The BtpA protein stabilizes the reaction center proteins of photosystem I in the cyanobacterium Synechocystis sp. PCC 6803 at low temperature

Abstract

Specific inhibition of photosystem I (PSI) was observed under low-temperature conditions in the cyanobacterium Synechocystis sp. strain PCC 6803. Growth at 20 degrees C caused inhibition of PSI activity and increased degradation of the PSI reaction center proteins PsaA and PsaB, while no significant changes were found in the level and activity of photosystem II (PSII). BtpA, a recently identified extrinsic thylakoid membrane protein, was found to be a necessary regulatory factor for stabilization of the PsaA and PsaB proteins under such low-temperature conditions. At normal growth temperature (30 degrees C), the BtpA protein was present in the cell, and its genetic deletion caused an increase in the degradation of the PSI reaction center proteins. However, growth of Synechocystis cells at 20 degrees C or shifting of cultures grown at 30 degrees C to 20 degrees C led to a rapid accumulation of the BtpA protein, presumably to stabilize the PSI complex, by lowering the rates of degradation of the PsaA and PsaB proteins. A btpA deletion mutant strain could not grow photoautotrophically at low temperature, and exhibited rapid degradation of the PSI complex after transfer of the cells from normal to low temperature.

Figure 1

Fluorescence emission spectra (77 K) from whole cells of wild-type (WT) and BP26 mutant strains grown at 30°C or 20°C. Excitation was at 600 ± 5 nm. The spectra were corrected for the wavelength characteristics of the emission monochromator and the response of the signal detector. See “Materials and Methods” for experimental details.

Figure 2

Presence of PSI and PSII proteins in wild-type and mutant strains grown at 30°C or 20°C. Membrane proteins from wild-type (WT), BP26, btpA-deletion (ΔbtpA), and psaA/B-deletion (ΔpsaAB) strains were fractionated on SDS-PAGE (50 μg protein per lane), transferred to nitrocellulose filters, and immunostained with antibodies against PsaA, PsaB, PsaC, PsaD, PsaF, PsaK, PsaL, CP43, and BtpA proteins, respectively.

Figure 3

Stability of PSI and PSII proteins at 30°C. Wild-type (WT), BP26, and btpA-deletion (ΔbtpA) strains were incubated in the presence of chloramphenicol (150 μg/mL), and aliquots of cells were collected immediately (0), and 3, 6, and 12 h later. Membrane proteins were fractionated on SDS-PAGE, transferred to nitrocellulose filters, and immunostained with antibodies against PsaA, PsaB, PsaL, CP43, D1, and BtpA proteins, respectively. Seventy micrograms of protein-containing sample was loaded in each lane. N/D, Not determined.

Figure 4

Stability of PSI and PSII proteins at 20°C. Wild-type (WT) and BP26 mutant strains were incubated in the presence of chloramphenicol (150 μg/mL), and aliquots of cells were collected immediately (0), and 3, 6, and 12 h later. Membrane proteins were fractionated on SDS-PAGE, transferred to nitrocellulose filters, and immunostained with antibodies against PsaA, PsaB, and PsaL proteins, respectively. Seventy micrograms of protein-containing sample was loaded in each lane.

Figure 5

Changes in the amounts of PSI proteins after a temperature shift from 30°C to 20°C. Wild-type (A) and BP26 (B) mutant cells were grown at 30°C and then transferred to 20°C for 24 h without (lanes 2) or with (lanes 3) chloramphenicol (150 μg/mL). Lanes 1, Control cells maintained at 30°C. Membrane proteins were separated on SDS-PAGE, transferred to nitrocellulose membranes, and probed with antibodies against PsaA and PsaB proteins, respectively. Eighty micrograms of protein-containing sample was loaded in each lane.

Figure 6

Changes in the amounts of PsaA, PsaB, and MSP proteins in the btpA-deletion (ΔbtpA) strain during temperature shift from 30°C to 20°C. The deletion mutant was grown at 30°C and then transferred to 20°C. Aliquots of cells were collected before transfer (0), and 3, 6, 12, and 24 h after incubation at 20°C. Membrane proteins were fractionated on SDS-PAGE, transferred to nitrocellulose membranes, and probed with antibodies against PsaA, PsaB, and MSP proteins, respectively. Eighty micrograms of protein-containing sample was loaded in each lane.