Renewable H-2 photoproduction by green algae such as Chlamydomonas reinhardtii is a promising system for solar fuels. However, large-scale application of the system has lagged virtually due to lack of high H-2-producing strains. We previously identified ferredoxin-NADP(+) reductase (FNR) among the 105 proteins differentially expressed in Chlamydomonas during sulfur-deprived H-2 photoproduction. In this work, we used an RNA interference (RNAi) approach to generate Chlamydomonas mutant strains with reduced levels of FNR. We found that fnr-RNAi strains exhibited higher rates of H-2 photoproduction (2.5-fold) than wild type under sulfur-deprived condition. To elucidate the basis for this increase, we analyzed the physiological characteristics of the fnr-RNAi strains under such condition. Major changes, due to the down-regulation of FNR, included the lower rates of photosynthetic O-2 evolution (44%), greater reduction of Rubisco amounts (60%) and higher rates of starch degradation (140%). These may result in an earlier onset of anaerobiosis and increased electron supply to the hydrogenases in the mutant strains. The results provide new information of FNR in regulating H-2 metabolism as well as potential strains for further improvement of the organism toward application in solar-powered systems. Copyright (C) 2013, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.