Localization of putative carbonic anhydrases in two marine diatoms, Phaeodactylum tricornutum and Thalassiosira pseudonana

Abstract

It is believed that intracellular carbonic anhydrases (CAs) are essential components of carbon concentrating mechanisms in microalgae. In this study, putative CA-encoding genes were identified in the genome sequences of the marine diatoms Phaeodactylum tricornutum and Thalassiosira pseudonana. Subsequently, the subcellular localizations of the encoded proteins were determined. Nine and thirteen CA sequences were found in the genomes of P. tricornutum and T. pseudonana, respectively. Two of the β-CA genes in P. tricornutum corresponded to ptca1 and ptca2 identified previously. Immunostaining transmission electron microscopy of a PtCA1:YFP fusion expressed in the cells of P. tricornutum clearly showed the localization of PtCA1 within the central part of the pyrenoid structure in the chloroplast. Besides these two β-CA genes, P. tricornutum likely contains five α- and two γ-CA genes, whereas T. pseudonana has three α-, five γ-, four δ-, and one ζ-CA genes. Semi-quantitative reverse transcription PCR performed on mRNA from the two diatoms grown in changing light and CO(2) conditions revealed that levels of six putative α- and γ-CA mRNAs in P. tricornutum did not change between cells grown in air-level CO(2) and 5% CO(2). However, mRNA levels of one putative α-CA gene, CA-VII in P. tricornutum, were reduced in the dark compared to that in the light. In T. pseudonana, mRNA accumulation levels of putative α-CA (CA-1), ζ-CA (CA-3) and δ-CA (CA-7) were analyzed and all levels found to be significantly reduced when cells were grown in 0.16% CO(2). Intercellular localizations of eight putative CAs were analyzed by expressing GFP fusion in P. tricornutum and T. pseudonana. In P. tricornutum, CA-I and II localized in the periplastidial compartment, CA-III, VI, VII were found in the chloroplast endoplasmic reticulum, and CA-VIII was localized in the mitochondria. On the other hand, T. pseudonana CA-1 localized in the stroma and CA-3 was found in the periplasm. These results suggest that CAs are constitutively present in the four chloroplastic membrane systems in P. tricornutum and that CO(2) responsive CAs occur in the pyrenoid of P. tricornutum, and in the stroma and periplasm of T. pseudonana.

Light and CO2/cAMP Signal Cross Talk on the Promoter Elements of Chloroplastic β-Carbonic Anhydrase Genes in the Marine Diatom Phaeodactylum tricornutum

“…PtCA2 is a paralogous CA to PtCA1, with an approximately 80% amino acid sequence identity whose transcription is also CO 2 -responsive, and they colocalize at the pyrenoid of P. tricornutum Tanaka et al, 2005;Tachibana et al, 2011). Both CAs are under redox regulation by the function of diatom plastidic thioredoxins (Kikutani et al, 2012) and the function of the promoters of both CA genes are responsive to CO 2 via the function of cAMP ( Fig.…”

“…Numerous CAs, a critical CCM component, were also found in the genomes of P. tricornutum and T. pseudonana and most of them have been localized Tanaka et al, 2005;Tachibana et al, 2011;Samukawa et al, 2014). Interestingly, there are significant differences in the location of CAs between these two species.…”

The place of diatoms in the biofuels industry

“…In diatoms, evidence for both C 3 and C 4 biochemical fixation exists, and the preference can apparently differ in relatively closely related species [58], which has led to confusion in attempts to make generalizations. P. tricornutum and T. pseudonana encode 9 and 13 CAs, respectively, but only P. tricornutum has b-CAs that are targeted to the chloroplast [59]. It is as yet unclear whether differences in CCMs relate to the different evolutionary time periods under which different classes of diatoms arose in relation to CO 2 levels.…”

Inorganic carbon utilization by aquatic photoautotrophs and potential usages of algal primary production

“…The Diatom CCM is also regulated by pCO 2 , and recent progress in molecular studies on the transcriptional control of CCM components in response to pCO 2 have revealed that cAMP is a second messenger ). There are redundant CA genes in genomes of two model marine diatoms, Phaeodactylum tricornutum, and Thalassiosira pseudonana (Tachibanal et al 2011). In P. tricornutum, all 5 a-CAs were localized at the four-layered chloroplast membrane system whereas the 2 b-CAs were localized in the pyrenoid and one c-CA in the mitochondria (Tachibanal et al 2011), which provide a set of data to support the predominant operation of a biophysical CCM in P. tricornutum.…”

“…There are redundant CA genes in genomes of two model marine diatoms, Phaeodactylum tricornutum, and Thalassiosira pseudonana (Tachibanal et al 2011). In P. tricornutum, all 5 a-CAs were localized at the four-layered chloroplast membrane system whereas the 2 b-CAs were localized in the pyrenoid and one c-CA in the mitochondria (Tachibanal et al 2011), which provide a set of data to support the predominant operation of a biophysical CCM in P. tricornutum. In T. pseudonana, one a-CA and one f-CA were localized to the stroma and the periplasm, respectively and these CAs were induced under CO 2 limitation (Tachibanal et al 2011).…”