The Columbia River (CR) is a powerful economic and environmental driver in the US Pacific Northwest. Microbial communities in the water column were analyzed from four diverse habitats: (1) an estuarine turbidity maximum (ETM), (2) a chlorophyll maximum of the river plume, (3) an upwelling-associated hypoxic zone, and (4) the deep ocean bottom. Three size fractions, 0.1–0.8, 0.8–3, and 3–200 μm were collected for each habitat in August 2007, and used for DNA isolation and 454 sequencing, resulting in 12 metagenomes of >5 million reads (>1.6 Gbp). To characterize the dominant microorganisms and metabolisms contributing to coastal biogeochemistry, we used predicted peptide and rRNA data. The 3- and 0.8-μm metagenomes, representing particulate fractions, were taxonomically diverse across habitats. The 3-μm size fractions contained a high abundance of eukaryota with diatoms dominating the hypoxic water and plume, while cryptophytes were more abundant in the ETM. The 0.1-μm metagenomes represented mainly free-living bacteria and archaea. The most abundant archaeal hits were observed in the deep ocean and hypoxic water (19% of prokaryotic peptides in the 0.1-μm metagenomes), and were homologous to Nitrosopumilus maritimus (ammonia-oxidizing Thaumarchaeota). Bacteria dominated metagenomes of all samples. In the euphotic zone (estuary, plume and hypoxic ocean), the most abundant bacterial taxa (≥40% of prokaryotic peptides) represented aerobic photoheterotrophs. In contrast, the low-oxygen, deep water metagenome was enriched with sequences for strict and facultative anaerobes. Interestingly, many of the same anaerobic bacterial families were enriched in the 3-μm size fraction of the ETM (2–10X more abundant relative to the 0.1-μm metagenome), indicating possible formation of anoxic microniches within particles. Results from this study provide a metagenome perspective on ecosystem-scale metabolism in an upwelling-influenced river-dominated coastal margin.