Colony Organization in the Green Alga Botryococcus braunii (Race B) Is Specified by a Complex Extracellular Matrix

Abstract

f Botryococcus braunii is a colonial green alga whose cells associate via a complex extracellular matrix (ECM) and produce prodigious amounts of liquid hydrocarbons that can be readily converted into conventional combustion engine fuels. We used quickfreeze deep-etch electron microscopy and biochemical/histochemical analysis to elucidate many new features of B. braunii cell/ colony organization and composition. Intracellular lipid bodies associate with the chloroplast and endoplasmic reticulum (ER) but show no evidence of being secreted. The ER displays striking fenestrations and forms a continuous subcortical system in direct contact with the cell membrane. The ECM has three distinct components. (i) Each cell is surrounded by a fibrous ␤-1, 4-and/or ␤-1, 3-glucan-containing cell wall. (ii) The intracolonial ECM space is filled with a cross-linked hydrocarbon network permeated with liquid hydrocarbons. (iii) Colonies are enclosed in a retaining wall festooned with a fibrillar sheath dominated by arabinose-galactose polysaccharides, which sequesters ECM liquid hydrocarbons. Each cell apex associates with the retaining wall and contributes to its synthesis. Retaining-wall domains also form "drapes" between cells, with some folding in on themselves and penetrating the hydrocarbon interior of a mother colony, partitioning it into daughter colonies. We propose that retaining-wall components are synthesized in the apical Golgi apparatus, delivered to apical ER fenestrations, and assembled on the surfaces of apical cell walls, where a proteinaceous granular layer apparently participates in fibril morphogenesis. We further propose that hydrocarbons are produced by the nonapical ER, directly delivered to the contiguous cell membrane, and pass across the nonapical cell wall into the hydrocarbon-based ECM.T he trebouxiophyte green fresh-to brackish-water alga Botryococcus braunii has a unique colonial organization: individual cells of the colony are embedded in an extracellular matrix (ECM) composed of polymerized and liquid hydrocarbons (LH) (29,35,(46)(47)(48). Although they may serve additional functions, these hydrocarbons notably allow B. braunii colonies to float, presumably to increase exposure to light for photosynthesis at the surfaces of ponds or lakes (4). The presence of B. braunii oils and ECM fossils in petroleum, coal deposits, and oil shale suggests that their hydrocarbon products once contributed to these reserves (1,2,8,10,16,17,34,36,52,62,63,69,70) and that these hydrocarbons could be used as a source of renewable energy. In fact, B. braunii hydrocarbons can be readily converted into petroleum equivalent, drop-in transportation fuels by conventional petroleum-processing techniques (22, 28).The three races of B. braunii are classified on the basis of the chemical nature of the liquid hydrocarbons they produce. The A race primarily produces alkadienes and alkatrienes derived from fatty acids (38,49,65,67); the B race (the focus of this study) produces two triterpenoids, tetramethylsqualene as a minor comp...

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