (2004).
Infection of E. huxleyi with EhV1 results in massive
mortality and nearly complete clearing of the culture (left image). Right image shows a scanning electron
micrograph of viral attachment.
Although the rationale for the
development of PCD in metazoans is clear, the reasons
for its development in unicellular phytoplankton remain elusive. Initially, the
thought of a unicellular organism opting to kill itself seems
counter-productive. However, if PCD could incur some
benefit to the population as a whole, it would become an evolutionary
advantage. One possible hypothesis is that PCD may
have evolved as a “virus exclusion” defense mechanism to limit the
spread of viruses during infection and to prevent catastrophic death of a
population. This type of response has been demonstrated
in the bacterium, Escherichia coli,
and humans, in the latter case involving the activation of caspases.
Furthermore, given that viral particles in the sea
exceed phytoplankton and bacterial abundance by an order of magnitude (or
more), this strategy would be extremely beneficial to clonal
phytoplankton populations. I
am currently examining if the PCD cellular machinery
plays a role in a viral-induced death cascade, using E. huxleyi and its virus (EhV1) as a model system.
Relevant publications:
BIDLE, K. D., and P. G. FALKOWSKI. 2004. Cell death in planktonic
photosynthetic microorganisms. Nature
Rev. Microbiol. 2: 643-655.
BERMAN-FRANK,