Physiologia Plantarum 133: 78–91. 2008 Copyright ª Physiologia Plantarum 2007, ISSN 0031-9317 Expression and regulation of carbonic anhydrases in the marine diatom Thalassiosira pseudonana and in natural phytoplankton assemblages from Great Bay, New Jersey Patrick J. McGinn* and Franc xois M. M. Morel Department of Geosciences, Guyot Hall, Princeton University, Princeton, NJ 08544, USA Correspondence *Corresponding author, e-mail: pmcginn@princeton.edu Received 3 October 2007; revised 19 November 2007 doi: 10.1111/j.1399-3054.2007.01039.x BLAST searches of expressed sequence tag libraries have revealed putative homologs of the archetypal diatom d-carbonic anhydrase (CA) TWCA1 (for Thalassiosira weissflogii CA) in a broad range of eukaryotic phytoplankton including haptophytes, prasinophytes and dinoflagellates. Four putative homologs of TWCA1 are also reported and described from a search of the genomic sequence of Thalassiosira pseudonana and designated TWCA Tp1–4 . The d-CA class is therefore more widely distributed in marine phytoplankton than previously thought. In particular, it is not restricted to the diatoms like the cadmium-containing enzyme, CDCA, seems to be (z-CA class). Zinc status strongly influences growth rate in marine diatoms. We observed a decrease in the specific growth rate of T. pseudonana from 2.1 to 1.3 day 21 when the unchelated Zn concentration (Zn#) was lowered from 20 to 5 pM. In the same cultures, we detected a drop in whole-cell CA activity of about 50% in the most Zn-limited cells that occurred simultaneously with a large downregulation of TWCA Tp1 , TWCA Tp2 and CDCA Tp gene transcripts and protein. These three genes were also found to be strongly upregulated by low pCO 2 in a manner typical of many algal CA enzymes. We have also conducted field experiments in diatom-dominated natural phytoplankton assemblages sampled from Great Bay of the coast of New Jersey. We observed increased total CA activity in parallel with increased expression of homologous twca and cdca gene transcripts in water incubated at increasingly higher pH values. Immunoblot and transcript expression analyses demonstrated a clear upregulation of CDCA transcript and protein as incubation pH increased both in the lab and in the field indicating that this CA is expressed under a broad range of environmental conditions and not restricted to low pCO 2 and low Zn. Introduction Carbonic anhydrases (CAs, EC 4.2.1.1) are a large and diverse collection of Zn metalloenzymes that catalyze the equilibration of dissolved inorganic carbon species according to the following equation: CO 2 1 H 2 O4HCO 2 3 1 H 1 The essential nature of this reaction in diverse biological systems has driven the evolution of several distinct and unrelated families of CAs (Hewett-Emmett and Tashian 1996). Animals possess a-CAs that are Abbreviations – CA, carbonic anhydrase; CDCA1, cadmium-containing z-class CA isolated from Thalassiosira weissflogii; cdca Tp / CDCA Tp , homologous cdca gene/protein in T. pseudonana; EST, expressed sequence tag; pCO 2 , partial pressure of CO 2 ; TWCA1 227 kDa, Zn(Co)-dependent d-class CA isolated from T. weissflogii; twca Tp / TWCA Tp , homologous twca gene/protein in T. pseudonana. 78 Physiol. Plant. 133, 2008