Accessible Requires Authentication Published by De Gruyter January 6, 2021

Sequence comparison and expression analysis of an inferred Na+/Pi cotransporter gene in the marine diatom Skeletonema tropicum

Shr-Hau Hung, Yung-Hsiu Lu, Chih-Ching Chung, Chi-Yu Shih, Gwo-Ching Gong and Jeng Chang
From the journal Botanica Marina


Unicellular algae have evolved to express many forms of high-affinity phosphate transporters, and homologs of these proteins are broadly distributed in yeast, fungi, higher plants, and vertebrates. In this report, an effort has been made to characterize such a transporter gene, StPHO, in the marine diatom Skeletonema tropicum. The primers used for polymerase chain reaction were designed by referring to a homologous gene in a prasinophyte, and the full-length (1692 bp) cDNA of StPHO was then cloned and sequenced. Sequence alignments and secondary structure prediction indicated that StPHO is a gene that encodes a type III Na+/Pi cotransporter (SLC20 family). To study the function of StPHO, specific concentrations of inorganic phosphate (Pi) were used to alter the physiological status of S. tropicum. In each treatment, samples were collected for the measurements of StPHO mRNA, [PO43−], cell abundance, the maximal photochemical efficiency of photosystem II (Fv/Fm), and alkaline phosphatase activity (APA). The results indicated that the ambient [PO43−] strongly affected the population growth and related physiological parameters of S. tropicum. The transcription of StPHO was fully repressed when the [PO43−] was greater than 1 μM but increased approximately 100-fold when the ambient [PO43−] decreased to 0.02 μM. Within this [PO43−] range, the regression equations are Y = −0.6644X + 0.9034 and Y = −0.5908X + 0.8054 for Pi-starved and Pi-limited treatments, respectively. This trend of gene expression suggested that StPHO plays an important role in the uptake of [PO43−], and StPHO may serve as a useful molecular biomarker for Pi-stressed diatom populations in marine ecosystems.

Corresponding author: Jeng Chang, Institute of Marine Biology, National Taiwan Ocean University, Keelung20224, Taiwan, ROC; Institute of Marine Environment and Ecology, National Taiwan Ocean University, Keelung20224, Taiwan, ROC; and Center of Excellence for the Oceans, National Taiwan Ocean University, Keelung20224, Taiwan, ROC, E-mail:

Funding source: Ministry of Science and Technology

Award Identifier / Grant number: MOST 108-2811-M-019-505, MOST 109-2611-M-019-003, NSC 99-2313-B-019-002


We deeply appreciate Drs. L.-K. Kang, S.-P. L. Hwang, and P.-H. Hsu for technical instructions and helpful discussions. We would like to thank Dr. F.-K. Shiah for providing the FIRe fluorometer, C.-Y. Chen for assistance in measuring the phosphate concentrations, and T.-H. Kuo and X.-B. Chen for monitoring the bacterial abundance. We are grateful to the Core Facility of the Institute of Cellular and Organismic Biology, Academia Sinica, for assistance with DNA sequencing.

  1. Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.

  2. Research funding: This study was supported by the grants MOST 109-2611-M-019-003 and NSC 99-2313-B-019-002 from the Ministry of Science and Technology (Taiwan, ROC). CYS was supported through MOST 108-2811-M-019-505.

  3. Conflict of interest statement: The authors declare no conflicts of interest regarding this article.


Armbrust, E.V. (2009). The life of diatoms in the world’s oceans. Nature 459: 185–192. Search in Google Scholar

Chung, C.-C., Hwang, S.-P.L., and Chang, J. (2003). Identification of a high-affinity phosphate transporter gene in a prasinophyte alga, Tetraselmis chui, and its expression under nutrient limitation. Appl. Environ. Microbiol. 69: 754–759. Search in Google Scholar

Chung, C.-C., Hwang, S.-P.L., and Chang, J. (2005). Cooccurrence of ScDSP gene expression, cell death, and DNA fragmentation in a marine diatom, Skeletonema costatum. Appl. Environ. Microbiol. 71: 8744–8751. Search in Google Scholar

Cruz de Carvalho, M.H., Sun, H.-X., Bowler, C., and Chua, N.-H. (2016). Noncoding and coding transcriptome responses of a marine diatom to phosphate fluctuations. New Phytol. 210: 497–510. Search in Google Scholar

Daram, P., Brunner, S., Rausch, C., Steiner, C., Amrhein, N., and Bucher, M. (1999). Pht2;1 encodes a low-affinity phosphate transporter from Arabidopsis. Plant Cell 11: 2153–2166. Search in Google Scholar

Dyhrman, S.T., Jenkin, B.D., Rynearson, T.A., Saito, M.A., Mercier, M.L., Alexander, H., Whitney, L.P., Drzewianowski, A., Bulygin, V.V., Bertrand, E.M., et al.. (2012). The transcriptome and proteome of the diatom Thalassiosira pseudonana reveal a diverse phosphorus stress response. PloS One 7: e33768. Search in Google Scholar

Falkowski, P.G. (2002). The ocean’s invisible forest. Sci. Am. 287: 54–61. Search in Google Scholar

Falkowski, P.G., Barber, R. T., and Smetacek, V. (1998). Biogeochemical controls and feedbacks on ocean primary production. Science 281: 200–206. Search in Google Scholar

Gallina, A.A., Chung, C.-C., and Casotti, R. (2015). Expression of death-related genes and reactive oxygen species production in Skeletonema tropicum upon exposure to the polyunsaturated aldehyde octadienal. Adv. Oceanogr. Limnol. 6: 13–20. Search in Google Scholar

Guillard, R.R.L. and Ryther, J.H. (1962). Studies of marine planktonic diatoms: I. Cyclotella nana hustedt, and Detonula confervacea (Cleve) Gran. Can. J. Microbiol. 8: 229–239. Search in Google Scholar

Hediger, M.A., Romero, M.F., Peng, J.B., Rolfs, A., Takanaga, H., and Bruford, E.A. (2004). The ABCs of solute carriers: physiological, pathological and therapeutic implications of human membrane transport proteins. Pflugers Arch. Eur. J. Physiol. 447: 465–468. Search in Google Scholar

Hung, S.-H., Chung, C.-C., Liao, C.-W., Gong, G.-C., and Chang, J. (2013). Sequence diversity and expression levels of Synechococcus phosphate transporter gene in the East China Sea. J. Exp. Mar. Biol. Ecol. 440: 90–99. Search in Google Scholar

Hwang, S.-P.L., Wang, S.K., Wei, S.F., Cheng, L.-C., and Chang, J. (1999). Identification and expression pattern of DNA polymerase α gene in a marine diatom, Skeletonema costatum. Mar. Biotechnol. 1: 200–206. Search in Google Scholar

Kang, L.-K., Gong, G.-C., Wu, Y.-H., and Chang, J. (2015). The expression of nitrate transporter genes reveals different nitrogen statuses of dominant diatom groups in the southern East China Sea. Mol. Ecol. 24: 1374–1386. Search in Google Scholar

Kester, D.R., Duedall, I.W., Connors, D.N., and Pytkowicz, R.M. (1967). Preparation of artificial seawater. Limnol. Oceanogr. 12: 176–179. Search in Google Scholar

Kumar, S., Stecher, G., Li, M., Knyaz, C., and Tamura, K. (2018). MEGA X: molecular evolutionary genetics analysis across computing platforms. Mol. Biol. Evol. 35: 1547–1549. Search in Google Scholar

La Roche, J., McKay, R.M.L., and Boyd, P. (1999). Immunological and molecular probes to detect phytoplankton responses to environmental stress in nature. Hydrobiologia 401: 177–198. Search in Google Scholar

Lu, Y.-H. (2004). Performance comparisons between polymerase chain reaction and dot blotting in differentiating phytoplankton genes from distinct species, Master’s thesis. Keelung, Taiwan, National Taiwan Ocean University. Search in Google Scholar

Martinez, P. and Persson, B.L. (1998). Identification, cloning and characterization of a derepressible Na+-coupled phosphate transporter in Saccharomyces cerevisiae. Mol. Gen. Genet. 258: 628–638. Search in Google Scholar

Młodzińska, E. and Zboińska, M. (2016). Phosphate uptake and allocation – a closer look at Arabidopsis thaliana L. and Oryza sativa L. Front. Plant Sci. 7: 1198. Search in Google Scholar

Pai, S.C., Yang, C.C., and Riley, J.P. (1990). Effects of acidity and molybdate concentration on the kinetics of the formation of the phosphoantimonylmolybdenum blue complex. Anal. Chim. Acta 229: 115–120. Search in Google Scholar

Parkhill, J.P., Maillet, G., and Cullen, J.J. (2001). Fluorescence-based maximal quantum yield for PSII as a diagnostic of nutrient stress. J. Phycol. 37: 517–529. Search in Google Scholar

Pedersen, B.P., Kumar, H., Waight, A.B., Risenmay, A.J., Roe-Zurz, Z., Chau, B.H., Schlessinger, A., Bonomi, M., Harries, W., Sali, A., et al.. (2013). Crystal structure of a eukaryotic phosphate transporter. Nature 496: 533–536. Search in Google Scholar

Perry, M.J. (1976). Phosphate utilization by an oceanic diatom in phosphorus‐limited chemostat culture and in the oligotrophic waters of the central North Pacific. Limnol. Oceanogr. 21: 88–107. Search in Google Scholar

Persson, B.L., Berhe, A., Fristedt, U., Martinez, P., Pattison, J., Petersson, J., and Weinander, R. (1998). Phosphate permeases of Saccharomyces cerevisiae. Biochim. Biophys. Acta 1365: 23–30. Search in Google Scholar

Rychter, A.M. and Rao, I.M. (2005). Role of phosphorus in photosynthetic carbon metabolism. In: Pessarakli, M. (Ed.). Handbook of photosynthesis. Taylor and Francis, Boca Raton, pp. 123–148. Search in Google Scholar

Scanlan, D.J., Mann, N.H., and Carr, N.G. (1993). The response of the picoplanktonic marine cyanobacterium Synechococcus species WH7803 to phosphate starvation involves a protein homologous to the periplasmic phosphate‐binding protein of Escherichia coli. Mol. Microbiol. 10: 181–191. Search in Google Scholar

Shih, C.-Y., Kang, L.-K., and Chang, J. (2015). Transcriptional responses to phosphorus stress in the marine diatom, Chaetoceros affinis, reveal characteristic genes and expression patterns in phosphorus uptake and intracellular recycling. J. Exp. Mar. Biol. Ecol. 470: 43–54. Search in Google Scholar

Theodorou, M.E. and Plaxton, W.C. (1993). Metabolic adaptations of plant respiration to nutritional phosphate deprivation. Plant Physiol. 101: 339–344. Search in Google Scholar

Van Mooy, B.A.S., Fredricks, H.F., Pedler, B.E., Dyhrman, S.T., Karl, D.M., Koblížek, M., Lomas, M.W., Mincer, T.J., Moore, L.R., Moutin, T., et al.. (2009). Phytoplankton in the ocean use non-phosphorus lipids in response to phosphorus scarcity. Nature 458: 69–72. Search in Google Scholar

Versaw, W.K. and Harrison, M.J. (2002). A chloroplast phosphate transporter, PHT2;1, influences allocation of phosphate within the plant and phosphate-starvation responses. Plant Cell 14: 1751–1766. Search in Google Scholar

Virkki, L. V., Biber, J., Murer, H., and Forster, I.C. (2007). Phosphate transporters: a tale of two solute carrier families. Am. J. Physiol. Ren. Physiol. 293: 643–654. Search in Google Scholar

Wawrik, B., Paul, J.H., and Tabita, F.R. (2002). Real-time PCR quantification of rbcL (ribulose-1,5-bisphosphate carboxylase/oxygenase) mRNA in diatoms and pelagophytes. Appl. Environ. Microbiol. 68: 3771–3779. Search in Google Scholar

Xu, H., Inouye, M., Missey, T., Collins, J.F., and Ghishan, F.K. (2002). Functional characterization of the human intestinal NaPi-IIb cotransporter in hamster fibroblasts and Xenopus oocytes. Biochim. Biophys. Acta 1567: 97–105. Search in Google Scholar

Zhang, S.-F., Yuan, C.-J., Chen, Y., Chen, X.-H., Li, D.-X., Liu, J.-L., Lin, L., and Wang, D.-Z. (2016). Comparative transcriptomic analysis reveals novel insights into the adaptive response of Skeletonema costatum to changing ambient phosphorus. Front. Microbiol. 7: 1476. Search in Google Scholar

Supplementary Material

The online version of this article offers supplementary material (

Received: 2020-05-29
Accepted: 2020-12-11
Published Online: 2021-01-06
Published in Print: 2021-02-23

© 2020 Walter de Gruyter GmbH, Berlin/Boston