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Soil microorganisms are considered C-limited, while plant productivity is frequently N-limited. Large stores of organic C in boreal forest soils are attributed to negative effects of low temperature, soil acidity and plant residue recalcitrance upon microbial activity. We examined microbial activity, biomass and community composition along a natural 90-m-long soil N supply gradient, where plant species composition varies profoundly, forest productivity three-fold and soil pH by three units. There was, however, no significant variation in soil respiration in the field across the gradient. Neither did microbial biomass C determined by fumigation-extraction vary, while other estimates of activity and biomass showed a weak increase with increasing N supply and soil pH. Simultaneously, a phospholipid fatty acid attributed mainly to mycorrhizal fungi declined drastically, while bacterial biomass increased. We hypothesize that low N supply and plant productivity, and hence low litter C supply to saprotrophs is associated with a high plant C supply to mycorrhizal fungi, while the reverse occurs under high N supply. This should mean that effects of N availability on C supply to these functional groups of microbes acts in opposing directions.