This Article Full Text Full Text (PDF) All Versions of this Article: 16/8/2151    most recent tpc.104.021972v1 Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Web of Science (28) Citing Articles via Google Scholar Google Scholar Articles by Posewitz, M. C. Articles by Ghirardi, M. L. Search for Related Content PubMed PubMed Citation Articles by Posewitz, M. C. Articles by Ghirardi, M. L. Agricola Articles by Posewitz, M. C. Articles by Ghirardi, M. L.

Hydrogen Photoproduction Is Attenuated by Disruption of an Isoamylase Gene in Chlamydomonas reinhardtii

^a National Renewable Energy Laboratory, Golden, Colorado 80401
^b University of California, Berkeley, California 94720

¹ To whom correspondence should be addressed. E-mail maria_ghirardi{at}nrel.gov; fax 303-384-6150.

DNA insertional transformants of Chlamydomonas reinhardtii were screened chemochromically for attenuated H₂ production. One mutant, displaying low H₂ gas photoproduction, has a nonfunctional copy of a gene that shows high homology to the family of isoamylase genes found in several photosynthetic organisms. DNA gel blotting and gene complementation were used to link this isoamylase gene to previously characterized nontagged sta7 mutants. This mutant is therefore denoted sta7-10. In C. reinhardtii, the STA7 isoamylase gene is important for the accumulation of crystalline starch, and the sta7-10 mutant reported here contains <3% of the glucose found in insoluble starch when compared with wild-type control cells. Hydrogen photoproduction rates, induced after several hours of dark, anaerobic treatment, are attenuated in sta7 mutants. RNA gel blot analysis indicates that the mRNA transcripts for both the HydA1 and HydA2 [Fe]-hydrogenase genes are expressed in the sta7-10 mutant at greater than wild-type levels 0.5 h after anaerobic induction. However, after 1.5 h, transcript levels of both HydA1 and HydA2 begin to decline rapidly and reach nearly undetectable levels after 7 h. In wild-type cells, the hydrogenase transcripts accumulate more slowly, reach a plateau after 4 h of anaerobic treatment, and maintain the same level of expression for >7 h under anaerobic incubation. Complementation of mutant cells with genomic DNA corresponding to the STA7 gene restores both the starch accumulation and H₂ production phenotypes. The results indicate that STA7 and starch metabolism play an important role in C. reinhardtii H₂ photoproduction. Moreover, the results indicate that mere anaerobiosis is not sufficient to maintain hydrogenase gene expression without the underlying physiology, an important aspect of which is starch metabolism.

This article has been cited by other articles:

				V. Chochois, D. Dauvillee, A. Beyly, D. Tolleter, S. Cuine, H. Timpano, S. Ball, L. Cournac, and G. Peltier Hydrogen Production in Chlamydomonas: Photosystem II-Dependent and -Independent Pathways Differ in Their Requirement for Starch Metabolism Plant Physiology, October 1, 2009; 151(2): 631 - 640. [Abstract] [Full Text] [PDF]

				T. Matthew, W. Zhou, J. Rupprecht, L. Lim, S. R. Thomas-Hall, A. Doebbe, O. Kruse, B. Hankamer, U. C. Marx, S. M. Smith, et al. The Metabolome of Chlamydomonas reinhardtii following Induction of Anaerobic H2 Production by Sulfur Depletion J. Biol. Chem., August 28, 2009; 284(35): 23415 - 23425. [Abstract] [Full Text] [PDF]

				M. Timmins, S. R. Thomas-Hall, A. Darling, E. Zhang, B. Hankamer, U. C. Marx, and P. M. Schenk Phylogenetic and molecular analysis of hydrogen-producing green algae J. Exp. Bot., April 2, 2009; (2009) erp052v1. [Abstract] [Full Text] [PDF]

				F. Jans, E. Mignolet, P.-A. Houyoux, P. Cardol, B. Ghysels, S. Cuine, L. Cournac, G. Peltier, C. Remacle, and F. Franck A type II NAD(P)H dehydrogenase mediates light-independent plastoquinone reduction in the chloroplast of Chlamydomonas PNAS, December 23, 2008; 105(51): 20546 - 20551. [Abstract] [Full Text] [PDF]

				S. Streb, T. Delatte, M. Umhang, S. Eicke, M. Schorderet, D. Reinhardt, and S. C. Zeeman Starch Granule Biosynthesis in Arabidopsis Is Abolished by Removal of All Debranching Enzymes but Restored by the Subsequent Removal of an Endoamylase PLANT CELL, December 1, 2008; 20(12): 3448 - 3466. [Abstract] [Full Text] [PDF]

				A. V. Nguyen, S. R. Thomas-Hall, A. Malnoe, M. Timmins, J. H. Mussgnug, J. Rupprecht, O. Kruse, B. Hankamer, and P. M. Schenk Transcriptome for Photobiological Hydrogen Production Induced by Sulfur Deprivation in the Green Alga Chlamydomonas reinhardtii Eukaryot. Cell, November 1, 2008; 7(11): 1965 - 1979. [Abstract] [Full Text] [PDF]

				R. Surzycki, L. Cournac, G. Peltier, and J.-D. Rochaix Potential for hydrogen production with inducible chloroplast gene expression in Chlamydomonas PNAS, October 30, 2007; 104(44): 17548 - 17553. [Abstract] [Full Text] [PDF]

				F. Mus, A. Dubini, M. Seibert, M. C. Posewitz, and A. R. Grossman Anaerobic Acclimation in Chlamydomonas reinhardtii: ANOXIC GENE EXPRESSION, HYDROGENASE INDUCTION, AND METABOLIC PATHWAYS J. Biol. Chem., August 31, 2007; 282(35): 25475 - 25486. [Abstract] [Full Text] [PDF]

				O. Kruse, J. Rupprecht, K.-P. Bader, S. Thomas-Hall, P. M. Schenk, G. Finazzi, and B. Hankamer Improved Photobiological H2 Production in Engineered Green Algal Cells J. Biol. Chem., October 7, 2005; 280(40): 34170 - 34177. [Abstract] [Full Text] [PDF]

				S. Fouchard, A. Hemschemeier, A. Caruana, J. Pruvost, J. Legrand, T. Happe, G. Peltier, and L. Cournac Autotrophic and Mixotrophic Hydrogen Photoproduction in Sulfur-Deprived Chlamydomonas Cells Appl. Envir. Microbiol., October 1, 2005; 71(10): 6199 - 6205. [Abstract] [Full Text] [PDF]