ALB3 Insertase Mediates Cytochrome b6 Co-translational Import into the Thylakoid Membrane

The cytochrome b(6)f complex occupies an electrochemically central position in the electron-transport chain bridging the photosynthetic reaction center of PS I and PS II. In plants, the subunits of these thylakoid membrane protein complexes are both chloroplast and nuclear encoded. How the chloroplast-encoded subunits of multi-spanning cytochrome b(6) are targeted and inserted into the thylakoid membrane is not fully understood. Experimental approaches to evaluate the cytochrome b(6) import mechanism in vivo have been limited to bacterial membranes and were not a part of the chloroplast environment. To evaluate the mechanism governing cytochrome b(6) integration in vivo, we performed a comparative analysis of both native and synthetic cytochrome b(6) insertion into purified thylakoids. Using biophysical and biochemical methods, we show that cytochrome b(6) insertion into the thylakoid membrane is a non-spontaneous co-translational process that involves ALB3 insertase. Furthermore, we provided evidence that CSP41 (chloroplast stem-loop-binding protein of 41 kDa) interacts with RNC-cytochrome b(6) complexes, and may be involved in cytochrome b(6) (petB) transcript stabilization or processing.