Isolation of an Algal Morphogenesis Inducer from a Marine Bacterium

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Science  11 Mar 2005:
Vol. 307, Issue 5715, pp. 1598
DOI: 10.1126/science.1105486


Ulva and Enteromorpha are cosmopolitan and familiar marine algal genera. It is well known that these green macroalgae lose their natural morphology during short-term cultivation under aseptic conditions and during long-term cultivation in nutrient-added seawater and adopt an unusual form instead. These phenomena led to the belief that undefined morphogenetic factors that were indispensable to the foliaceous morphology of macroalgae exist throughout the oceans. We characterize a causative factor, named thallusin, isolated from an epiphytic marine bacterium. Thallusin induces normal germination and morphogenesis of green macroalgae.

Ulva (sea lettuce) and Enteromorpha (gut weed) are cosmopolitan, intertidal marine algae that display a foliaceous morphology in their natural environment. However, these green macroalgae lose their natural morphology during short-term cultivation under aseptic conditions and during long-term cultivation in nutrient-added seawater (17). This observation has led to the belief that undefined morphogenetic factors are present in the ocean that are indispensable for the foliaceous morphology of macroalgae (2).

The alga Monostroma oxyspermum (Monostromataceae, Ulvales) has a leafy morphology in its natural state and looks like thin Ulva. However, under aseptic conditions, it grows as loose aggregates of single cells (7). We have shown previously that these cells can redifferentiate upon reinfection with marine bacteria from the Cytophaga-Flavobacterium-Bacteroides (CFB) group that were isolated from the green algal surface (8). We have also demonstrated that bacterial extracellular metabolites can induce not only the normal morphogenesis of M. oxyspermum but also the normal germination of germfree spores of Ulva pertusa and Enteromorpha intestinalis (8).

We report here the isolation of a highly potent differentiation inducer, termed thallusin (Fig. 1A), from the epiphytic marine bacterium strain YM2-23 (deposition no. MBIC 04683), belonging to the CFB group that was isolated from Monostroma sp. (8). The structure of thallusin was determined by spectral and single-crystal x-ray structure determination of a thallusin derivative, Me1H1W4 (fig. S1) (9).

Fig. 1.

(A) The structure of thallusin. Only the relative configuration was determined. (B) Morphology of axenic M. oxyspermum, after 7 days of cultivation. Inset: A loose culture mass consisting of single cells. (C) The effect of thallusin on M. oxyspermum (an initial concentration of 1 ng ml–1). Inset: A typical monostromatic gametophyte. (D) Dedifferentiation given a lack of thallusin (an initial concentration of 1 pg ml–1). Inset: Protruding cells from disintegrating thalli. (E) Small callus-like morphology of U. pertusa, after 2 months of cultivation under aseptic conditions. Inset: Colorless protrusions from the lateral cell walls. (F) The effect of thallusin (1 ng ml–1) on U. pertusa. Inset: A foliaceous distromatic blade. Scale bars, (B) to (D), 100 μm; (E) and (F), 4 mm; insets, 20 μm.

Pure thallusin strongly induced the differentiation of M. oxyspermum (Fig. 1, B and C). The minimum effective concentration (8) in a cell differentiation assay with M. oxyspermum was between 1 fg ml–1 and 1 ag ml–1, depending on the lighting conditions, the assay duration (usually 4 days), and the initial number of M. oxyspermum cells (usually 20 cells per ml). When thallusin was added at a concentration of 100 μg l–1, no inhibitory activity was apparent in the cell differentiation or growth of M. oxyspermum. The morphogenesis of M. oxyspermum into the thallus with thallusin addition and the gradual disintegration of the thallus into single cells with thallusin deprivation (Fig. 1D) suggest that thallusin must be continuously supplied by an epipythic marine bacterium to maintain algal body.

Thallusin was also able to induce the germination of such other green macroalgae as U. pertusa (Fig. 1, E and F) and E. intestinalis (10), after they had been cultured under aseptic conditions. Previous studies have also reported that these green algae did not develop as normal foliate gametophytes under aseptic conditions (15). Because thallusin was effective toward U. pertusa and E. intestinalis, in addition to M. oxyspermum, thallusin-dependent morphogenesis may be a common feature in Monostromataceae and Ulvaceae (8).

The mode of action of the algal response to thallusin is unknown, but a bacterial supply of thallusin functions in the development of algal communities. Other differentiation inducers may be found in nature. The role of thallusin shown here exemplifies the fundamental symbiotic chemical communication between macroalgae and epiphytic bacteria in the marine environment.

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Fig. S1

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