22. Kassen, R., Buckling, A., Bell, G. & Rainey, P. B.Diversity peaks at intermediate productivity in a laboratory microcosm. Nature 406, 508–512 (2000). 23. Ayala, F. J. & Campbell, C. A. Frequency-dependent selection. Annu. Rev. Ecol. Syst. 5, 115–138 (1974). 24. Rosenzweig, M. L. Species Diversity in Space and Time (Cambridge Univ. Press, Cambridge, 1995). 25. Buckling, A. & Rainey, P. B. Antagonistic coevolution between a bacterium and a bacteriophage. Proc. R. Soc. Lond. B 269, 931–936 (2002). 26. Lenski, R. E. & Levin, B. R. Constraints on the coevolution of bacteria and virulent phage—a model, some experiments, and predictions for natural communities. Am. Nat. 125, 585–602 (1985). 27. Lande, R. Statistics and partitioning of species diversity, and similarity among multiple communities. Oikos 76, 5–13 (1996). 28. Holt, R. D., Grover, J. & Tilman, D. Simple rules for interspecific dominance in systems with exploitative and apparent competition. Am. Nat. 144, 741–771 (1994). 29. Leibold, M. A graphical model of keystone predators in food webs: trophic regulation of abundance, incidence and diversity patterns in communities. Am. Nat. 147, 784–812 (1996). 30. Simpson, E. H. Measurement of diversity. Nature 163, 688 (1949). Acknowledgements We thank M. Brockhurst, L. Hurst and S. West for comments on the manuscript. This work was supported by NERC (UK) and the Royal Society. Competing interests statement The authors declare that they have no competing financial interests. Correspondence and requests for materials should be addressed to A.B. (e-mail: bssagjb@bath.ac.uk). .............................................................. Calcium activation of BK Ca potassium channels lacking the calcium bowl and RCK domains Rebecca Piskorowski & Richard W. Aldrich Department of Molecular and Cellular Physiology, and Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, California 94305-5345, USA ............................................................................................................................................................................. In many physiological systems such as neurotransmitter release, smooth muscle relaxation and frequency tuning of auditory hair cells, large-conductance calcium-activated potassium (BK Ca ) channels create a connection between calcium signalling path- ways and membrane excitability 1–4 . BK Ca channels are activated by voltage and by micromolar concentrations of intracellular calcium. Although it is possible to open BK Ca channels in the absence of calcium 5–9 , calcium binding is essential for their activation under physiological conditions. In the presence of intracellular calcium, BK Ca channels open at more negative membrane potentials 5,10–14 . Many experiments investigating the molecular mechanism of calcium activation of the BK Ca channel have focused on the large intracellular carboxy terminus, and much evidence supports the hypothesis that calcium-binding sites are located in this region of the channel. Here we show that BK Ca channels that lack the whole intracellular C terminus retain wild-type calcium sensitivity. These results show that the intra- cellular C terminus, including the ‘calcium bowl’ and the RCK domain, is not necessary for the calcium-activated opening of these channels. Several experimental results imply that the intracellular C termi- nus of the BK Ca channel mediates Ca 2þ binding and/or activation. The channel’s pore-forming subunit is shown in Fig. 1a. Mutations in a conserved group of aspartic acid residues, called the ‘calcium bowl’, change the ability of low concentrations of Ca 2þ to activate the channel 15 . Additional experiments indicate that the last 400 residues of the C terminus (which includes the calcium bowl) are important in Ca 2þ activation of the BK Ca channel 16 . Removal of these amino acids yields a non-functional BK Ca channel, and co- expression of the 400 amino acid fragment with a shortened channel restores normal behaviour 16,17 . Replacement of this region by the corresponding region of a BK Ca channel homologue that lacks Ca 2þ sensitivity (Slo3) results in a channel with greatly decreased Ca 2þ sensitivity 16 . In addition, this region of the C terminus binds Ca 2þ in 45 Ca 2þ -overlay protein blot assays 18,19 , and combinations of point mutations in the calcium bowl and RCK domains eliminate Ca 2þ activation 20,21 . The structure of a bacterial K þ channel gated by millimolar Ca 2þ has been solved in the presence of 200 mM Ca 2þ (ref. 22), and Ca 2þ -binding sites are clearly evident in the C-terminal RCK domain. Taken together, these results have led to the commonly accepted conclusion that the calcium bowl and the RCK domain, either separately or together, contain Ca 2þ -binding sites that are involved in activation of the channel by micromolar concentrations of Ca 2þ . If this idea is correct, then removing these domains should greatly alter the Ca 2þ activation of the channel. Counter to this prediction, we found that a truncated channel that lacks the whole intracellular C terminus (made by deleting the amino acids after residue 323; Fig. 1a) retains Ca 2þ sensitivity (Fig. 2). Single-channel current traces (Figs 1b and 2a) and quantified voltage- and Ca 2þ -dependent activity (Fig. 2b, c) of wild-type and truncated channels showed that the truncated channel is activated over the same range of Ca 2þ concentrations as the full-length channel. Thus, a pore-forming domain with an intact C-terminal domain is not required for voltage- and Ca 2þ -dependent gating. Given the variability in gating between the different channels (Fig. 2c), it was unclear whether the apparent change in voltage dependence (Fig. 2b) is a genuine property of the truncated channel. The behaviour of the truncated and full-length channels was not identical: single-channel kinetics showed that the truncated channel has a shorter mean open time (Fig. 3), indicating that the transition energy of the open state of the channel has been altered. Expression of the truncated channel, as determined by electro- physiological recordings, was roughly a thousand-fold less than that Figure 1 The truncated BK Ca channel retains normal function. a, The truncated BK Ca channel. The different domains of the intracellular C terminus and the aspartic acids in the calcium bowl are indicated. The ‘tail’ corresponds the to last 400 amino acids of the C terminus that have been proposed to be important in Ca 2þ activation of BK Ca channels. b, Single-channel currents measured at 100 mM Ca 2þ from full-length and truncated channels heterologously expressed in Xenopus oocytes. letters to nature NATURE | VOL 420 | 5 DECEMBER 2002 | www.nature.com/nature 499 © 2002 Nature Publishing Group